Research Report 2010 2011 - Helmholtz-Zentrum für ...

RESEARCH REPORT 2006/2007
2010/2011

FOREWORD 04 The Helmholtz Centre for Infection Research – a portrait
07 Foreword
08 Orbituary – Jürgen Wehland
FOCUS 12 The German Centre for Infection Research (DZIF) – a great chance for the HZI
15 Highlights 2010-2011
RESEARCH REVIEWS 22 Frontier runners of Hepatitis C Virus
30 The aging of the immune system: challenges and perspectives
36 Novel nanoparticle-based technology platform for the delivery of vaccine antigens
42 Mycobacterial phagosomes and innate immunity
SPECIAL FEATURES 48 Where would we be without DNA sequences?
52 Nuclear magnetic resonance spectroscopy, a major player in the arsenal
of platform technologies available in the HZI
60 International cooperation of Helmholtz Centre for Infection Research with
India and China: a success story
SCIENTIFIC REPORTS 64 INFECTION AND IMMUNITY
67 Microbial Pathogenesis
69 Structural analysis of virulence factors
70 Virulence factors of streptococci and pneumococci
71 Molecular mechanisms of intracellular trafficking,survival and persistence of streptococci
72 Analysis of protein networks induced by early host-pathogen interactions
73 Structural and mechanistical analysis of functional amyloids
74 Microtubule dynamics and bacterial pathogenesis
75 Function and regulation of Yersinia virulence factors
76 Structural characterisation of pathogen defence factors
77 Mycobacterial phagosomes and immunity
78 Molecular mechanisms of host-cell pathogen interactions
79 Signalling to actin dynamics
80 Generation and exploitation of DNA sequence data
81 Host Pathogen Interactions
84 Pathogenesis of chronic Pseudomonas aeruginosa infections
85 Unravelling mechanisms of host defence against Gram-positive pathogens in the mouse model
86 Microbial communication
87 Systems genetics of infection and immunity
88 The structural basis of mammalian prion transmission barriers
89 Biofi lm communities
90 Metabolic diversity

SCIENTIFIC REPORTS 91 Infection and Immunity
92 Development and functional properties of Foxp3 + regulatory T cells
93 Mucosal immunity and infl ammation
94 Immuneffectors: molecules, cells and mechanisms
95 Interferons in viral defence and immunity
96 Cell-death mechanisms in immunity
97 Infl ammation and regeneration
98 Cellular models for infection
99 Strategies for Prevention and Therapy
102 Molecular mechanisms of infection and replication in the hepatitis C virus
103 Interaction between innate and adaptive immunity
104 Antigen delivery systems and vaccines
105 Chronic infection and cancer
106 Therapeutic cellular vaccines
107 Molecular diagnostics of mircobial pathogens
108 Pharmaceutical Research
110 Microbial diversity and natural product discovery
111 Medicinal chemistry of natural products
112 Chemical biology of infectious disease
113 Identifi cation of molecular targets of antiinfectives
114 New Project Groups
115 Regulation and herpesviral immune modulation of toll-like receptor signalling
116 Systems immunology
117 The National (“Helmholtz“) Cohort
118 Immune aging
119 Development of novel diagnostic and therapeutic tools for tuberculosis control and prevention
120 Exploring and exploiting microbial proteomes
121 POF II INDEPENDANT RESEARCH
122 Functional genomics and niche specifi ty
123 Structural biology of the cytoskeleton
124 Structural analysis of the innate immune system
125 TECHNOLOGICAL PLATFORMS
126 Central animal facility
127 Recombinant protein expression
128 Gene expression analysis
129 Peptide synthesis
130 Histo-pathology platform
131 Analytical instruments
132 The Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS)
138 TWINCORE, Centre for Experimental and Clinical Infection Research GmbH
148 Publications 2010-2011
FACTS AND FIGURES 172 Facts and Figures

4 PORTRAIT
The Helmholtz Centre for Infection Research
Our name reflects the scientific programme: At the
Helmholtz Centre for Infection Research in Braunschweig
approximately 350 scientists and 350 employees from
technology and administration lay the foundations for new
prevention methods, diagnostic procedures, medicines and
active agents with which infectious diseases can be better
treated or more effectively prevented. The paths that lead to
this goal are as varied as the paths by which the pathogens
enter our bodies. Microbiologists investigate how bacteria
and viruses manage to enter our bodies, how bacteria
communicate with one another and how exactly they make
us ill. Geneticists investigate our genetic make-up in search
of reasons why one person falls ill with flu, for example,
whilst his neighbour does not. Immunologists investigate
how organisms react to an intruder and resist it. Structural
biologists research the molecular structures of key molecules
with all of their interactions. Chemists use this
knowledge to investigate and develop new agents that can
in turn be employed to combat pathogens. Vaccine researchers
have their sights set on the best way to combat germs:
they aim to prevent these from making us ill at all.
New diagnostic approaches, vaccines or medicine can only
be successfully developed when the mechanisms of
infectious diseases have been properly understood. A
starting point from which researchers at the Helmholtz
Centre for Infection Research approach the complex
network of “infection” is the cell - both that of the host and
that of the bacteria. One example: opportunistic infections.
These are a problem in hospitals. In a place where patients
with weakened immune systems are treated, bacteria
transform themselves from unobtrusive companions to
dangerous aggressors. In extreme cases they ensconce
themselves permanently in our bodies by forming a
so-called biofilm, causing chronic illness under certain
circumstances. In biofilms the bacteria are surrounded by a
protective sheath that protects them very effectively against
attacks from the immune system or antibiotics. But what
has to occur in order for a seemingly harmless germ to
become an aggressor? How do bacteria communicate with
one another and the host? Only when scientists have
understood how the pathogens and their host cells interact
and which are the mechanisms behind these interactions
they can disrupt the communication between bacteria in a
targeted manner.
Infections are always caused by molecular interactions
between humans and pathogens and between the pathogens
themselves. Central element of this interplay are the
proteins involved in infections. These consequently play a
key role in the comprehension of infectious diseases. The
microorganisms organise and catalyse their life with
thousands of proteins. The goal of infection researchers is
not only to describe these proteins but also to understand
their functions. The proteins on the cell surface, in particular,
are responsible for the contact with the host – and the
question hanging over the research remains: how do bacteria
manage to infect us? The arsenal of these different pathogenity
factors, with which bacteria, viruses and other microbial
pathogens interfere with the process of human cells is a
modest one – however, knowing where these are located and
being able to name these factors does not yet mean that they
are understood. Proteins are very large molecules with
complex structures and it is precisely in these structures that
the secret of their success lies. The scientists at the HZI take
a close look at their structure - atom for atom. They look for
niches and hooks with which they brush alongside one
another, cling to and release one another.
Once the structural biologists have familiarised themselves
with these decisive parts of the molecular structure the
chemists can begin to put this knowledge to further use.
Their goal is to develop tailored molecules that fit this
structure exactly - and subsequently have the opportunity
to interrupt an infection. To this end chemists look for
natural inhibitors or create new synthetic ones, using these
to block the functions of the pathogen proteins in a targeted
approach.
The basis for new active agents, which can then attack the
weak points of the bacterial proteins, are often natural
agents. These originate in traditional, recently rediscovered
healing plants, from fungi or bacteria. A special role in the
HZI research is played by the myxobacteria. These live in
the soil and defend themselves against bacterial com petitors
with a range of chemical substances - substances that are
highly effective when precisely analysed by infection
researchers and then developed into medicines. Microbial
active agent principles for combating infectious diseases
are therefore seen as the optimal initial substances for
developing new therapies against bacteria for use in
hospitals.

PORTRAIT
5
The task of the chemists in infection research is easily
described but difficult to implement: chemists search for
effective components that enable myxobacteria to defend
themselves against other microorganisms or turn a plant
into a healing plant. And in these molecules they search for
decisive structural characteristics and chemical groups in
order to recreate these and even improve their effectiveness.
Synthesising or modifying natural agents to create
potent medicines is a discipline somewhere between pure
research and medicinal application for the HZI researchers.
An important step on the way for the application of natural
products is the further development of these compounds.
New and often multi-resistant pathogens demand an
efficient development of new antiinfectives. The Helmholtz
Institute for Pharmaceutical Research Saarland (HIPS), a
branch of the HZI and established in 2009, deals with that
task. Scientists at HIPS use genetic and genome-analytical
processes to optimize on one side the producers of natural
products and to further develop molecules, which are
potential candidates for an antibiotic treatment, for medical
and clinical purposes. With regard to this they are looking
for methods able to better transport such natural products
or their derivatives through biological barriers to the place
of their action and they are developing new and further
optimized formulations for those natural products.
The vaccine researchers at the HZI also tread a fine line
between basic and clinical research. Vaccines are held to be
the most effective and economic method for protecting
humans and animals against infectious pathogens.
Although the scientists are obviously in search of new
vaccines against illnesses such as AIDS or flu, one of their
specialities is the optimisation of vaccines. Adjuvants are
the key. These agents have no effect on infectious pathogens
themselves, but help the vaccines to develop their full
effectiveness.
imperviously to an infection. In this, the genes influence
one another, with the consequence that a defective gene
may be compensated by another gene or defects in genetic
material reinforce one another. The system-genetic investigation
of such interrelations will lead to new therapeutic
approaches. The instruments for this are not limited to the
petri dish, microscope or mouse, but are above all computers.
This research bridge, which the Helmholtz Centre for
Infection Research spans from the molecular interaction
between pathogen and host to new active agents and
prevention is aimed to lead directly to humans. Cooperation
with the Hannover Medical School has resulted in the
TWINCORE in Hannover – a translation centre where
clinical research and basic science meet. Medical personnel
and pure scientists are working there together under one
roof, drawing up joint solutions for the problem of infection
in doctors’ surgeries and clinics. The involvement in the
new research centres enables the Helmholtz Centre for
Infection Research to reinforce the health research
capabilities of the Braunschweig-Hannover district. And,
step by step, progress is being made towards resolving an
old - and yet current - problem: protecting people against
infectious diseases.
The Helmholtz Centre for Infection Research
Inhoffenstraße 7
38124 Braunschweig, Germany
Tel: +49 (0)531-61 81-0
Fax: +49 (0)531-61 81-2655
info@helmholtz-hzi.de
www.helmholtz-hzi.de
An infection requires the pathogens, the organism concerned
and environmental factors. As the environmental
factors have not proven readily comprehensible thus far, the
HZI researchers are focusing upon the interaction between
pathogen and host. The research field that aims to address
the associated, highly-complex questions is in the first
stages of development: system genetics. A whole series of
genes determine whether a host responds sensitively or

6 Guests at HZI events
Participants of the opening ceremony of the symposium “Networking and Technology Transfer”, organized by HZI, InWEnt,
and Thai Red Cross Photo: QSMI/HZI
Photos: HZI

FOREWORD | Prof. Dr. Dirk Heinz
7
Foreword
Prof. Dr. Dirk Heinz | Acting Scientific Director
The past two years have comprised a phase of change and transition for the Helmholtz Centre for Infection Research (HZI). After
its successful scientifi c refocusing on infection research, the Scientifi c Director Rudi Balling left in 2009 - fi rst for a scientifi c sabbatical
to the Broad Institute and shortly after permanently for a new position as Director of the Luxembourg Centre for Systems
Biomedicine. With Jürgen Wehland, former head of the Cell Biology Department, an excellent successor for this crucial position
was appointed in January 2010. Keenly aware of the research performed at the HZI, he was a natural choice for steering the centre
and for shaping its profi le and scientifi c strategy. After less than one year in offi ce, his tragic death on 16th August 2010 saw the
loss of a highly respected colleague and dear friend to many of us. In spite of this overshadowing event, and with the third Scientifi
c Director in offi ce in less than three years, the HZI has continued to emerge as a centre of excellent infection research. In the
spirit of the changes initiated by Rudi Balling, continued and realigned by Jürgen Wehland, a number of important projects have
been launched and we are now successfully implementing these further, both on the HZI campus and beyond:
• In 2009, the Helmholtz Institute for Pharmaceutical Research Saarbrücken (HIPS) was founded in collaboration with the University
of Saarbrücken. The pharmaceutical know-how at the HIPS ideally complements the expertise in infectious disease and
natural compound research at the HZI.
• Together with the Fraunhofer Institute for Toxicology and Experimental Medicine (ITEM), the Hannover Medical School (MHH),
the Leibniz-University Hannover, the University of Veterinary Medicine Hannover and the University of Braunschweig, the HZI
founded the “Translationsallianz in Niedersachsen” (TRAIN) in 2008 to promote translational drug and vaccine research. In May
2011, construction of the Hannover Centre for Translational Medicine (HCTM) will start. The HCTM is a joint project of the TRAIN
partners ITEM, MHH and HZI and will serve as a platform for early clinical studies.
• The HZI, together with its partners in the Braunschweig-Hannover region, successfully took part in the competitive selection of
founding members of the German Centre for Infection Research (DZIF), following an initiative of the Bundesministerium für Bildung
und Forschung. Together with 26 other research institutions from all over Germany, the HZI is now actively participating in
the foundation process. The DZIF will join forces in infection research and places emphasis on the translation of basic research
into medical application. The central DZIF administrative offi ce will be located at the HZI.
• Working together with several partner institutions in northern Germany, the HZI took the lead in the development of a concept
for the Centre for Structural Systems Biology (CSSB). The CSSB will bring together expertise in structural and systems biology to
allow the analysis of the molecular processes taking place during infection and other diseases on a hitherto unattainable level of
tempo-spatial resolution. The founding agreement for the CSSB was signed early in 2011.
• A number of new buildings on the HZI campus have been completed or are about to be fi nished:
· The new animal facility (Tierhaus II) commenced operation in 2010.
· The administrative annex building was set-up at the end of 2009.
· The novel BSL3-building is now complete and will be operational in 2011.
· The office-annex for building D was topped out in spring 2011 and will allow establishment of new lab space upon completion.
As a consequence of these successful projects and collaborations, the HZI is now on course for the next round of the Helmholtz
Programme-oriented Funding (PoF III) in 2013.The HZI is well prepared for the challenges posed in infection research through
its clear strategic focus on the elucidation of infection processes complemented by its anti-infectives research and its on-going
efforts in the translation of basic research in collaboration with its academic and industrial partners. In future we will further
strengthen our scientifi c profi le by fostering outstanding projects and recruiting additional scientifi c expertise.
Prof. Dr. Dirk Heinz

8 OBITUARY | Jürgen Wehland
… during a staff outing
… a visit of politicians
… during a PhD commemoration
… during the visit of Minister Prof. Schavan
… during a PhD commemoration
… during the TRAIN event
… during the TRAIN event
All Photos: HZI

OBITUARY | Jürgen Wehland
9
Jürgen Wehland †
Scientific Director 2009-2010 | Head of the Division of Cell and Immune Biology 1997-2009 |
Scientist at the HZI since 1989
The Helmholtz Centre for Infection Research mourns Jürgen Wehland who suddenly and unexpectedly passed away on
August 16, 2010, while on vacation in Scandinavia.
Jürgen Wehland was a key fi gure in the HZI for many years and in numerous aspects:
As an excellent scientist with international standing, he made seminal contributions to the fi elds of cell and infection biology.
Using Listeria monocytogenes as a model organism, he could show how the human pathogen utilizes the host’s actin
cytoskeleton to promote infection. Together with other leading scientists in the fi eld, he laid the foundation for a new discipline,
cellular microbiology, tackling host-pathogen interactions at various molecular levels, without which modern infection biology
would simply be unthinkable.
As an outstanding scientifi c manager, he not only developed his own department into one of the most successful ones of the
HZI but also represented the mission and vision of the entire centre by skilfully shaping the research programme, by fostering
strategic alliances with our partner institutions and by promoting a number of crucial recruitments to Braunschweig and our
subsidiary institutions. Even though only a short time in offi ce as the Scientifi c Director of the HZI, he was leading the centre
towards exciting and important future developments.
As a caring supervisor and an inspirational mentor he actively supported the scientifi c careers of numerous young scientists
within the HZI and beyond.
As a dear colleague and friend to so many of us his door was always open for scientifi c and personal advice. He just always
took the time.
Beside his many great accomplishments, Jürgen Wehland remained a remarkably modest character, a great example to all of us.
We sadly miss him!

FOCUS RESEARCH REVIEWS SPECIAL FEATURES
Photos from left to right: The representatives of the HZI PhD-student organization announcing the award for the best PhD-supervisor
of the year 2010: Prof. Dr. Eva Medina. | Thomas Gazlig, the head of the HGF department of Public Relations, during a talk at the
exhibition “Wunderkammer der Wissenschaft” at the Braunschweig “Schloss-Akarden” | Prof. Dr. Kurt Wütherich, Nobel Laureate,
with Maxi Scheiter discussing her results of the poster presentation Photos: HZI, Hübner (le) | HZI (ce) | HZI, Hübner (ri)

SCIENTIFIC REPORTS
FACTS AND FIGURES
12 The German Centre for Infection Research (DZIF) –
a Great Chance for the HZI – an Interview
15 Highlights 2010–2011

12 FOCUS | The German Centre for Infection Research (DZIF) – a Great Chance for the HZI – an Interview
The German Centre for Infection Research (DZIF) – a Great
Chance for the HZI
INTERVIEW WITH | Prof. Dr. Dirk Heinz | Acting Scientific Director of the Helmholtz Centre for Infection
Research – HZI
Professor Heinz, in November 2010 the HZI was selected
as a partner institution for the German Centre for Infection
Research. What does this decision mean for the
centre in Braunschweig?
Dirk Heinz: We are proud to be included in the small circle
of leading research facilities that have been assigned to
jointly established the German Centre for Infection Research
(Deutsches Zentrum für Infektionsforschung, DZIF).
The DZIF is part of a nation-wide initiative of the Federal
Ministry of Education and Research. Several German centres
for health research will be launched in order to take a
more targeted course of action against the major diseases.
The aim is to consolidate research efforts, to improve exchange
between participating institutions and to accelerate
the transfer of innovation from basic research into the
clinic – the so-called translation.
Which diseases are the focus of attention for the Federal
Government?
DH: In the context of this initiative, four German centres for
health research are to be founded: the German consortium
for translational cancer research as well as German centres
for research on cardiovascular diseases, pulmonary diseases
and, of course, infection research. The prerequisites
for selection of the sites were proven scientific expertise,
and the ability to expedite translation research. Both infrastructures
as well as personnel will be financed for this
purpose on a sustained basis.
How will the national health research centres be
organized?
DH: In most of the centres, and the DZIF as well, it is a
matter of inter-site constructs. No new major research facilities
are being built, but leading institutions are brought
together in order to facilitate networking and coordination
of work under one common organisational roof. This is also
the case for the DZIF with seven locations in Hannover-
Braunschweig, Hamburg-Lübeck-Borstel, Bonn-Cologne,
Tübingen, Munich, Heidelberg and Gießen-Marburg-Langen.
The central administrative office, which coordinates the
work of the various research projects within the DZIF, will
be located in Braunschweig on the HZI campus.
The participating institutes of the DZIF are located in the cities,
marked in the map. Further details see Interview.
Which topics and which pathogens are to be investigated at
the DZIF?
DH: It is a primary goal to tackle important health issues
relevant to Germany, which clearly not excludes major infectious
diseases relevant to the rest of the world. Therefore,
chronic viral diseases such as HIV/AIDS and hepatitis are,
among others, a major topic. Another focus are gastrointestinal
infections by Helicobacter, Yersinia and other
pathogenic bacteria. These are areas to which our consortium
will also make considerable contributions. Furthermore,
tuberculosis and malaria are highly relevant topics in
infection research to be addressed by the DZIF. One should
not forget hospital-acquired infections with persistently
reoccurring problems of antibiotic resistance and biofilm
formation. MRSA and pseudomonads, in particular, play
a role here. Moreover, the researchers at the DZIF will be
dealing with overriding issues of infection research. These
will include control of pathogens by the immune system,
improved vaccinations and, most importantly, new active
compounds against infections.

FOCUS | The German Centre for Infection Research (DZIF) – a Great Chance for the HZI – an Interview
13
What is the advantage of such a large alliance like the DZIF
over the numerous cooperations that we already have –
nationally as well as internationally?
DH: It is a different approach. In the future we will not carry
out research and cooperate as before. Rather, we will initiate
new approaches. Together the strong partner institutions
can, through consolidation in the DZIF, make use of common
infrastructures and expertise. This will allow a more consistent
translation from fundamental research to clinical
studies. The ultimate target is the development of new therapeutic
agents or preventative measures such as new vaccines
and vaccination strategies. We have to work together and
synergistically make our respective expertise available, so
that optimal results can be achieved for public health.
What shall be the task – the special function – of the HZI
within the DZIF alliance?
DH: On the basis of our special expertise, research on
natural bioactive compounds is the central theme for the HZI
researchers in the DZIF. In this regard TWINCORE and the
Helmholtz Institute for Pharmaceutical Research Saarland
(HIPS), in addition to the activities on the HZI campus, play
an important role. Together we are constantly expanding
the pool of available natural substances from myxobacteria.
They have proven to be a highly potent source for a diversity
of bioactive compounds. Currently 80 percent of all antibiotics
found on the market originate from natural substances.
This shows that, in the search for new anti-infective agents,
we have to revert back to the existing, albeit insufficiently
developed, pool of natural substances. Furthermore, novel
compounds can be produced by means of targeted genetic
modification of production strains, or previously known
structures can be produced in larger quantities. The existing
bank of natural substances at the HZI will now be continuously
expanded. We will make it accessible to the various
consortia for research on completely different pathogens. Our
chemists enable the transition from the screening hit to lead
structure candidates. Moreover, we are handling topics such
as drug delivery procedures. It is in this context that our
pharmaceutical research at HIPS comes into play. Without it,
we could not make use of our bio-active compounds as pharmaceutical
products that can then be used in clinical studies.
Academic basic research as a basis for the development of
new drugs: this aspect of our DZIF application represents a
unique characteristic. It shows great promise for the future.
This is, in addition to our experience in translation research,
a main criterion for our funding.
What about organisation? You mentioned that the administrative
office will be located in Braunschweig…
DH: For all of the German centres for health research
there is a Helmholtz Centre, through which the funds from
the Federal Ministry of Education and Research flow into
the respective consortia. This means that we will take
the responsibility for both distribution of funds and the
controlling. That is why the subsidy management has to
be residing within the HZI. The DZIF administrative office
will work closely with the department for subsidy management
- another important reason for its location at the HZI.
However, the office itself will be independent of the HZI and
our own projects.
What level of funding are we talking about?
…Together the strong partner institutions can, through consolidation
in the DZIF, make use of common infrastructure and
expertise…. Photo: HZI; Gramann
DH: In the final developmental stage from 2015 on, almost
40 M€ per year have been planned for the complete DZIF.
The core estimated funding begins at developmental levels
this autumn. The portion of our own consortium will
amount to several million euros per year in the final stage.

14 FOCUS | The German Centre for Infection Research (DZIF) – a Great Chance for the HZI – an Interview
The scientists whose studies have gone into the application
process for the DZIF are already employed. They have been
working on their scientific objectives for many years now.
And today the objectives of the DZIF are added to this. Are
resources being restocked? What will the finances from the
DZIF be used for?
DH: First of all, money is being used to build up infrastructure
and expertise. This will allow us to do justice to
the claim that we work and live scientifically according to
translation principles. For example we shall, by means of
specific appointments, strengthen the research in bioactive
compounds on the HZI campus. This means that colleagues
will be specifically financed through the DZIF. An appointment
in bio-informatics would be equally conceivable. We
are investing new financial means primarily into this. Of
course, this does not forbid colleagues to incorporate their
expertise into the major thematic topics. A few examples
are: Eva Medina in staphylococci, Thomas Pietschmann in
hepatitis and Susanne Häußler in resistant gram-negative
bacteria. And, by the way, for this purpose we are bringing
a considerable portion of capital resources into the DZIF.
Let us return to the topic of translation, which represents a
primary concern of the DZIF. The HZI has already achieved
much in this regard. We already have the Translation Alliance
in Lower Saxony, TRAIN, and there is the TWINCORE
Centre for Experimental and Clinical Infection Research.
There are partners here who shall become a part of the
DZIF and who are already connected with one another in
their translation activities. What will interaction with the
DZIF look like?
DH: Without TRAIN, we would conceivably not have TWIN-
CORE either. It is possible that the expert evaluators would
have perceived the basic research here in Braunschweig and
the clinical research in Hannover as isolated site activities
that only collaborate on small projects. Giving money to
such facilities would probably not raise expectations that
they would be doing research together in a superordinate
structure in five years. TRAIN and TWINCORE thus were
the pre-conditions for the fact that we are forming, jointly
with the MHH, such a momentous double site within the
DZIF. I view the DZIF as a consistent continuation of our
translation activities. We are currently expanding beyond
local activities, with topics that are specified within the
DZIF, and make use of the strong local structures for this
purpose. One should realize that TRAIN and TWINCORE
are dealing with other projects as well, as opposed to those
which are planned in the DZIF.
… I view the DZIF as a consistent continuation of our translation
activities…. Photo: HZI; Gramann
What does the work at the DZIF specifically mean for the
HZI? Will the employees here on campus notice that the
DZIF exists?
DH: Some employees will strongly notice, but certainly not
all. We did not involve all departments directly in our application
to the DZIF. We placed our emphasis on host-pathogen
interactions and functional genome research, but most of all
on active compounds. Colleagues who are involved in these
areas will actively participate in the corresponding topics.
They will become part of a translation pipeline.
And when will it all begin?
DH: Presumably in the summer of 2011, but in the autumn
at the latest. After appraisal of the joint application in April,
a decision will be made. We will then know whether all
specifically-announced research projects within the DZIF
will be funded in their entirety*. When all these decisions
have been made, we can start. We are already looking forward
to working with all of our partners.
The interview was performed through the Public Relations
Department of the HZI in March 2011.

FOCUS | Highlights 2010-2011
15
Highlights 2010-2011
AUTHORS | Manfred Braun | Head of the Department of Public Relations | mbn@helmholtz-hzi.de |
Dr. Bastian Dornbach | Department of Public Relations | bad@helmholtz-hzi.de
Helmholtz Touring Exhibition in Braunschweig The
exhibition “Wunderkammer Wissenschaft” (“Chamber
of Scientific Wonders”) was a special attraction in the
Braunschweig “Schloss-Arkaden” from 8 to 16 April 2010.
More than 335,000 visitors attended the touring exhibition
and gathered information regarding research at the
Helmholtz-Association. About 500 images and multi-media
offerings in huge opened trunks provided insights into
miniscule nano-worlds and gigantic large-scale devices at
the Helmholtz Centres. The spectrum ran the gamut from a
close-up image of mould-fungus to photos of distant planets
and appealed to a broad audience from pupils to adults. The
photos – in part animated, but always animating in their
effect – led to intensive discussions with the exhibit attendants
from the HZI.
Herbert Waldmann 2010 awardee of the Inhoffen Medal
Winner of the Inhoffen award for 2010 was Prof. Dr. Herbert
Waldmann from the Max-Planck-Institute for Molecular
Physiology in Dortmund. Herbert Waldmann received the
award donated by the “Friends of the Helmholtz Centre”
(Förderverein des HZI) for his syntheses of new active
compounds. The biochemist Herbert Waldmann sought
after models, in nature, for new potential medications.
These are biologically-active molecules, that are produced
by plants, microorganisms and animals as, for example,
defence mechanisms or as messenger substances between
cells and organs. Many natural substances affect not only
their originally-intended targets but also certain human
metabolic processes. Almost half of the medications now
available are based on natural substances or natural
substance-like structures. Herbert Waldmann and his
co-workers were successful through their efforts of many
years in synthesizing a variety of new substances. A few
of them have a fundamental effect on the growth of breast,
ovary and stomach tumours, while others are able to regulate
the blood-sugar levels or participate in the conduction
of nerve stimuli.
A view of the exhibition “Wunderkammer Wissenschaft” at the
Schloss-Arkaden Braunschweig. Photo: HZI, Ritter
Science Shopping – Science late into the night Under
the auspices of the “Moonlight Shopping” in the Braunschweig
city-centre, the HZI presented, as one of 13 regional
research facilities, their “Science late into the night” exhibit
(“Wissenschaft bis in die Nacht”). Visitors were able to
gather information at four different learning stations
regarding infectious diseases and carry out experiments
with scientists in the afternoon until 11 p.m. The topics:
How does vaccination actually work and how does our
immune system function? How come the influenza virus
does not know borders? What are prions? Information
boards displayed the influence of infectious diseases upon
contemporary art. Young researchers were able to find out
for themselves why it is that red cabbage is also called blue
cabbage. The event was organised by the local research institutions
in cooperation with Braunschweig City Marketing
and business proprietors in the Braunschweig city-centre.
The HZI participating in the event “Moonlight Shopping” at
Braunschweig city-centre within the part “Science late into the
night”. Photo: HZI, Morczinietz

16 FOCUS | Highlights 2010-2011
Prof. Dr. Herbert Waldmann (middle) with the Inhoffen Medal
2010 together with Prof. Dr. Jürgen Wehland (†, at that time
Scientific Director of HZI, left) and Prof. Dr. Joachim Klein
(President of the HZI Förderverein, right) Photo: HZI, Gramann
Summer holiday course “Everything about Sugar”
Learning about biochemical principles through experiments
and result reportage was the objective of a four-day
summer holiday course for pupils, which the School Lab
“BioS” and the Public Relations Department of the HZI
organized jointly. The topic: sugar in all its manifold forms.
Pupils became acquainted with biochemical verification
methods for common sugar and got a first impression of
how laboratory studies are carried out. In the second part
of the course, pupils learned various journalistic formats
and processed their knowledge of topics such as diabetes
and lactose intolerance. This combination of biochemical
studies and reflection regarding scientifically-applied topics
was quite successful. Not only did it introduce pupils
the concept of having fun while working but they also
learned journalistic approaches to the technique of asking
suitable questions to find relevant information.
HZI at NDR Info: Big fear of small viruses – Tracking
down pathogens NDR Info, the Northern German news
radio station, introduced in March 2010 in collaboration with
the “Braunschweig Newspaper” and the House of Science
(“Haus der Wissenschaft”) in Braunschweig, the new discussion
and broadcast series “Logo – Science from Braunschweig”.
Braunschweig has, according to an EU study, the most
active research region in Europe with the highest concentration
of scientists. More than 15,000 individuals work in 26
scientific organisations and research facilities and there are
about 250 light industries within the high-tech sector. For
the second broadcast of this series in June, the Logo moderator
Regina Methler discussed with experts from the HZI and
with the audience topics concerned with bacteria and viruses
as well as our fears and the opportunities and limitations
inherent in infection research. These included: “When will
we experience the next pandemic?” and “Are animal experiments
still necessary in infection research?”
CSSB agreement Infection researchers and physicists in
Northern Germany are blazing new trails together in their
research of infectious diseases; the new “Centre for Structural
Systems Biology” (CSSB) is being developed on the
campus of the German Electron-Synchrotron (DESY) in Hamburg-Bahrenfeld
with scientific coordination from the HZI.
The CSSB is building a bridge between structural biology
and systems biology. In this context, biologists, chemists,
health and medical professionals, physicists and engineers
are studying the interaction between pathogens and their
hosts. For this purpose, unique tools are being made available
to them Germany-wide through DESY: the storage ringbased
X-ray source PETRA III, the free-electron laser FLASH
and the X-ray laser European XFEL, which is presently being
built. A federal government and state agreement will make
50 million euros available for construction of the CSSB.
A group of pupils that participated in the Summer holiday
course at the BioS-Laboratory. On the right side Dr. Iris
Eisenbeiser from the BioS-Laboratory. Photo: HZI, Fischer
After signing of the CSSB agreement at the DESY facilities in
Hamburg. From left to right: Prof. Dr. Helmut Dosch, President
of the DESY Board of Directors; Prof. Dr. Johanna Wanka,
Minister of Science and Culture of Niedersachsen; Christoph
Ahlhaus, Lordmayor of the Freie und Hansestadt Hamburg;
Prof. Dr. Annette Schavan, Minister of Education and Research,
BMBF; Dr. Herlind Gundelach, Senator of Science and
Research of Hamburg; Prof. Dr. Dirk Heinz, acting Scientific
Director of the HZI. Photo: DESY, Reipka

FOCUS | Highlights 2010-2011
17
Peter Seeberger receives the Inhoffen Award for 2011
For his fully automatic carbohydrate-syntheses, Prof. Dr.
Peter Seeberger received the 2011 Inhoffen Award. The
department head and managing director at the Max-Planck
Institute for Colloids and Interfaces in Golm near Potsdam
was successful in his attempts to produce completely
synthetic complex carbohydrates, which play a significant
role in almost all physiological processes. Carbohydrates,
or sugar molecules, fulfil multifarious tasks in and on our
cells. They deliver energy, form the foundation for surface
structures such as phlegm, but are also used by pathogens.
They play, therefore, a decisive role in infections and
immuno-reactions and are attractive target molecules for
medical research. However, a chemical synthesis method
had been missing hitherto for producing biologically important
sugars in large amounts – a gap that Peter Seeberger
was able to fill.
During the press conference of the State of Niedersachsen with
the participating institutions of the German Centres for Health
Care Research in Niedersachsen. 2 nd from the left side: Prof. Dr.
Dirk Heinz, acting Scientific Director of HZI. Photo: MHH, Kaiser
State Secretary Dr. Georg Schütte as a guest at the HZI
On Monday, January 17, 2010, Dr. Georg Schütte State
Secretary from the Federal Ministry for Education and Research
(BMBF) was a guest at the HZI to become informed
of the research at the HZI. In addition to information
regarding the world of viruses and microbes, he had the opportunity,
through extensive discussions with the management
board, to find out about the challenges and research
goals of the HZI. He then visited the often awarded School
Lab “BioS”, where he was able to gain insights regarding
the facility for promotion of a rising generation of young
scientists in biotechnological research.
Prof. Dr. Peter Seeberger (middle) with the Inhoffen Medal
2011 together with Prof. Dr. Joachim Klein (President of
the HZI Förderverein, left) and Prof. Dr. Dirk Heinz (acting
Scientific Director, HZI, right) Photo: HZI, Hübner
HZI participating in the German Centre for Infection Research
To investigate pathogens and to develop new strategies
against them – these are the core tasks of the “German
Centre for Infection Research” (DZIF), whose research
programme will start in the summer of 2011. The initiative
of the Federal Ministry for Education and Research (BMBF)
comprises numerous partners from university and nonuniversity
research groups. The HZI is participating in
DZIF as part of the Hannover-Braunschweig consortium.
The administrative centre of the DZIF will also be located
at the HZI. Partner facilities in the region Hannover-Braunschweig
are, in addition to the HZI, the Medical School of
Hannover, the University of Vetenary Medicine Hannover
(TiHo), the Technical University of Braunschweig (TU BS),
the German Collection of Microorganisms and Cell-Cultures
(DSMZ) and TWINCORE, the Centre for Experimental
and Clinical Infection Research.
State Secretary Dr. Georg Schütte (BMBF) during his visit
at the HZI and BioS. Here talking with a pupil at the BioS
Laboratory. Prof. Dr. Dirk Heinz (acting Scientific Director
of HZI, right) and Ulf Richter (Administrative Director,
background) Photo: HZI, Hübner

18 FOCUS | Highlights 2010-2011
First “North-Regio-Day on Infection“ at the HZI On 1 st
and 2 nd October 2010, scientists from universities and
research facilities in Northern Germany met at the HZI on
the occasion of the first “North-Regio-Day on Infection“;
the researchers presented up-to-date research results,
became familiar with new technical developments and
discussed important issues and future challenges in infection
research. The focus was on bacterial pathogens. The
meeting was coordinated jointly by the HZI, the Technical
University of Braunschweig and the Robert-Koch-Institute
in Wernigerode (RKI) under the auspices of the HZI Graduate
School. The goal of the “NORDI” symposia in the future
will be the promotion of exchanges between researchers in
northern Germany and promotion of junior scientists.
Prof. Les Baillie from the Welsh School of Pharmacy of the
Cardiff University, UK, during his presentation. Photo: HZI, Oumard
The organizers of the symposium. From left to right: Antje
Flieger (Robert-Koch Institute, Berlin), Petra Dersch (HZI) and
Michael Steinert (TU Braunschweig). Photo: HZI, Dornbach
“Day on deadly killers“ On the “Day on Deadly Killers“
on March 3, 2011, international experts discussed the current
status of infection research. The pathogens for such
dreaded epidemics as cholera, anthrax, rabies and AIDS
represented the focus of the symposium at which experts
from Europe, the USA and Asia presented their research
results and discussed new therapeutic approaches against
infectious diseases.
Gene laboratory and school The workshop “Genlabor &
Schule” was a guest at the HZI from 24 to 25 September
2010. The Society for Biochemistry and Molecular Biology
(GBM) and the Braunschweig School Lab “BioS” invited
participants to the fifth workshop of the Network “Genlabor
& Schule”, a unique event in the German-speaking countries.
These people responsible for almost 40 school labs in
Germany, Austria and Switzerland have met annually or
bi-annually since 2002. The workshop “Genlabor & Schule”
offered an extensive programme consisting of technical
lectures, poster presentations and panel discussions. It is
a forum for everybody who deals with the subject of junior
scientists in the natural sciences. Scientists, directors and
employees of the school lab, experts from the field of biology,
as well as representatives from the Cultural Ministries
of Hessen and Niedersachsen exchanged their experiences
with the goal of developing the school lab as an extracurricular
place of learning.
Among the speakers was the renowned US scientist Dr.
Henry F. Chambers from General Hospital in San Francisco,
specialist for antibiotic-resistant hospital infections.
Dr. Iris Eisenbeiser (left) giving some explanations during the
workshop “Gene lab & School” Photo: HZI

FOCUS | Highlights 2010-2011
19
Nobel Laureate Prof. Kurt Wüthrich guest at the HZI
An international star of science came to visit Braunschweig
on April 4, 2011. Prof. Dr. Kurt Wüthrich visited the HZI to
discuss research with colleagues and to exchange findings.
The Swiss-born scientist is currently supervising a laboratory
at the Scripps Institute in La Jolla, USA, as well as a
second one at the Federal Technical University (Eidgenössische
Technische Hochschule) in Zürich. His research
focus has a direct connection with research at the HZI
– clarification and examination of the spatial structure of
biological molecules. Kurt Wüthrich developed, collectively
with other colleagues, the technology of the so-called “multi-dimension
NMR spectroscopy” as a powerful analytical
tool for the structural elucidation of large biomolecules. For
his pioneering work Kurt Wüthrich was awarded the Nobel
Prize for Chemistry in 2002.
Visit of Nobel Laureate Prof. Wütherich at HZI. From left to right:
Prof. Dr. Christiane Ritter, Dr. Torsten Lührs (both HZI), Prof.
Dr. Kurt Wüthrich (Scripps Institute La Jolla, USA), Prof. Dr.
Dirk Heinz (acting Scientific Director HZI). Photo: HZI, Hübner
Prof. Dr. Rolf Müller, HZI Photo: HZI, Gramann
Cells meet Surface In May 2010, scientists took a deeper
look at cell surfaces: “Cells meet Surface”, a two-day symposium,
organized by the German Association of Transfusion
Medicine (“Berufsverband Deutscher Transfusionsmediziner”,
BDT), Fhg IST, SFB 578, the InnoNet project
”InnoSurf“, the Helmholtz Centre for Infection Research
(HZI) and the DECHEMA, took place at the HZI Forum.
For a long time the interaction between cells and (artificial)
surfaces was not taken into account. However, scientists
learned that this interaction is essential for the well-being
of a cell and that the cultivation of cells can be influenced
positively or negatively by physical or chemical modifications
of surfaces. The scope of this meeting was to present to clinicians
and clinical-oriented scientists the latest developments
in this field. Main emphases of this symposium were put on
the modification of surfaces and the technical possibilities
to prove these and to influence the characteristics of cells.
Furthermore, new ways of cell analysis based on magnetic
signals, the successful use for the generation of cell products
and initial attempts in clinical practice were presented.
Rolf Müller awarded the DECHEMA Prize of the Max-
Buchner Research Foundation Prof. Rolf Müller received
the 20,000 euro DECHEMA Prize of the Max-Buchner-Research
Foundation on 26 November 2010 for his research in
biologically active natural materials that are produced from
bacteria living in the soil and used to develop new medications.
Rolf Müller is Managing Director of the Saarbrücken
branch of the HZI, known as HIPS, and teaches at the
University of Saarland. The department led by Rolf Müller
concentrates on analysis and production of molecules derived
from myxobacteria and actinomycetes with biological
activity. An example for a microbial active ingredient made
from myxobacteria is epothilone, which has been successfully
deployed in cancer therapy in the USA.

FOCUS RESEARCH REVIEWS SPECIAL FEATURES
Photos from left to right: Niña S. Cortina, HIPS, feeding a Genetix clone picking robot. | Dr. Maximiliano Gutierrez
acquiring confocal images of eukaryotic cells infected with mycobacteria | Wiebke Ginter, Twincore, during the preparation
of a new experiment Photos: HIPS/HZI (le) | HZI, Bierstedt (ce) | Twincore/HZI (ri)

SCIENTIFIC REPORTS
FACTS AND FIGURES
22 Frontier Runners of Hepatitis C Virus
30 The Aging of the Immune System: Challenges
and Perspectives
36 Novel Nanoparticle-based Technology Platform
for the Delivery of Vaccine Antigens
42 Mycobacterial Phagosomes and Innate Immunity

22 RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
Frontier Runners of Hepatitis C Virus
CORRESPONDING AUTHOR | Prof. Dr. Thomas Pietschmann | Department of Experimental Virology |
TWINCORE | thomas.pietschmann@twincore.de | tpi07@helmholtz-hzi.de
CO-AUTHORS | Dr. Sandra Ciesek | Dr. Eike Steinmann | both at Department of Experimental Virology | TWINCORE
Transmission of hepatitis C occurs via direct blood-to-blood contact: prior to 1990 primarily blood transfusions were the problem,
however, today transmission by means of blood products, at least in Germany, no longer plays a role due to highly sensitive and
accurate screening procedures. Risk factors to date are non-sterile handling of injection utensils in non-industrialised nations
and intravenous drug consumption in industrialised nations. In the case of tattooing, piercing, acupuncture and medical interventions,
application of insuffi ciently sterilised instruments can lead to transmission of the virus. It is actually quite seldom that
hepatitis C virus (HCV) infections are attributed to sexual intercourse with hepatitis-C-positive sexual partners. Transmission of
the infection from an HCV-positive mother to her child prior to or during birth occurs in no more than 4% of cases. How ever, for
approximately one-third of HCV-patients, the origin of the transmission is no longer traceable.
With currently approximately 130 million virus carriers,
hepatitis C virus (HCV) infection is one of the most widespread
infectious diseases worldwide. According to data
from the Robert-Koch Institute, there are ca. one-half
million virus carriers living in Germany. In the USA and
Europe it is estimated that 1.5% of the population are
infected, while in Egypt and Central Africa the ratio of up
to 20% is significantly higher. The virus is highly mutable
and is, therefore, able to evade, time and time again, the
immune system. On the basis of sequence analyses, the
viruses are separated into seven genotypes, which deviate
from one another by 30% and respond to medications with
varying success.
The natural course of a HCV infection can be roughly
devided into two stages: an acute and a chronic phase.
During the first six months after being infected, the patient
will experience the acute, mostly symptom-free phase. A
segment of the patients is able to control virus replication
during this acute phase and the infection is spontaneously
cleared. But for the majority of patients (50% to 90%), the
virus is not eliminated and a chronic infection ensues.
Occasionally, non-specific symptoms will emerge such as
fatigue, depression, nausea, pains in the upper abdomen
and indigestion – often times the infection is noticed and
confirmed many years after the initial contagion. Of these
chronically-infected patients, up to 40% develop a progressive
liver disease with formation of cirrhosis of the liver
(Figure 1). HCV cirrhosis is, furthermore, one of the main
risk factors for emergence of a heptocellular carcinoma
(HCC). The effects of a chronic hepatitis C virus infection
are considered among the most frequent indications for
liver transplantation 3 . In light of the world-wide dispersal
of the HCV infection, considerable clinical significance is
attributed to this pathogen on the grounds of its frequency
and morbidity.
Fig. 1. Comparison of a cirrhotic liver (above) with a healthy
liver (beneath). Photo: Twincore

RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
23
Currently, world-wide there are only insufficient options
available for treatment of hepatitis C. Since the beginning
of the 90’s chronic HCV infection has been treated with
interferon-alpha, a cytokine of our immune system, which
triggers mechanisms in the cell to control a virus infection.
In recent years this therapy has been substantially
optimised. The application of interferon alpha (IFN-alpha)
derivatives with an increased half-life time (pegylated
interferon) and in combination with ribavirin, a nucleoside
analogue, the response rates have improved considerably.
With this combination it is possible to achieve a long-term
virological response with 50% of the patients with the HCV
genotype 1, while patients with genotype 2 and 3 viruses
are even able to attain long-term therapeutical success in
more than 80% of the cases 12 . New and directly effective
antiviral medications will increase the options for therapy
by the end of 2011. However, these novel therapeutics are
unfortunately not suitable for the treatment of all viral
genotypes. Since the virus is able to become resistant very
quickly, following application of these combinations without
further medications (mono-therapy), these preparations
supplement standard therapy, but are not able to replace the
IFN-ribavirin treatment with its multiple side-effects. This
leaves us with continued demand for the development of new
forms of therapy with preferably diminished side-effects,
genotype-overlapping effectiveness and a high barrier
against viral resistance.
In this tug-of-war between fundamental questions generated
by the virus and the clinical challenges of therapy, we
at TWINCORE are focusing on HCV. Our team is composed
of natural-science scientists and physicians. We work
closely with the Department of Chemical Biology of the HZI
(Helmholtz Centre for Infection Research - Dr. Ronald Frank
und Dr. Florenz Sasse), and with the Clinic for Gastroenterology,
Hepatology and Endocrinology (Prof. Dr. Michael
Manns) of the MHH (Hannover Medical School). This close
collaboration between TWINCORE scientists, HCV experts
in the clinic and natural scientists at the HZI fosters the
successful operation of a translational research programme.
Convenient proximity to the MHH is quite helpful since
it facilitates pragmatic implementation of the vision of
translational research – for example, when young assistant
physicians oversee research projects within their respective
medical environment at TWINCORE.
The basis for our collaborative efforts is a cell-culture
system developed by us for HCV. With this groundwork,
we can examine the mechanisms of virus-replication in
human liver cells in the Petri dish, identify new targets for
therapy, and search for new active compounds that are able
to inhibit propagation of the virus. We also use this model
in order to be able to, on the one hand, better estimate
HCV transmission risks within the clinical environment
or amongst the drug milieu, for example, and, on the other
hand, to define disinfection agents and hygienic procedures
that can safely inactivate the virus.
The development of HCV cell-culture models As opposed
to bacteria or fungi, viruses are obligatory intra-cellular
parasites. They have no self sufficient metabolism of their
own and are therefore dependent upon suitable host cells.
HCV-replication can therefore only be examined in cell
cultures – however, for many years scientists had been
unsuccessful in their attempts to develop such systems for
the propagation of HCV.
It was not until ten years after the discovery of HCV that
Lohmann and Bartenschlager were able to achieve some
success on the way to establishing a cell-culture model suitable
for HCV propagation 11 . They constructed HCV “minigenomes”,
so-called sub-genomic HCV replicons. These replicons
possess all viral proteins that are necessary for the
propagation of the virus genome. However, in place of the
viral structural proteins that are required for the formation
of new virus particles, they carry a resistance gene. This
resistance gene is required to separate cells that propagate
the replicons from other cells that do not: after transfection
of the replicons into the human hepatoma-cell-line Huh-7,
these cells are treated with an antibiotic that is inactivated
by the replicon-encoded resistance gene. Consequently,
only those cells survive that have assumed the replicons
and effectively propagate them (Figure 2). This is a system
with two essential limitations: The formation and release of
viruses as well as the infection process itself could not be
examined, since the sub-genomic replicons were missing
the structural proteins.
The discovery that retroviruses, from which the cognate
envelope protein had been genetically removed, are able
to incorporate the HCV envelope proteins E1 and E2 in a
functional fashion in their virus envelope placed one more
important milestone on the pathway to establishing a cellculture
system for HCV. By using this technique Bartosch
and her colleagues were able to show in 2003 that these

24 RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
Viral pseudoparticle
(HCVpp)
E2
HCV E1
Retroviral
Genome
Retroviral
Capsid
Structural
proteins
HCV Complete genome
Resistence
gene
IRES
Non-structural
proteins
Non-structural
proteins
Manipulation of
HCV Genome
(1) DNA with cloned HCV Genome (2) In vitro Transcription
(4) Transfection
T7-RNA Polymerase
(3) HCV RNA Transcripts
(5) Defection infected cells
(IFM, qRT-PCF)
HCV Replicon
96 h 72 h
Infection
Model for the
HCV
Infection process
Transfection
Selection
HCV Replicon cells
Model for the
HCV
RNA Replication
Huh-7 Hepatoma cells
Manipulation of
the host cell
(e.g. RNAi)
Analysis of
Phenotypes
IFM
α-NS3
Wild type
Mutant
Fig. 2. Important HCV cell-model systems prior to development
of the JFH1 infection system. IRES, internal ribosome entry
point. Graphic: Twincore
Fig. 3. Experimental procedure for examination of HCV replication
with the aid of the JFH1 infection system. IFM, immunofluorescence;
qRT-PCR, quantitative Reverse-Transcriptase Polymerase
Chain Reaction; RNAi, RNA-interference. Graphic: Twincore
mixed-viruses (“HCV-pseudo-particles” = HCVpp), with a
retroviral core and an HCV-typical virus envelope, preferentially
infect liver cells 1 . Since the early stages of infection
are conveyed essentially by the proteins in the virus
envelope (the HCV E1 and E2 proteins, for the HCVpp case),
a system was made available that permitted analysis of the
HCV infection process by means of molecular-biological
technology (Figure 2). It was the developments of 2005 to
2006 that ultimately rendered the methods and approaches
of classic virology accessible to HCV. Together with Wakita
and other colleagues, we have been able to show that a
new HCV-isolate taken from a Japanese patient with a case
of fulminant hepatitis (“Japanese fulminant hepatitis 1” =
JFH1) could not only efficiently replicate in Huh-7 cells, but
it could also yield virus particles in the culture medium 15 .
More over, we were also able to show that these particles
(“cell culture grown HCV” = HCVcc) are infectious not only
in cell cultures, but in vivo as well. By constructing JFH1
variations with structural proteins from other virus isolates,
we substantially enhanced the efficiency level of the infection
system 13 . Further HCV chimeras have been constructed
in the mean time by us and other groups. In this manner,
comparative examinations between different isolates are
now possible as regards the route of transmission and the
mechanisms of virus formation and virus release.
With the aid of molecular-biological techniques we can now
manipulate, in a targeted manner, either the virus genome
or the host cell and subsequently characterise the influence
of these modifications on virus propagation (Figure 3). In
this fashion we gain essential knowledge concerning the
virus’ essential interactions with the host cell, its method of
propagation within the cell, and we can identify new targets
for development of a direct antiviral therapy. Moreover, new
therapy procedures can be evaluated regarding their
effectiveness. Our HCV cell-culture model represents
thereby the central key to solving clinically relevant issues
that are current in HCV research.
The pathway of the virus through the clinic An important
aspect in the management of hepatitis C is prevention,
since the mode of transmission from blood to blood should
be controllable with suitable hygienic measures. Particularly,
transmissions within the hospital environment,
the stability of the virus under variable environmental
conditions and its sensitivity to chemical disinfection
agents are in the focus of our efforts. Examinations and
experiences heretofore have been based, due to a lack of
HCV in vitro models, almost exclusively on studies with the
bovine diarrhoea Virus (BVDV), which is a relative of the
HCV and which has been cultivated for a fairly long time to
date. However, these studies with the surrogate virus for
HCV allow only a semi-dependable estimation of the
infectiousness of the HCV.

RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
25
With the aid of our HCV infection model, we were able to
show that HCV still remains infectious after 28 days at
room temperature, and at 4°C the virus is still infectious
even after 150 days 5 . Since HCV in vivo is associated with
human serum, the influence of human serum from healthy
patients on HCV stability was analysed. The presence of
serum, however, had no influence upon the stability of HCV
particles. Incubation of HCV on various surfaces such as
plastic, steel and gloves indicated a comparable half life of
the virus between these materials.
It is a limiting factor of these studies that it is currently
unknown whether cell-culture viruses and naturally
occurring viruses possess precisely the same characteristics.
However, since in vivo animal studies based on
chimpanzees are controversial due to high costs and ethical
concerns and as small animal models for HCV replication
are still not broadly available, cell culture studies with cell
culture derived HCV represent the optimal model to address
these important questions.
One important finding of our study was that detection of
HCV-RNA does not necessarily correlate with the viral
infectiousness of HCV. Several different published studies
are based nevertheless on a verification of HCV-RNA by
means of a PCR (polymerase chain reaction). Knowledge
concerning the missing correlation between detection of
HCV-RNA and infectiousness must therefore be taken into
consideration in the interpretation of such studies. Moreover,
to evaluate the efficacy of HCV inactivation procedures,
various alcohols and seven commercial disinfection agents
were tested regarding their virucidal characteristics. We
were able to show in this context that certain disinfection
agents led to complete virus deactivation only with
undiluted application 14 . These findings provide a valuable
frame work to define safe hygiene procedures for handling
of HCV-positive materials in the clinical daily routine. We
are currently working on other studies that concern
stability of HCV in body fluids such as breast milk and
semen, since these could also potentially contain either
virucidal substances or factors that may facilitate virus
transmission. Furthermore, it has been planned to examine,
by means of special carrier experiments, the behaviour and
stability of HCV on various surface areas after drying.
This knowledge will contribute to the assessment of
transmission risks and in turn help to prevent new HCV
infections.
The influence of immuno-suppressive drugs on HCV
Once HCV has been transmitted, the infection process
proceeds through the stages described above, frequently
leading to chronic liver disease. HCV-associated liver
failure is one of the most frequent reasons for liver
transplantation 3 . Since in practice neither virus-neutralising
antibodies nor potent antivirals have been made
available that could hinder re-entry of the virus into the
liver, re-infection of the transplanted liver can currently not
be avoided. Consequently, the graft liver is almost universally
re-infected by HCV. Unfortunately, disease progression
after transplantation is usually more rapid than before,
often leading to severe liver disease. In fact, the time span
between transplantation and development of liver cirrhosis
has unfortunately become shorter in the last ten years 2 .
This means that patients who underwent transplantations
in 1988/89 experienced deterioration of liver function by
one fibrosis grade on average within approximately 5 years.
For patients who underwent transplantation in 1996, this
time span amounted to only one year.
In the meantime several factors that may account for this
development have been identified 2,8,9. It must also be
mentioned that patients with a chronic hepatitis C virus
infection after liver transplantation experience a worse
development than patients with other liver diseases. They
have a worse prognosis in comparison to patients with
alcohol- or hepatitis B-associated liver cirrhosis, or primary
sclerosing cholangitis (PSC = a liver disease which involves
destruction of the bile ducts), or autoimmune hepatitis. In
this context the particular kind of immuno-suppressive
drug administered after the transplantation plays a role. An
HCV-specific and optimised immuno-suppression could
contribute to an improvement in the disease process for
these patients. It was a goal in this study to systematically
examine all currently and routinely administered immunosuppressive
drugs as regards their influence on the overall
life-cycle of HCV. We thus examined the entry of the virus,
the RNA replication and the release in the presence of these
drugs utilizing our new HCV in-vitro-system 6 .
With regard to RNA replication, both cyclosporine A and
mycophenolic acid as well as FTY720 act in an inhibitive
manner in the infection model 6,7 . Other frequently used
immuno-suppressive drugs such as tacrolimus, everolimus,
basiliximab and prednisolone had no influence on HCV-
RNA replication. When we examined the complete viral
lifecycle, we discovered important correlations: incubation

26 RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
with higher doses of prednisolone and other steroids
increased HCV infectivity. By means of time-kinetic
experiments, we were able to prove that HCV under
treatment of steroids can penetrate the liver cells more
quickly. With the aid of retroviral pseudotypes, we could
show that the increase in infectiousness was specific for
HCV. Further mechanistic studies revealed that prednisolone
treatment increased the mRNA and the protein expression
of SR-BI and occludin, two proteins that are essential
components of the HCV receptor complex.
It was possible to ablate this steroid effect through the use
of the glucocorticoid receptor inhibitor RU-486, which
indicates that this signalling cascade is responsible for the
influence on the receptor expression and the HCV cell entry.
Moreover, we noted that prednisolone not only facilitated
virus cell entry and spread in the human hepatoma-cell-line
Huh-7.5: the incubation with prednisolone also increased
viral dissemination in dimethyl-sulphoxide-differentiated
liver cells and primary human hepatocytes. This result
correlates with the clinical experience in the patient:
Various clinical studies had reported that administration of
high dosages of steroids, for example in the context of acute
organ rejection treatment, was associated with increased
viral load and augmented liver inflammation 8 . Correspondingly,
avoidance of highly-dosed steroids could contribute to
an improvement in clinical course after HCV-induced liver
transplant.
Hepatitis C – not only in the liver? To date research on
HCV has been primarily focussed on the liver – an obvious
focus, since the symptoms can be attributed predominantly
to damage of this organ. Upon more precise investigation
however, one can see that infection with HCV generates a
series of other symptoms as well, and these symptoms are
not necessarily in direct relation to the liver infection.
Patients with a chronic HCV infection often times present
with various nervous system-associated symptoms. Chronic
fatigue syndrome, depression and cognitive disorders
frequently appear. It is however unclear whether these
symptoms can be attributed to a direct HCV replication in
the brain and the peripheral neuronal cells, or to indirect
effects upon the central and peripheral nervous system. For
this reason we investigated whether host cells from these
tissues – among them human neuroblastoma and glioblastoma
cell-lines and microglial cells – are susceptible to HCV
infection and can propagate the virus 4 . In this context we
identified the human peripheral neuroblastoma cell-line
SKNMC, which expresses all four essential HCV-entry
factors and was susceptible to cell entry by our HCV
pseudo-particles (HCVpp). Our investigations did however
show as well that the virus cannot efficiently propagate in
these cells. Nevertheless, our results point to the fact that
HCV can for all intents and purposes penetrate into
non-hepatic cell types. These interactions may in turn
influence the development of the disease and particularly
extra-hepatic disease manifestations.
Small molecules up against an unsolved problem The
options available for therapy against HCV are currently still
insufficient. Although a series of direct antiviral substances
are in clinical development, undesirable side-effects,
resistance development and genotype-specific antiviral
activity still pose substantial challenges. For this reason, the
on-going search for appropriate therapy measures continues.
The focus is now on combinations of well-tolerated active
compounds that act at different steps within the HCV
replication cycle. In this manner, a combination therapy
with drugs acting via distinct mode of action should prevent
rapid development of viral resistance.
In order to identify substances that disturb the various
steps in the HCV replication cycle, we have developed a new
dual-reporter-gene assay, and adapted it in collaboration
with scientists at the HZI to a high-through-put-screening
procedure (Figure 4). This system encompasses the entire
HCV life-cycle and, therefore, enables identification of
inhibitors against each individual step in viral propagation 10 .
The assay is based on an HCV reporter virus that carries a
gene for a luciferase-enzyme of the firefly (F-Luc; firefly
luciferase). The host cells, which are used for propagation of
HCV, were equally furnished by us with a luciferase gene
from the deep-sea crab gaussia (G-Luc; gaussia luciferase).
Since the two luciferase enzymes use different substrates
and do not influence one another, propagation of the HCV
and the vitality of the cells are simultaneously determined.
In this manner we are able to differentiate between
antiviral molecules and inhibitors of cell growth and
cytotoxic compounds, i.e. separate the wheat from the chaff.
Subsequently we validated the system with the aid of
known HCV inhibitors for cell entry, replication and virus

RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
27
production. Finally, we tested compounds of myxobacteria
taken from the HZI compound library. A series of candidates
emerged from this screening that inhibit either the
virus’ entry into the liver cells or the respective propagation
and exit from the cell. With these molecules it is our
goal, in collaboration with our partners at the HZI and the
myxobacteriologists working with Rolf Müller at HIPS, to
develop lead molecules for new HCV inhibitors. Parallel to
this, we want to use these compounds to better understand
how HCV propagates in liver cells (Figure 5). In this regard
the myxobacterial natural materials provide us with a
unique gateway to facilitate manipulation of cell-biological
processes, and to be able to observe how HCV is influenced
thereby. For these purposes, also those compounds that augment
propagation of HCV are by all means quite important.
Once we have come to understand precisely which processes
in the cell are responsible for these processes, we will be
able to find ways in the future to use this interaction
between virus and cell for therapeutic purposes.
A
Firefl y
”F-Luc”
Deep sea crab
HCV Reportervirus
Liver cell with Reportergene
Virus production
+/- Substance
Fig. 5. Influence of small molecules with antiviral activity
against HCV on the formation of cellular lipid droplets in
liver cells. HCV infected cells were incubated with solvent or
an inhibitor. Lipid droplets were coloured green with an anti-
ADRP antibody (left) or a lipid colouring agent (small picture).
HCV core was detected with a core-specific antibody (on the
right and marked with red in the small picture). Incubation
with the inhibitor leads to expansion and aggregation of lipid
droplets, as well as to the enrichment of HCV core on the
surface. Graphic: Twincore
B
”G-Luc”
Measurement of
reporter activity
Fig. 4. HZI robot in the S3* safety laboratory for execution of
high-throughput screenings. Graphic & Photo: Twincore

28 RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
Prof. Pietschmann and his work group. Photo: Twincore/HZI
Sandra Ciesek born 1978 in Goslar, studied medicine in
Göttingen and Hannover and received her M.D. degree with
honours under Prof. M. P. Manns and Dr. H. Wedemeyer
at the Clinic for Gastroenterology, Hepatology and Endocrinology
at the MHH (Hannover Medical School). Sandra
Ciesek received numerous prizes for her doctoral thesis and
has been working since then as Assistant Physician in the
Department Gastroenterology, Hepatology and Endocrinology
and as a scholarship-funded scientist at TWINCORE with
Prof. Thomas Pietschmann. Under the auspices of an individually
contracted position funded by the German Research
Foundation (DFG), she is now doing research at TWINCORE
on the influence of cyclosporine A on the hepatitis C nonstructural
protein NS2.
Thomas Pietschmann born in 1971 in Würzburg, studied
biology with emphasis on biochemistry, animal physiology,
virology and immunobiology at the University of Würzburg
and the Duke University (Durham, NC, USA). After completing
his studies in 1996, he received his Ph.D. degree
in biology at the Institute for Virology of the University
of Würzburg and subsequently worked as postdoc at the
Institute for Virology in Mainz and in the Department for
Molecular Virology in the University Clinic of Heidelberg.
Thomas Pietschmann established there an independent
research group that investigated the mechanisms of morphogenesis
and cell entry of the hepatitis C virus. From the
year 2006 his group was supported by an Emmy Noether
fellowship from the German Research Community (Deutsche
Forschungsgemeinschaft). In the spring of 2007 he was
appointed with his work group to TWINCORE. He now leads
the Department for Experimental Virology there.
Eike Steinmann born 1978 in Bremen, studied biology at
Leibniz University Hannover, with emphasis on Virology,
Microbiology and Molecular Biology. After a DAAD scholarship
for study at Northeastern University in Boston, he
completed his diploma dissertation under Prof. Herrler at
the Institute for Virology of the Veterinary University of
Hannover. For his Ph.D. thesis Eike Steinmann changed to
the Department for Molecular Virology at the University
Clinic of Heidelberg and did research in the group of Prof.
Bartenschlager regarding the function of p7 protein in the
HCV replication cycle. With his advisor, Prof. Thomas
Pietsch mann, he then was appointed to TWINCORE. His
research is now concentrated on various aspects of the HCV
assembly process and its release, as well as a search for
new antiviral targets. Furthermore, he is examining the
environmental stability and susceptibility of HCV to disinfection
agents.

RESEARCH REVIEWS | Frontier Runners of Hepatitis C Virus
29
Literature
1. Bartosch, B., Dubuisson, J., & Cosset, F. L. (2003) Infectious
hepatitis C virus pseudo-particles containing functional
E1-E2 envelope protein complexes. Journal of Experimental
Medicine 197, 633-642.
2. Berenguer, M., Prieto, M., San Juan, F., Rayon, J. M.,
Martinez, F., Carrasco, D., Moya, A., Orbis, F., Mir, J., &
Berenguer, J. (2002) Contribution of donor age to the recent
decrease in patient survival among HCV-infected liver transplant
recipients. Hepatology 36, 202-210.
3. Brown, R. S. (2005) Hepatitis C and liver transplantation.
Nature 436, 973-978.
4. Bürgel B, Friesland M, Koch A, Manns MP, Wedemeyer H,
Weissenborn K, Schulz-Schaeffer WJ, Pietschmann T, Steinmann
E & Ciesek S (2010) Hepatitis C virus enters human
peripheral neuroblastoma cells - evidence for extra-hepatic
cells sustaining hepatitis C virus penetration. Journal of Viral
Hepatology Jun 23 [Epub ahead of print].
5. Ciesek, S., Friesland, M., Steinmann, J., Becker, B., Wedemeyer,
H., Manns, M. P., Steinmann, J., Pietschmann, T.,
& Steinmann, E. (2010) How stable is the hepatitis C virus
(HCV)? Environmental stability of HCV and its susceptibility
to chemical biocides. Journal of Infectious Diseases 201, 1859-
1866.
6. Ciesek, S., Steinmann, E., Iken, M., Ott, M., Helfritz, F. A.,
Wappler, I., Manns, M. P., Wedemeyer, H., & Pietschmann, T.
(2010) Glucocorticosteroids increase cell entry by hepatitis C
virus. Gastroenterology 138, 1875-1884.
7. Ciesek, S., Steinmann, E., Wedemeyer, H., Manns, M. P.,
Neyts, J., Tautz, N., Madan, V., Bartenschlager, R., von Hahn,
T., & Pietschmann, T. (2009) Cyclosporine A inhibits hepatitis
C virus nonstructural protein 2 through cyclophilin A.
Hepatology 50, 1638-1645.
9. Gane, E. J., B. C. Portmann, N. V. Naoumov, H. M. Smith,
J. A. Underhill, P. T. Donaldson, G. Maertens, and R. Williams.
1996. Long-term outcome of hepatitis C infection after liver
transplantation. New England Journal of Medicine 334:815-
820.
10. Gentzsch, J., Hinkelmann, B., Kaderali, L., Irschik, H.,
Jansen, R., Sasse, F., Frank, R., & Pietschmann, T. (2011).
Hepatitis C virus complete life cycle screen for identification
of small molecules with pro- or antiviral activity. Antiviral
Research 89(2), 136-148.
11. Lohmann, V., Körner, F., Koch, J. O., Herian, U., Theilmann,
L., & Bartenschlager, R. (1999) Replication of subgenomic
hepatitis C virus RNAs in a hepatoma cell line.
Science 285,110-113.
12. Manns, M. P., Wedemeyer, H., & Cornberg, M. (2006).
Treating viral hepatitis C: efficacy, side effects, and complications.
Gut 55, 1350-1359.
13. Pietschmann, T., Kaul, A., Koutsoudakis, G., Shavinskaya,
A., Kallis, S., Steinmann, E., Abid, K., Negro, F., Dreux, M.,
Cosset, F. L., & Bartenschlager, R. (2006) Construction and
characterization of infectious intragenotypic and intergenotypic
hepatitis C virus chimeras. Proceedings of the National
Acdademy of Sciences USA 103, 7408-7413.
14. Steinmann, J., Becker, B., Bischoff, B., Paulmann, D.,
Friesland, M., Pietschmann, T., Steinmann, J., & Steinmann,
E. (2010) Virucidal activity of 2 alcohol-based formulations
proposed as hand rubs by the World Health Organization.
American Journal of Infection Control 38, 66-68.
15. Wakita, T., Pietschmann, T., Kato, T., Date, T., Miyamoto,
M., Zhao, Z., Murthy, K., Habermann, A., Krausslich, H. G.,
Mizokami, M., Bartenschlager, R., & Liang, T. J. (2005) Production
of infectious hepatitis C virus in tissue culture from
a cloned viral genome. Nature Medicine 11, 791-796.
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& Lucey, M. R. (2002) The association between hepatitis C
infection and survival after orthotopic liver transplantation.
Gastroenterology 122, 889-896.

30 RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
The Aging of the Immune System: Challenges and
Perspectives
AUTHORS | Priv.-Doz. Dr. Eva Medina | Research Group Infection Immunology | eme@helmholtz-hzi.de |
Dr. Dr. Luka Cicin-Sain | Junior Research Group Immune Aging and Chronic Infections | lcs09@helmholtz-hzi.de
How old is old? Looking back through human history, the average life expectancy during the Stone Age was 20 to 35 years, just
enough time to allow procreation and ensure the continuation of the human race. In the late 19th and the 20th centuries, however,
life expectancy in industrialized nations exploded, nearly doubling over the last 100 years. The average life expectancy is now ≈75
years for men and somewhat over 80 years for women, with a still upwardly trend and nobody is able to predict where it may end
(Figure 1). Although the news that we are living longer is positive, this fact presents new challenges to both individuals and to
society. The increase in the number of people with advanced age is accompanied by an epidemic of chronic diseases, including
Alzheimer disease, arthritis, atherosclerosis or diabetes. A common denominator of all these conditions is their infl ammatory
pathogenesis, indicating that aging results in a dysbalance of the immune function referred to as “immunosenescence”. This
age-associated deterioration of the immune system is responsible for the increased prevalence and severity of infectious diseases,
as well as the low effi cacy of vaccination in the elderly. Indeed, the elderly population is particularly susceptible to infection and
vulnerable in case of disease (Gavazzi and Krause, 2002). For example, deaths resulting from infl uenza and pneumonia represent
the sixth leading cause of death among persons aged 65 or older in developed countries (Stupka et al., 2009). Vaccinations could
help to overcome this increased risk of infectious death in elderly persons. However, the protective effect of vaccination is also
partially lost in the elderly population (Chen et al., 2009). Understanding the basic mechanisms of immune dysfunction that occur
with age will help to develop interventions to delay or even reverse the detrimental effects of immunosenescence, and thereby
ensure a better protection of the vulnerable elderly population and a “healthy aging”.
Development of pathogen diagnostics: Mechanisms of
immunosenescence The aging process can be viewed
as a progressive breakdown of the homeostatic balance in
an individual. As we age, it becomes increasingly difficult
to react to external stress and restore the homeostasis.
Therefore, the aging host requires increasingly more time
and energy to recover its homeostatic footing, which makes
them comparatively less fit than younger individuals. Systems
that allow an individual to adapt to its environment –
such as the nervous system, which allows the adaptation to
the immediate physical environment or the immune system,
which allows the host to adapt to its microbiological environment
– are particularly affected by the aging process,
because their rapid response to external stimuli is essential
for host survival.
The immune system can be divided into an innate part,
consisting mainly of monocytes, neutrophils, natural
killer and dendritic cells, as well as into an adaptive part,
represented by B and T lymphocytes. The innate immune
system allows prompt reactions to pathogen-associated
molecular patterns (PAMP) but responds indiscriminately
to an infectious threat. The adaptive immune system is able
to respond in a more pathogen-specific manner and has the
ability to remember novel moieties associated with an infec-
tious agent and rapidly recognize them upon reinfection.
Lymphocytes constitute a population of millions of clonally
diverse cells, which can each recognize a different molecular
moiety that is not cognate to the host (antigen). Once a
naïve lymphocyte clone recognizes a novel antigen, it proliferates
to form a large population of cells recognizing and
neutralizing pathogens carrying this moiety. A number of
these lymphocyte clones persists after infection resolution
as memory cells in the host, conferring immunity against
recurrent infections with the same pathogen.
Generally, the aging process can affect both branches of the
immune system, yet the functional losses of the adaptive
branch of the immune system have been exhaustively documented,
while the documented age-related changes of the
innate immune system are less exhaustive and sometimes
contradictory (Nikolich-Zugich and Cicin-Sain, 2010). While
a number of studies showed an increase in the activity of
the innate immune system and the inflammation associated
with aging has been prominently termed “inflammaging”
(Franceschi and Bonafè, 2003), it remains unclear if this is
due to a compensation of the progressively weaker function
of the adaptive immune branch or an intrinsic increase in
the activity of the innate immune system.

RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
31
Fig 2. Loss of naive cytotoxic T-lymphocytes in aged
individuals (adapted from Cicin-sain et al. PNAS 2007). The
population of naive lymphocytes, characterized by low CD95
and high CD28 expression, is typically abundant in young
adult rhesus monkeys, but severely diminished in old ones.
This results in relative increases of memory subsets. Graphic: HZI
Fig. 1. The male (blue histogrammes) or female (red histogrammes)
general population has been traditionally
distributed across age categories to form a pyramid, where
the most numerous individuals were present in the youngest
age cohorts. In Germany, the demographic changes resulted
in an altered, mushroom shaped, distribution. If current
trends persist, in 25 years the population between 65 and 75
years of age will be the most frequent one. Graphic: HZI
On the other hand, there is a clear consensus that aging
adversely affects the adaptive branch of the immune system,
and in particular the response to emerging infections
occurring in individuals that have reached an advanced age
(Nikolich-Zugich and Rudd, 2010). In contrast, the memory
responses to pathogens and vaccines encountered earlier in
life are maintained even in very advanced age (Ammana et
al., 2007). Consequently, clonally diverse naïve lymphocytes
are more profoundly affected by aging than clonally restricted
memory subsets. Therefore, the loss of clonal diversity in
the lymphocyte pool is a prominent change associated with
immunosenescence.
The causes of immunosenescence are multiple and not entirely
understood. Among the well characterized contributors
to immune aging are the thymic involution and the consequent
loss of newly generated naïve T cells (Figure 2) as
well as metabolic stress, in particular oxidative stress (Altmeyer
and Hottiger, 2009). It remains less clear if inflammatory
conditions contribute to immune aging or are merely
a consequence of immunosenescence. Similarly, the role
of chronically persisting infections in immunosenescence
is not entirely understood (Virgin et al. 2009) and it is not
clear if persistent infections contribute to immune aging or
if immunosenescence results in chronic infections. These
questions can be answered by experimental modelling of
the immune processes in aging animals.
Immunosenescence has multiple clinical consequences.
The elderly show a significant increase in the susceptibility
to infections that are controlled by naïve lymphocyte
populations, such as emerging influenza strains, West Nile
Virus infections or pneumococcal infections. This problem
is complicated by the inability of naïve cell populations to
generate protective immunity upon vaccination in elderly
individuals. On the other hand, the elderly are also more
vulnerable to a variety of other conditions, particularly
chronic inflammatory conditions including arthritis,
inflammatory bowel disorders, atherosclerosis or conditions
with a strong inflammatory component like diabetes or
Alzheimer disease.
Our understanding of the cellular and molecular changes
that underlie the decline in immune function with age has
increased significantly in recent years, and clinical trials to
evaluate methods by which to augment immunity in elderly
individuals are now underway.

32 RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
Centenarians as a model for healthy aging Centenarians,
which are by definition very old individuals, are the best
model in which to study human longevity. The centenarians
are persons not only older than 100 years, but also in good
mental and physical condition. Centenarians were once
considered to be exceedingly rare; however this is no longer
the case as their numbers are dramatically increasing. Thus,
according to published predictions, those surviving longer
than 95 or 100 years will soon represent a considerable
population (Olshansky et al., 1990; Barinaga M, 1991). Centenarians
are the best example of successful healthy aging
since they have avoided and/or survived the most important
pathologies that are responsible for the morbidity and
mortality of aged people. In order to be capable of avoiding or
postponing major age-related diseases, centenarians should
be equipped with well preserved and efficient immune and
defence mechanisms and have optimal combinations of appropriate
lifestyle and genetic background (Franceschi and
Bonafè, 2003). Studies of their immune system have revealed
parameters that follow the deteriorating trend often present
in aged people (e.g. reduction of B and T lymphocytes),
whereas other parameters are well preserved (e.g. chemotaxis,
phagocytosis). Whether these components, rather than
environmental and genetic determinants, are responsible for
reaching such as an advanced age still remains to be determined.
Thus, the study of centenarians, and particularly that
of healthy centenarians, is not only of broad biological and
medical interest, but can also help in identifying genes that
prevent the above-mentioned age-related diseases.
Animal models for the study of immunosenescence
Ethical concerns pose considerable limitations on our
ability to perform experiments in humans. Hence, much of
what we have learned is owed to the utility of experimental
cellular or animal models of aging. A number of animal
models have been established to examine the age-associated
effects in the immune function and specific pathways that
have been shown to be altered by aging. Invertebrates offer
pragmatic benefits for such studies as they generally have
short generation times, short life spans and can be raised in
large numbers. However, the translation of the knowledge
gained in invertebrate organisms is complicated by the fact
that invertebrates rely solely on an innate immune response
and lack the components of the adaptive immune system
found in vertebrates. Nevertheless, the lack of components
of the adaptive immune response has the advantage that it
allows the study of effects of age on the innate immune system
while excluding the confounding interactions between
the innate and adaptive immune mechanisms. The most
extensively used invertebrates for aging studies have
been Drosophila melanogaster and Caenorhabditis elegans
(Kurz and Tan, 2004).
Fig 3. The mouse model has proved to be a very useful tool
for studying the ageing process as well as for elucidating
the basis for the increased susceptibility to infection that
accompanies advance age. Photos: HZI
The mouse models (Figure 3), on the other hand, have
provided significant data on the changes in the adaptive immune
response with advancing age (Maue et al., 2009). The
immunosenescence models include mice with alterations in
telomerase activity, tumour suppressor function, oxidative
stress, hormone expression and various other molecules
associated with immune development and differentiation as
well as longevity. A number of unique findings have been
made in these models regarding basic immune function and
the relationship between cellular networking and signalling
pathways associated with longevity and immune function.
Furthermore, the mouse model represents a very useful tool
for evaluating strategies to prevent or reverse the immune
aging process, such as those aimed at improving the ability
of aged animals to successfully respond to vaccination or to
combat pathogen challenge.
One major limitation of extrapolating the findings obtained
with the mouse model of immune aging to immunosenescence
in human medicine has been the relatively short
lifespan of mice (≈2.5 years). A more suitable alternative is
offered by studies performed on aging rhesus non-human
primates (NHP), which are closer to humans both from the
phylogenetic perspective and also in terms of their lifespan
clock (average life expectancy in rhesus monkeys ≈25 years).
On the other hand, research in primate models is limited by
ethical concerns and by technical restrictions. One can only
house and use a limited number of NHP in an experiment
and the advanced genetic models which are freely available
in experimental mice cannot be practically implemented in

RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
33
long-lived hosts. To this end, different models offer varying
benefits in the study of immunosenescence. Identifying the
comparative advantages of individual experimental models
and their use in the appropriate context is key to successful
research.
Immune rejuvenation: more than just a dream? As we
learn more about the cellular and molecular changes that
underlie aging of the immune system, it is likely that new approaches
to reverse this process will be discovered. As immunosenescence
is associated with a decline in naïve B and T
cell production, the repopulation of the immune system by B
and T cells derived from pluripotent stem cells might provide
a means to revert immunosenescence in elderly patients,
provided that adult somatic cells may be reprogrammed into
pluripotent stem cells to raise naïve lymphocytes. Another
potential strategy that provided encouraging results in animal
models is based on the restoration of the thymic epithelium
function and the consequent delay of thymic involution
after treatment with keratinocyte growth factor (Min et al.,
2007). An additional promising strategy for rejuvenation of
the aged immune system is to target lymphocyte production
to increase the number of naïve B and T cells that migrate
to secondary lymphoid organs. Increases in the numbers of
new B and T cells in the circulating pool may improve the
ability of elderly individuals to mount a response to new or
recall antigens. Several interventions, including exercise,
have also been proposed to restore immune function in older
populations. The findings from some, but not all studies, support
the possibility that exercise may attenuate immunosenescence
and, therefore, may be an efficacious therapy for
restoring immune function in the elderly. Another possible
strategy to counteract immunosenescence is to reduce the
antigenic load represented by pathogens, such as influenza
virus and cytomegalovirus (CMV). From this point of view, a
systematic search for chronic viral infections in the elderly
and the establishment of safe procedures to eradicate them
would be likely to have a beneficial impact on longevity.
Because the aging process varies widely between individuals,
it will be important to develop ways of identifying those
patients who would benefit most from immunomodulatory
treatments. Therefore, it would be useful to have simple
biomarkers with which to accurately measure the effects of
aging on the immune system. Progress in this area is already
underway.
Aging research at the HZI Aging research is currently
conducted at the HZI by two Research Groups: Infection
Immunology (INI) and Immune Aging and Chronic Infections
(IMCI). The research efforts of these groups are focused on
identifying age-related changes in the immune function in
the hope of developing intervention strategies to delay or prevent
the decline in the functionality of the immune system.
Using a mouse model of aging, the researchers from the
Group Infection Immunology have investigated specific
deficiencies in the immune function that lead to age-related
increased susceptibility to infection with the bacterium
Streptococcus pyogenes (Goldmann et al., 2010). This pathogen
is an important cause of severe, life-threatening infections
among the elderly population. They found that, similar
to humans, young animals (aged two to three months) are
able to combat the infection successfully while aged mice
(older than 20 months) died even if they were infected with
fewer bacteria (Figure 4).
The immune mechanisms affected by immunosenescence
that make aged mice more susceptible to bacterial infections
are also a focus of research in this Group. They centre their
studies on macrophages because these cells are the first line
of defence for combating bacterial infections. They have found,
for example, that the amount of macrophages is highly reduced
in the tissue of aged mice compared to young animals
(Figure 5). As the amount of macrophages in the organs depends
on the production of a specific growth factor (M-CSF),
the researchers have investigated whether treatment with
this growth factor could induce repopulation of resident tissue
macrophages in aged mice and increase their resistance
during infection. Indeed, the treatment with M-CSF made
aged mice much more resistant and they were able to fight
the infection much better. The outcome of these investigations
indicates that repeated prophylactic administration of M-
CSF can help to maintain the macrophage compartment in
the elderly and the fitness of the immune system.
Fig 4. Mice exhibit the age-related loss of resistance to
Streptococcus pyogenes seen in humans. This figure shows
that young mice can recover and survive when they are
infected with S. pyogenes (open symbols). On the other
hand, aged mice (closed symbols) are very susceptible
to streptococcal infection and all mice died shortly after
bacterial inoculation. Graphic: HZI

34 RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
Fig 5. Tissue macrophages are very important cells for
combating S. pyogenes infection. This figure shows that
aged mice had a significantly lower number of resident tissue
macrophages (indicated by an arrow) than young animals.
This can be one of the reasons for the elevated susceptibility
of aged mice to S. pyogenes infection. Graphic: HZI
The newly established Immune Aging and Chronic Infections
Group focuses on the mechanisms and effects of immune
aging. In a recent collaborative study, the Group was
able to show that aging NHP (≈18-25 yrs old) show significantly
weaker immune responses to vaccination than adults
(≈7-10 yrs old), and that this response could be predicted by
the relative and absolute count of naïve cells in the blood of
monkeys (Cicin-Sain et al., 2010) (Figure 6). The vaccination
was performed with an attenuated virus (modified vaccinia
Ankara – MVA) that may be realistically considered for vaccinations
in elderly populations. Interestingly, the naïve cell
counts and vaccine responses correlated only if the fraction
of naïve cells as a pool of total lymphocytes dropped below
a threshold of approximately 20%. Therefore, the clonal
diversity of naïve T cells appeared to define the intensity of
responses to the vaccine only if the clonality was a limiting
factor. In a follow up study submitted for publication the
authors show that the same population of NHP can mount
vigorous memory responses to a pathogen to which they
were exposed early during their life (Rhesus cytomegalovirus
– RhCMV). The Group now focuses on the effects that
the persistent infections exert in a chronically infected host
and their contribution to the immune aging process.
Conclusions To conclude, at present aging must be considered
an unavoidable end point of the life history of each
individual. Nevertheless, our increasing knowledge about
the mechanisms regulating aging and immunosenescence
allows us to envision many different strategies to cope with
and delay the processes in order to endow everybody with a
long and healthy life.
Fig 6. Vaccination with an attenuated poxvirus (Modified
vaccinia Ankara strain) results in detectable immune
responses of tissue resident cytotoxic T-lymphocytes in adult
rhesus monkeys, but not in aged ones. This loss of immune
responses correlates directly to the loss of naive cells in aged
hosts (see figure 2). Graphic: HZI
Eva Medina is the head of the Infection Immunology
Research Group at the HZI. Eva Medina graduated in Biological
Sciences at the University of Seville (Spain) in 1987
and obtained her PhD at the Medical School of Seville in
1990. She worked as a postdoctoral researcher from 1991 to
1993 in the Immunology Department at the Royal Free Hospital
School of Medicine in London. In 1993, she undertook a
postdoctoral fellow position at The Trudeau Institute in New
York studying several aspects of the host response to infection
with Mycobacterium tuberculosis. From 1997 to 2004 she
worked as a postdoc at the Department of Microbial Pathogenesis
and Vaccine Research in the HZI. On 2004 Eva Medina
was appointed as Head of the Infection Immunology
Research Group in the same institution.

RESEARCH REVIEWS | The Aging of the Immune System: Challenges and Perspectives
35
Franceschi, C., & Bonafè, M. (2003) Centenarians as a model
for healthy aging. Biochemical Society Transactions 31(2),
457-61.
Gavazzi, G., Krause, K. H. (2002) Ageing and infection. Lancet
Infectious Diseases 2(11), 659-66.
Luca Cicin-Sain born in 1972, is head of the junior research
group Immune Aging and Chronic Infections at the Helmholtz
Centre for Infection Research (HZI). He studied at the
University of Rijeka, Croatia, and obtained his MD in 1996
and his PhD in 2006. From 2001 to 2004 he worked at the
Max von Pettenkofer Institute in Munich and then moved to
the Oregon Health and Science University in Portland, OR,
USA, where he worked as a postdoctoral DFG fellow until
2007 and as a Research Assistant Professor until 2009. In
2009 he moved back to Germany and joined the HZI in his
current position. In 2011 he was awarded the European Research
Council Starting Grant to expand his research focus
on the molecular and cellular mechanisms of herpesviral
antigenic induction and on the effects of persistent infections
on immune homeostasis and aging.
Literature
Altmeyer, M., & Hottiger, M. O. (2009) Poly(ADP-ribose)
polymerase 1 at the crossroad of metabolic stress and inflammation
in aging. Aging (Albany NY) 1(5), 458-69.
Amanna, I. J., Carlson, N. E., & Slifka, M. K. (2007) Duration
of humoral immunity to common viral and vaccine antigens.
New England Journal of Medicine 357(19), 1903-15.
Barinaga, M. (1991) How long is the human life-span? Science
254(5034), 936-8.
Chen, W. H., Kozlovsky, B. F., Effros, R. B., Grubeck-Loebenstein,
B., Edelman, R., & Sztein, M. B. (2009) Vaccination in
the elderly: an immunological perspective. Trends in Immunology
30(7), 351-9.
Cicin-Sain, L., Smyk-Paerson, S., Currier, N., Byrd, L., Koudelka,
C., Robinson, T., Swarbrick, G., Tackitt, S., Legasse, A.,
Fischer, M., Nikolich-Zugich, D., Park, B., Hobbs, T., Doane,
C. J., Mori, M., Axthelm, M. T., Lewinsohn, D. A., & Nikolich-
Zugich, J. (2010) Loss of naive T cells and repertoire constriction
predict poor response to vaccination in old primates.
Journal of Immunology 184(12), 6739-45.
Goldmann, O., Lehne, S., & Medina, E. (2010) Age-related
susceptibility to Streptococcus pyogenes infection in mice:
underlying immune dysfunction and strategy to enhance immunity.
Journal of Pathology 220(5), 521-9.
Kurz, C. L., & Tan, M. W. (2004) Regulation of aging and innate
immunity in C. elegans. Aging Cell 3(4), 85-93.
Maue, A. C., Yager, E.J., Swain, S. L., Woodland, D. L., Blackman,
M. A., & Haynes, L. (2009) T-cell immunosenescence:
lessons learned from mouse models of aging. Trends in Immunology
30(7), 301-5.
Min, D., Panoskaltsis-Mortari, A., Kuro-O., M., Holländer, G.
A., Blazar, B. R., & Weinberg, K. I. (2007) Sustained thymopoiesis
and improvement in functional immunity induced
by exogenous KGF administration in murine models of aging.
Blood 109(6), 2529-37.
Nikolich-Zugich, J. & Cicin-Sain, L. (2010) Aging of the Immune
System Across Different Species. In: Comparative Biology
of Aging (Wolf, N., ed.), Springer Science and Business
Media, New York, pp 232-249.
Nikolich-Zugich, J., & Rudd, B. D. (2010) Immune memory
and aging: an infinite or finite resource? Current Opinion in
Immunology 22, 535-40.
Olshansky, S. J., Carnes, B. A., & Cassel, C. (1990) In search
of Methuselah: estimating the upper limits to human longevity.
Science 250(4981), 634-40.
Stupka, J. E., Mortensen, E. M., Anzueto, A., & Restrepo, M. I.
(2009) Community-acquired pneumonia in elderly patients.
Aging Health 5(6), 763-774.
Virgin, H. W., Wherry, E. J., & Ahmed, R. (2009) Redefining
chronic viral infection. Cell 138(1), 30-50.

36 RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
Novel Nanoparticle-based Technology Platform for the
Delivery of Vaccine Antigens
CORRESPONDING AUTHOR | Prof. Dr. Claus-Michael Lehr | Department of Drug Delivery, Helmholtz-Institute
for Pharmaceutical Research Saarland – HIPS | cle09@helmholtz-hzi.de | Chair for Biopharmaceutics and
Pharmaceutical Technology, Saarland University, Saarbruecken | lehr@mx.uni-saarland.de
CO-AUTHORS | Dr. Steffi Hansen | Research Group Transdermal Drug Delivery – HIPS | Prof. Dr. Ulrich F. Schäfer |
Chair for Biopharmaceutics and Pharmaceutical Technology, Saarland University, Saarbruecken | Prof. Dr. Dr. Carlos
A. Guzmán | Department of Vaccinology and Applied Microbiology – Helmholtz Centre for Infection Research – HZI
Infections are responsible for approximately one third of all deaths occurring each year in the world. In addition infectious agents
are also directly involved in the pathogenesis of many cancers and chronic diseases. Furthermore, infections are often the fi nal
cause of death in patients affected by other primary diseases (e.g. trauma, cardiovascular or respiratory syndromes). Thus, it is of
the upmost importance to develop strategies to prevent and treat infectious diseases. In this context vaccines are the most
cost-effi cient tool to prevent infections. Moreover, their therapeutic application for both infectious and non-infectious diseases is
attracting general interest.
Classical vaccines were based on live attenuated or inactivated
pathogens. However, due to their complex and
ill-defined nature, such vaccines can vary in quality from
batch to batch and can induce adverse events. In recent
years we have seen the advent of subunit vaccines. These
preparations are based on the use of well-defined subcellular
components of the corresponding pathogen which are
in turn critical for the stimulation of a protective response.
Subunit vaccines can be prepared with native components
derived from the pathogen or obtained by DNA-recombinant
technologies or in vitro synthesis (e.g. recombinant proteins,
synthetic peptides, capsular carbohydrates, etc.). Classical
vaccines were traditionally very immunogenic, due to the
complex nature of these formulations and the presence of
pathogen-derived components with built-in adjuvant properties.
In contrast, purified components are usually very
poor immunogens rendering essential the incorporation of
adjuvants in the formulation. Adjuvants do not only allow
to improve the overall strength of the elicited responses but
also to reduce the amount of antigen needed and the time
required to achieve a threshold of protective immunity as
well as to modulate the quality and expand the breadth of
the elicited response. Finally, adjuvants enable the stimulation
of long lasting memory responses, thereby reducing the
need for frequent boost vaccinations.
Most of the traditional vaccines have been administered
by subcutaneous or intramuscular injection. However, the
use of this route is associated to lack of acceptance by the
public and safety issues (e.g. risk of contamination). It also
requires skilled health personnel which in turn represents
a logistic constraint. In addition, an injection (e.g. intramuscular)
does not deliver the vaccine optimally to antigen
presenting cells (APCs) which are the relevant target to
prime naïve T cells to initiate an efficient adaptive immune
response. In fact, only a limited number of APC are present
in the muscles. In contrast, dendritic cells are present in
high densities in the dermis, and Langerhans cells are
located in the epidermis accumulating around the hair
follicles. Therefore, it is highly attractive to target vaccines
to the skin immune system by chemical, mechanical or
nano-technological means and devices thereby promoting
access of vaccine antigens to these APCs. In fact, intradermal
immunization can induce strong mucosal and systemic
immune responses, as well as protection against infections.
Fig. 1. Penetration of methylene blue into porcine skin.
The dye penetrates deep into the hair follicle. Photo: HIPS/HZI

RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
37
At the same time the skin and especially the horny layer of
the stratum corneum provides nearly perfect protection not
only against the invasion of chemicals or pathogens but also
against therapeutics topically applied to the skin. Thus, the
highly compact structure of the stratum corneum with alternating
hydrophilic and lipophilic areas presents, indeed, a
very effective first line defense, in particular against large
and hydrophilic molecules. Furthermore, purified antigens
are highly unstable when applied in their native state to
the skin. Innovative delivery strategies are urgently needed
(i.e. suitable carrier devices or formulations), which enable
antigen stabilization and facilitate permeation.
Pathways for passive diffusion of small molecules have been
identified, among them the inter- and intra-cellular paths,
as well as permeation via the appendages, such as hair follicles
and pores. The trans-appendage pathway is especially
interesting for the permeation of larger molecules since
it allows sidestepping the tight stratum corneum. Cornified
cells are present only in the upper thirds of the hair
follicles, whereas the lower part is lined with an epithelium
containing tight junctions. However, Langerhans cells accumulating
around the follicles build up a second immunologic
line of defense in places where the mechanical barrier is
weakened. Although Langerhans cells represent only about
1% of the total cell population in the skin, they cover a large
surface area of approximately 20% through their horizontal
orientation and long interdigitating protrusions. 2
Transdermal Vaccination – Innovative Approaches
Various approaches have been pursued to deliver antigens
across the skin. New technologies are in progress to ensure
the targeting of vaccine compounds to the APCs present in
the epidermis. Membrane permeabilization agents such as
cholera toxin used on patches that were combined with high
doses of antigens induced significant immune responses. 5
New approaches which include micro-needles 6 , Gene gun or
PowderJect technologies 7 , electroporation, iontophoresis and
dermal abrasion have been proposed to facilitate DNA and
protein vaccine delivery by the percutaneous route. 8
In a clinical phase 1 trial including 11 healthy volunteers,
the normal seasonal flu vaccine was applied either by
conventional intramuscular injection or transcutaneously to
an area of 16 or 32 cm 2 , respectively, on the upper arm after
weakening the horny layer barrier. Previously the horny
layer at the application site had been pretreated with superglue
and adhesive tape. The procedure was generally well
tolerated leading to a slight erythema in a single volunteer.
The immune response to both immunization methods was
Fig 2. Penetration of methylene blue into mouse skin. The dye
penetrates deep into the living skin layers and the hair follicle
(arrow). Photo: HIPS/HZI
comparable. Transcutaneously immunized volunteers
exhibited both CD4+ and CD8+ T-cell responses, whereas the
latter was missing in conventionally immunized volunteers. 9
The common principle of these permeability enhancing
methods is that the mechanical action helps to overcome the
very effective stratum corneum barrier to facilitate the access
of the antigen to the APCs. It was further hypothesized that
the mechanic action leads to unspecific pre-activation of
immune cells, thereby improving antigen-specific immune
responses. A historical example where this concept was
successfully used is the immunization against smallpox.
An effective and strong immune response was induced
subsequent to the degradation of the skin barrier as a result
of scarification.
Several such physical permeability enhancing methods
have been combined with innovative formulations of vaccines
in order to achieve an improved immune response,
reduce the risk of severe side effects and lower the required
dose. To this end different delivery vectors such as liposomes,
virosomes, transferosomes, nano- or micro-beads
and viral vectors have already been tested in animal
models. 2 It has been demonstrated both in vitro and in vivo
that microparticles with a size below 5 μm are ingested by a
wide variety of phagocytic cells. 10 For submicron particles,
there are contradictory reports in the literature with regard
to the differential uptake of nanoparticles by APCs, as
compared to microparticles, depending on the nature of the
particle, the involved antigen, the route of delivery and the
study endpoint. There is indeed evidence that nanoparticles
show increased uptake by APCs, which suggests that they
may be better inducers of immune responses than micro-

38 RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
particles. 11 In addition, it was shown that carrier size could
play a role in determining the type of response induced.
Virus-like particles induced IFN-γ and cell-mediated type
1 responses, whereas larger beads mostly induced type 2
responses. 12
However, strategies for transdermal vaccination that reduce
the protective barrier function of the stratum corneum for a
significant period of time are intrinsically problematic due
to the risk of invasion of pathogens. This imposes a limit to
such methods for mass vaccination campaigns in countries
with critical hygienic conditions and calls for alternatives
that leave the stratum corneum intact. Therefore, the transappendage
pathway seems to be an appealing alternative to
circumvent the stratum corneum barrier and directly target
Langerhans cells via the hair follicles.
Fig 3. Overlay of transmission and fluorescence image of
a longitudinal section of skin from a porcine ear. Green
fluorescent nanoparticles (ca. 500 nm) applied to the ear
accumulate in the opening of the hair follicle. Photo: HZI
Development of pollen-mimetic vaccine particles
Scientists from HIPS and Saarland University, in collaboration
with the Charité Universitätsmedizin Berlin¸
recently showed that nanoparticles accumulate selectively
in the openings of hair follicles. 13 These nanoparticles were
made of the biodegradable and biocompatible polymer
polylactic-co-glycolic acid (PLGA) using polyvinyl alcohol
as a stabilizer.
A plain aqueous nanoparticle suspension as well as a
hydrogel formulation that was massaged into pig ear skin
in vitro, permeated deeper and to a greater extent into hair
follicles than an aqueous solution applied with similar
physical force. It was hypothesized that this is due to a passive
targeting effect, elicited by the nano-scale size of the
delivery device. We could further show that such particles
may release different encapsulated compounds that then
enter the stratum corneum.
As outlined above, the trans-follicular route appeals as a
very attractive and safe strategy for the delivery of vaccine
antigens to APCs. Hair bulbs are also an excellent reservoir,
being only slowly cleared by hair growth and sebum production.
16 An activation of the APCs via this route is a commonly
occurring phenomenon in allergic contact dermatitis
in people allergic to pollen antigens. In fact, pollen can be
considered as a micrometer sized carrier that accumulates
in the follicle opening, sebaceous gland or dermatoglyphs
(skin folds). Antigen release from the pollen is triggered by
a moist atmosphere with sufficient humidity, such as occurs
on the skin by sweating. The idea is now to mimic pollen
delivery in order to vaccinate across the intact skin barrier.
Therefore, we are currently working to create pollen-mimetic
carriers allowing antigens to be stably encapsulated in
order to target the hair follicles. This would allow releasing
the antigen at the site of action in a controlled fashion in
order to activate skin APCs, thereby eliciting an effective
humoral and cellular immune responses. Co-encapsulation
of different adjuvants would enable to strengthen the
overall responses, as well as to modulate them according to
the specific needs. First experiments show that the model
antigens can be encapsulated into PLGA particles. These
particles can be freeze-dried to be stored in a stable form.
They can then be re-dispersed in water directly before the
experiment to be tested in vitro or in vivo. The particles shall
later be formulated as a gel or lotion making them easy to
manufacture, acceptable to patients and safe to use.

RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
39
The research group at the Department of Prof. Dr. Claus-Michael Lehr at the University of Saarbrücken / HIPS Photo: HZI
The research group of Prof. Dr. Dr. Carlos Guzmán at HZI
Photo: HZI
Claus-Michael Lehr studied pharmacy at the universities of
Mainz and Hamburg. After that, he wrote his PhD thesis at
the Leiden University in the Netherlands. In 1991, he started
as a postdoc at the University of Southern California in Los
Angeles, USA, and returned to the Netherlands in the following
year, where he worked at the Leiden/Amsterdam Centre
for Drug Research. In 1993, Claus-Michael Lehr became professor
for pharmaceutical technology at the Phillips University
Marburg and, since 1995, is the head of the department
Biopharmaceutics and Pharmaceutical Technology at the
Saarland University in Saarbrücken. He was engaged in the
founding of the Across Barriers GmbH Saarbrücken in 1998
and co-founded the Centre for Bioinformatics Saarbrücken in
2000, where he still is the deputy director. His successful research
was rewarded with numerous awards, for example the
International Prize of the Belgian Society of Pharmaceutical
Sciences (2009) and the APV Research Award for Outstanding
Achievements in the Pharmaceutical Sciences (2006).
In 2010 he was became „Fellow“ of the American Association
of Pharmaceutical Scientists (AAPS). So far only 5 scientists
from Germany have received this recognition.
Since 2009 Claus-Michael Lehr leads the department Drug
Delivery of the newly founded Helmholtz Institute for Pharmaceutical
Research Saarland (HIPS) and is co-founder of
the PharmaBioTec GmbH in Saarbrücken.

40 RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
In 2005 he was appointed as Head of the new HZI Department
of Vaccinology. He has been working in the field of
Vaccinology since 1989. His work has been instrumental for
the development of new adjuvants and the establishment of
Salmonella spp. as a delivery system for DNA vaccines and
therapeutic molecules.
Steffi Hansen studied pharmacy at Ernst-Moritz-Arndt
Universität, Greifswald and graduated 2004 with a diploma
in pharmacy. 2004 she also received her approbation as a
pharmacist. From 2005 to 2009 she joined the department of
Prof. Claus-Michael Lehr at Saarland University to prepare
her doctoral thesis entitled „Development of a physiology
based diffusion model to predict dermal absorption using
experimental input parameters“. Subsequently Steffi Hansen
started a postdoc at University of Cincinnati, USA sponsored
by the German Academic Exchange Service (DAAD). She
came back to Saarland to the newly founded Helmholtz-Institute
for Pharmaceutical Research in 2010 where she works as
a project leader on new strategies for transdermal vaccinations
on immune homeostasis and aging.
Carlos A. Guzmán born in 1959, is Head of the Department
of Vaccinology and Applied Microbiology at the Helmholtz
Centre for Infection Research (Braunschweig, Germany) and
APL-Professor at the Medical School of Hannover. He is also
external Lecturer of the Doctorate in Biotechnology from the
University of Catania (Italy). He graduated in Medicine at
the National University of Rosario (1981) and obtained his
Board Certification in Medical Bacteriology (1986) in Argentina.
In 1987, he moved to the Institute of Microbiology from
the University of Genoa (Italy), as Research Fellow from the
Italian Foreign Office Ministry. In Italy he also graduated as
Doctor of Medicine and Surgery and obtained his Doctorate
of Research in Microbiological Sciences. In 1994 he moved
to Germany, where he became Head of the Vaccine Research
Group at the German Research Centre for Biotechnology.
Ulrich Schäfer studied Pharmacy at Kiel University
from 1971 to 1974 and qualified as pharmacist in 1974. He
obtained his Ph.D. at the Saarland University in 1980 for his
thesis work on dissolution processes of poorly soluble
substances. In 1994, he was nominated as a so-called
‚Fachapotheker‘ for Pharmaceutical Technology. In 1995, he
was permitted to further educate Pharmaceutical Technology
as a professional training. Dr. Schäfer is member of the
CRS, the APV, and the DPhG. His research interests are the
biopharmaceutical investigations of semisolid dosage forms
for dermatological applications, the development of ex vivo
models for the determination of penetration of drugs and
preservatives into human skin, the optimization of (trans)
dermal dosage forms, and the solubility improvement of
poorly soluble drugs. In December 2010, he was appointed
APL Professor from the President of Saarland University as
appreciation for his commitment and achievements in the
Pharmaceutical Sciences.

RESEARCH REVIEWS | Novel Nanoparticle-based Technology Platform for the Delivery of Vaccine Antigens
41
Literature
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Compans, R. W., & Skountzou, I. (2010) Adjuvanted influenza
vaccine administered intradermally elicits robust long-term
immune responses that confer protection from lethal challenge.
PLoS ONE 5(5), e10897.
2. Babiuk, S., Baca-Estrada, M., Babiuk, L. A., Ewen, C., &
Foldvari, M. (2000) Cutaneous vaccination: the skin as an
immunologically active tissue and the challenge of antigen
delivery. Journal of Controlled Release 66(2-3), 199-214.
3. Hansen, S., Henning, A., Naegel, A., Heisig, M., Wittum,
G., Neumann, D., Kostka, K. H., Zbytovska, J., Lehr, C. M.,
& Schaefer, U. F. (2008) In-silico model of skin penetration
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68(2), 352-367.
4. Mitragotri, S. (2003) Modeling skin permeability to hydrophilic
and hydrophobic solutes based on four permeation
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5. Glenn, G. M., Rao, M., Matyas, G. R., & Alving, C. R. (1998)
Skin immunization made possible by cholera toxin. Nature
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6. Koutsonanos, D. G., Martin, M. d. P., Zarnitsyn, V. G., Sullivan,
S. P., Compans, R. W., Prausnitz, M. R., & Skountzou,
I. (2009) Transdermal influenza immunization with vaccinecoated
microneedle arrays. PLoS ONE 4(3).
7. Arora, A., Prausnitz, M. R., & Mitragotri, S. (2008) Microscale
devices for transdermal drug delivery. International
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for transcutaneous vaccination. Comparative Immunology,
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10. O’Hagan, D. T., Singh, M., & Ulmer, J. B. (2004) Microparticles
for the delivery of DNA vaccines. Immunological
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11. Foged, C., Brodin, B., Frokjaer, S., & Sundblad, A. (2005)
Particle size and surface charge affect particle uptake by human
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12. Fifis, T., Gamvrellis, A., Crimeen-Irwin, B., Pietersz, G. A.,
Li, J., Mottram, P. L., McKenzie, I. F., & Plebanski, M. (2004)
Size-dependent immunogenicity: therapeutic and protective
properties of nano-vaccines against tumors. Journal of Immunology
173(5), 3148-3154.
13. Lademann, J., Richter, H., Teichmann, A., Otberg, N.,
Blume-Peytavi, U., Luengo, J., Weiß, B., Schaefer, U. F., Lehr,
C.-M., Wepf, R., & Sterry, W. (2007) Nanoparticles - An efficient
carrier for drug delivery into the hair follicles. European
Journal of Pharmaceutics and Biopharmaceutics 66(2),
159-164.
14. Luengo, J., Weiss, B., Schneider, M., Ehlers, A., Stracke, F.,
König, K., Kostka, K.-H., Lehr, C.-M., & Schaefer, U. F. (2006)
Influence of nanoencapsulation on human skin transport
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190-197.
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F., & Schneider, M. (2006) Multiphoton microscopy for the
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2233.
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U., Teichmann, A., Otberg, N., & Sterry, W. (2006) Hair follicles
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180(3), 1482-1489.

42 RESEARCH REVIEWS | Mycobacterial Phagosomes and Innate Immunity
Mycobacterial Phagosomes and Innate Immunity
AUTHOR | Dr. Maximiliano G. Gutierrez | Junior Research Group Phagosome Biology | mgg08@helmholtz-hzi.de
Although potential pathogens are encountered routinely, only on rare occasion they do cause diseases. The vast majority of
pathogenic agents are eliminated rapidly by innate defenses operating in our bodies.
In 1882, Ilya Metchnikoff in a revealing experiment observed phagocytes surrounding and attempting to avidly eat a splinter he had
introduced into a transparent starfi sh larva. At that time, he postulated that phagocytes are critical part of the host immune
defense and elaborate the theory of phagocytosis and the evolutionary perspective from eating for nutrition to eating for defense.
In 1908, in recognition of his work on immunity, he was awarded the Nobel Prize in Physiology or Medicine, jointly with Paul
Ehrlich. Metchnikoff’s theory identifi ed and established the basis of our current knowledge of phagocytosis and the innate immune
response (reviewed in Tauber, 2003).
Phagocytosis Phagocytosis is the process by which professional
and non-professional phagocytes and other cells
ingest particles whose size exceeds 1 μm. The resulting
intracellular vacuoles, termed phagosomes, go through
dynamic changes that modify the composition of both their
limiting membrane and their contents, by a sequence that
resembles the progression of the endocytic pathway. This
process is referred to as phagosome maturation, and confers
the vacuole with degradative properties, which are central
to its microbicidal function, representing the first line of defense
against infection that vertebrates possess (Haas, 2007).
Phagosome maturation and immunity Advances in the
understanding of phagosome biology have been possible by
the use of the latex bead system. This model was originally
introduced by Weisman and Korn in the late 1960s and rediscovered
in the early 1990s (Desjardins et al, 1994). Latex
beads are a versatile system for both in vitro and in vivo
analyses of many phagosomal functions. This system allows
an easy approach to understand the very complex process
of phagocytosis (Figure 1). Because the latex floats in
sucrose gradients, latex bead phagosomes are much simpler
to isolate in pure fractions in order to analyse their content
(Figure 2). Much of what we understand today about phagocytosis
and phagosome biology in cellular systems has
been obtained from studies from many groups that use the
latex-bead phagosome system (Desjardins & Griffiths, 2003).
Moreover, it is possible to coat these beads with different
proteins or with specific ligands, which selectively bind to
cellular receptors to study the impact during internalisation.
Recently, it has been shown that particular substrates can
also be coupled to the beads to monitor enzymatic activity
in real-time (Yates et al, 2005).
By ingesting microbial pathogens, phagocytic cells achieve
two essential immune functions orchestrated by the phagosome.
First, phagosomes initiate microbial elimination, in
part by directing ingested microbial pathogens to lysosomes
in a process known as phagosome maturation pathway.
Second, phagocytic cells make use of phagocytic pathway
to direct processed protein and lipid antigens to the Major
Histocompatibility Complex (MHC) class I, MHC class II
and CD1 positive compartments. This loading and exposure
of antigens on plasma membrane allows the development
of a more customised adaptative immune response. Thus,
phagosomes play a dual role: as an innate immune effector
and as a bridge between the innate and acquired immune
system (Figure 3). Actually, phagosome maturation allows
the shaping of a cellular compartment where killing,
degradation and antigen processing take place in a very
organised manner (Jutras & Desjardins, 2005). The study of
phagosome biology is evolving rapidly and reflects recent
advances in a broad range of disciplines including cell biology,
proteomics, and immunology. A better understanding
of the mechanisms of phagosome formation and maturation
and its interactions with intracellular compartments will
Fig. 1. Phagocytosis of latex beds. RAW 264.7 macrophages
expressing Rab34-GFP (green) ingesting avidly IgG-coated
latex beads (red). Nucleus is shown in blue. This process
mimics the IgG-opsonised bacteria that also enter macrophages
via Fcγ receptors (FcγRs). A. Beads at different stages
of internalisation. B. 3D diagram of the localisation of Rab34
on the phagosomal membrane. Photo: HZI, Kasmapour

RESEARCH REVIEWS | Mycobacterial Phagosomes and Innate Immunity
43
allow the development of novel strategies to modulate the
innate and adaptive immune response.
Fig. 2. Latex bead phagosome purification. The scheme shows
the purification of latex bead phagosomes from macrophages
for biochemical and proteomics analysis. Briefly, latex beads
are internalised and then isolated in a sucrose gradient using
its property of flotation. Electron microscopy pictures show
the different steps during the process of purification. Isolated
LBP are used for different biochemical assays and proteomics.
Photo & Graphic: HZI
Mycobacterial phagosomes: the subverted compartment
More than 1.5 million people die every year from tuberculosis
(WHO, TB report 2008) and the appearance of multiand
extensive-drug-resistant strains in eastern Europe is
now moving swiftly towards western European countries
(www.eurotb.org). Although 3.2 billion people are infected
with Mycobacterium tuberculosis, only in 10% of those people
the bacteria do cause disease. The other 90% control the disease
and rely on innate and acquired immune processes of
the body that contain and restrict bacterial growth (Korbel et
al, 2008). Therefore, in most of the cases the innate immune
system is able to effectively eradicate the infection with M.
tuberculosis. The aim of our research is to give insights into
the host factors that facilitate this. The exact mechanisms
that govern this initial sterilising reaction are not completely
understood. Therefore, understanding how host cells
eliminate mycobacteria is important to find possible novel
therapeutic strategies that enhance this natural response.
To live within eukaryotic cells, through evolution Mycobacterium
tuberculosis has developed an impressive set
of mechanisms to subvert phagosome maturation. Many
clinical manifestations and problems during treatment of
tuberculosis are a direct consequence of a population of
intracellular bacilli (Russell, 2001). The key event during
M. tuberculosis infection is the ability of this pathogen to
survive within phagosomes in host cells (Figure 4). This
Fig 3. The phagosome links the innate and adaptative immune
response. The phagosome of M. tuberculosis plays two functions.
The first one is the continuous acquisition of bactericidal
properties. This important step in the development of the
innate immune response is achieved mainly by the interaction
of the phagosome with late endosomes and lysosomes (left
side). On the other hand, in the phagosome bacteria are
degraded and lipids and peptides are presented in the context
of MHC class I, class II and CD1 molecules. This activates
different populations of lymphocytes that participate in the
onset of the adaptive immunity. Some of these pathways are
actually impaired by M. tuberculosis at different levels but
are omitted for clarity.
Fig. 4. Visualising mycobacterial phagosomes in macrophages.
RAW264.7 macrophages infected with mycobacteria.
Cells were visualised using confocal microscopy (Green:
mycobacteria; Blue: nucleus; Red: actin cytoskeleton). Same
infected macrophages can be observed by electron microscopy.
In this case the lysosomes are loaded with BSA-Gold. As
shown, lysosomes are not fusing with the phagosomes
containing mycobacteria. Photo: HZI, Gutierrez

44 RESEARCH REVIEWS | Mycobacterial Phagosomes and Innate Immunity
capability is linked to the aptitude of the live pathogen to
manipulate phagosome maturation. The strategy by which
M. tuberculosis manipulates the cellular trafficking machinery
is likely to be both multifactorial and complex. In spite
of the significant progress in the last few years, a mechanistic
understanding of how M. tuberculosis infection proceeds
at the cellular level and how the first steps of host defenses
are organised against this pathogen is still elusive.
Our research focuses on the mechanisms whereby M. tuberculosis
arrests phagosome maturation and avoids killing
by macrophages. Towards this goal, we are studying the
intracellular transport of non-pathogenic and pathogenic
mycobacteria in macrophages. We have identified novel
factors involved in vesicular trafficking and protein sorting,
particularly phago-lysosome fusion, during mycobacterium
infection. These proteins are promising candidates for being
involved in the lysosomal-mediated killing of mycobacteria,
as well as in the molecular events linking innate and adaptive
immune responses (Gutierrez et al., 2009; Gutierrez et
al., 2008).
The most promising candidate proteins are rab proteins and
lysosomal receptors. The first ones are the molecular switches
that regulate membrane trafficking and spatio-temporal
organisation of vesicular trafficking (Stenmark, 2009). The
second ones are receptors for some important lysosomal com -
ponents and enzymes in charge of pathogen degradation.
Rab proteins and phagosomes Rab proteins are proteins
that have a key role in organising membranous compartments
facilitating transport and regulating vesicular
trafficking. A complex network of Rab GTPases controls the
dynamic changes of phagosome maturation. We have identified
some Rabs potentially implicated in response to mycobacterial
infection previously not related to infection with
intracellular pathogens (Gutierrez et al, 2008). However, the
exact role of many of these proteins in phagosome maturation
is unknown. We are now defining the function of those
proteins during phagocytosis and evaluating the impact on
the innate immune response to mycobacterial infection.
Lysosomal protein receptors and phagosomes For one
of the identified proteins, sortilin, we showed its role during
phagosome maturation. Up to now, the possibility of a
direct delivery pathway for some lysosomal enzymes from
the Trans Golgi Network (TGN) to the phagosome has been
suggested. However, the pathways involved in this process
and the identity of the enzymes delivered is unknown. Our
findings with sortilin in macrophages identified a novel
and important contribution of this protein to the delivery
of a sub-class of lysosomal proteins from the Golgi to the
phagosome (Wähe et al, 2010). Therefore, our observations
raise the question of the regulation of the sortilin-pathway
functions in the context of phagosomes containing intracellular
pathogens.
Perspectives Although in vitro and in vivo studies have
shed light on some aspects of tuberculosis pathogenesis,
we still do not completely understand how M. tuberculosis
manages to survive in eukaryotic cells and why host cells
are in some cases able to eradicate this pathogen. Therefore,
a deep understanding of the very fundamental biology of
M. tuberculosis that considers the intracellular lifestyle is
crucial for developing new therapies towards tuberculosis
control.
The study of the cell biology and cellular immune response
of macrophages to mycobacteria is a very productive area of
research and its understanding is critical for the understanding
of mycobacterial pathogenesis. These studies will
shed light on fundamental questions of the host to pathogen
response and will also provide knowledge of basic characteristics
of cellular and innate immune functions. With our
studies we expect to contribute to a better understanding
of how macrophages kill mycobacteria and other infectious
intracellular pathogens.
Maximiliano Gabriel Gutierrez born in 1976 in Mendoza,
Argentina, is head of the Junior Research Group Phagosome
Biology at the Helmholtz Centre for Infection Research,
Braunschweig. He studied at the University of San Luis,
Argentina and obtained a biochemistry and molecular biology
degree from the university in 2001. In 2005, he was also
awarded a PhD from the University of San Luis, Argentina.
During his PhD studies he spent some time as a research
visitor in the US and Germany. After that, he spent almost
three years as a postdoctoral scientist at EMBL, Heidelberg,
Germany, as a fellow of the von Humboldt Foundation and
EMBO. Since November 2008 he has been the head of the
Junior Research Group Phagosome Biology. The main area
of research in his group is the cell biology of host-pathogen
interactions with a focus on mycobacteria.

RESEARCH REVIEWS | Mycobacterial Phagosomes and Innate Immunity
45
Literature
Desjardins, M., Celis, J. E., van Meer, G., Dieplinger, H.,
Jahraus, A., Griffiths, G., & Huber, L. A. (1994) Molecular
characterization of phagosomes. The Journal of Biological
Chemistry 269, 32194-32200.
Desjardins, M., & Griffiths, G. (2003) Phagocytosis: latex
leads the way. Current Opinion in Cell Biology 15, 498-503.
Gutierrez, M. G., Gonzalez, A. P., Anes, E., & Griffiths, G.
(2009) Role of lipids in killing mycobacteria by macrophages:
evidence for NF-kappaB-dependent and -independent killing
induced by different lipids. Cellular Microbiology 11, 406-420.
Gutierrez, M. G., Mishra, B. B., Jordao, L., Elliott, E., Anes,
E., & Griffiths, G. (2008) NF-kappaB activation controls
phagolysosome fusion-mediated killing of mycobacteria by
macrophages. Journal of Immunology 181, 2651-2663.
Haas, A. (2007) The phagosome: compartment with a license
to kill. Traffic (Copenhagen, Denmark) 8, 311-330.
Jutras, I., & Desjardins, M. (2005) Phagocytosis: at the crossroads
of innate and adaptive immunity. Annual Review of Cell
and Developmental Biology 21, 511-527.
Korbel, D. S., Schneider, B. E., & Schaible, U. E. (2008) Innate
immunity in tuberculosis: myths and truth. Microbes and
Infection / Institut Pasteur 10, 995-1004.
Russell, D. G. (2001) Mycobacterium tuberculosis: here today,
and here tomorrow. Nature Reviews 2, 569-577.
Stenmark, H. (2009) Rab GTPases as coordinators of vesicle
traffic. Nature Reviews 10, 513-525.
Tauber, A. I. (2003) Metchnikoff and the phagocytosis theory.
Nature Reviews 4, 897-901.
Wähe, A., Kasmapour, B., Schmaderer, C., Liebl, D., Sandhoff,
K., Nykjaer, A., Griffiths, G., & Gutierrez, M. G. (2010)
Golgi-to-phagosome transport of acid sphingomyelinase and
prosaposin is mediated by sortilin. Journal of Cell Science
123, 2502-2511.
Yates, R. M., Hermetter, A., & Russell, D. G. (2005) The kinetics
of phagosome maturation as a function of phagosome/
lysosome fusion and acquisition of hydrolytic activity. Traffic
(Copenhagen, Denmark) 6, 413-420.

FOCUS RESEARCH REVIEWS SPECIAL FEATURES
Photos from left to right: During the official retirement of Dr. Helmut Blöcker. Dr. Siggi Weiss giving some reflections on
the work of Helmut Blöcker at the GBF/HZI | Dr. Kathrin Dinkla, Prof. Dr. G. Singh Chhatwal, Katja Mummenbrauer,
PD Dr. Manfred Rohde, Dr. Susanne Talay (from left to right) during a discussion on streptococci | Prof. Dr. Christiane Ritter
preparing for a new NMR experiment Photos: HZI, Dornbach (le) | HZI, Ammerpohl (ce) | HZI, Gramann (ri)

SCIENTIFIC REPORTS
FACTS AND FIGURES
48 Where Would We Be without DNA Sequences?
52 Nuclear Magnetic Resonance Spectroscopy,
a Major Player in the Arsenal of Platform Technologies
available in the HZI
60 International Cooperation of Helmholtz Centre for
Infection Research with India and China: A Success Story

48 SPECIAL FEATURES | Where Would We Be without DNA Sequences?
Where Would We Be without DNA Sequences?
AUTHOR | Dr. Helmut Blöcker | Department of Genome Analysis | hbl@helmholtz-hzi.de
Three weeks of hard work in the radioactivity laboratory, in the darkroom, in the biology lab and having snacks always impatiently
in between. Then it was done. Just when my colleagues returned from the cafeteria, I had developed the crucial X-ray fi lm, rinsed
it under running water and had a fi rst glance at it. Together we were able to actually read a sequence of 33 nucleotides! A team of
undergraduates, graduate students and postdocs had worked together in the early seventies to produce chemically and enzymatically
the world’s fi rst structural gene coding for a polypeptide. Around three years work of the team, and I was fi rst to hold the laboriously
produced evidence for the existence of the target compound in my hands: 33 bands on X-ray fi lm.
How can it be that today one is able to sequence complete human genomes with billions of DNA building blocks, whereas, within a
single researcher’s life, at that time happiness was in a short sequence of 33 nucleotides?
The development of sequencing technology (1) During
my doctoral work, in the years before the 33-base success,
sequencing procedure of choice was the “wandering spot
method”. Maximum read length was about 13 nucleotides.
Just good enough to check the chemically synthesized
oligonucleotides for the construction of the said 33mer
duplex. After a working visit at MIT, Hans Gross had the
method established at the MPI for Biochemistry in Martinsried/München
for RNA analysis. Together we tested it there
successfully for DNA sequences.
To check longer sequences, the development of base-specific
chemical cleavage of DNA, the method of Maxam and
Gilbert, proved advantageous. But how to get the details of
the tricky and not yet published method? Without the
existence of the internet, e-mail, pdf attachment, or even a
fax machine? Six weeks later, an airmail letter to Allan
Maxam at MIT brought permission to take a copy of a still
secret protocol of Heinz Schaller’s at DKFZ. So that’s the
33-base success.
Even a hard-working chemist does not like to work with
toxic and volatile substances such as in the Maxam/ Gilbert
protocol. It is, therefore, not surprising that this method
was soon replaced completely by the dideoxy method of
Sanger (“chain termination synthesis”). Soon you could
achieve read lengths of several hundred nucleotides and
analyse together several tens of samples over slab gel
electrophoresis devices. We had about a dozen of such
devices, so we had established a room especially for the
preparation of electrophoresis cassettes (pouring gels, clean
glass plates, etc.) including a special dishwasher.
This drudgery was over when commercial sequencers were
available, in which the separation of sequencing reaction
mixtures were no longer done next to each other on slab
gels, but performed on capillaries filled with separating
material. In addition, the capillaries were loaded automatically
with samples from microtiter plates. This generation
of devices formed the technical basis for the first complete
sequence of the human genome as it was developed by the
international public human genome consortium including
our department.
The sequencers and the procedures around them were so
sophisticated that one could quite get read lengths of 1,300
nucleotides per separation capillary at up to 384 capillaries
per device. For aesthetes, the gray boxes were not a
revelation - that has not changed until today. Our many
visitors were impressed at the most with the cost (several
hundred thousand Euros per unit). At the beginning of this
decade hardly any of the laboratory staff could imagine that
the entire Sanger-based technology would become hopelessly
inferior in a few years and would be ready for the
museum.
Already at the heyday of the Sanger technology experts
were informed at congresses and in particular from
international review documents that soon a whole new
generation of devices and procedures would be on the
market (“next generation sequencing”). Today, the three
main types of techniques and equipment of this generation
are marketed by the companies Roche, Life Technologies
and Illumina. In 2007, we opted for the Illumina devices
that are now dominant. It is surprising that within about
two years the sequencing productivity of our equipment
could be raised to about 1,500 percent through comprehensive
optimization. Today one can generate on a single
sequencer, the raw sequence of a complete human genome,
at a moderate cost of several thousand Euros. Remember
that the first estimates for the cost of the public Human
Genome Project was around two billion U.S. dollars!

SPECIAL FEATURES | Where Would We Be without DNA Sequences?
49
Fig. 1. Automated cluster analysis from Illumina sequencer.
Within about two years overall sequencing productivity could
be raised to about 1,500 percent through comprehensive
optimization. Right-hand side: blow-ups. Photo: HZI
An amazing journey from “handmade” 33 base pairs to the
machines of today. However, no deep knowledge is necessary
to imagine that this extremely rapid development will
continue. You can find a graphic explanation of general
lines of technology development in Clayton Christensen’s
book “The Innovator’s Dilemma” (2).
Fig. 2. Clayton Christensen. How low-end disruption occurs over
time. (http://en.wikipedia.org/wiki/Disruptive_technology)
In fact, there are announcements of the first devices that
are smaller, highly integrated, very much lower in acquisition
cost and can operate at extremely low cost. In future, it
will be possible to routinely sequence single molecules. The
DNA material will be used free of artificial markers, and
read lengths of tens of thousands of nucleotides are only a
few years (if not months) ahead. A focus will be on developments
in nanotechnology and microfluidics. The first
sequencers for around 50 kEuros are on the market. A
British company announced a disposable sequencer the size
of a USB stick they want to develop in the coming years.
Simply breathtaking!
No wonder that with such a price perspective, DNA
sequencing will soon find its way into daily routine
analysis, in hospitals, the pharmaceutical industry, food
industry, breeding research, and health and food inspection
agencies, and perhaps on farms and in doctors’ offices.
Compared to the period before the present devices, almost
nothing remained as it used to be. This is true for efficiency
and also the vastly greater versatility of new devices and
procedures developed for them. It is no longer merely about
the control of cloning reaction or de novo sequences of
genomes, but also, for example, the analysis of transcripts,
metagenomics, genome-wide methylation patterns, gene
duplications, and SNPs. It is this methodical expansion of
sequencing technology, which not only cannibalizes
existing chip technology and reduces it to pure niche
technology, but makes sequencers become an indispensable
part of any molecular biology laboratory for a very long
time. In future, a PS (“Personal Sequencer”) in the laboratory
will have come up to standard. Just like a PCR machine.
It would be an undue exaggeration to argue that the current
and next sequencers could be operated by well-trained apes.
But at least the next devices will demand less from the
technical personnel and will nevertheless produce almost
unimaginable amounts of data. We should strongly and
immediately adjust to the situation that we need intelligent
bioinformaticians and experts with a broad molecularbiological
and medical knowledge that can deal with mass
data. Despite enthusiasm for the rapid development of pure
data generation, this has been our permanent reminder
– not least in my HZI internal paper of 2006 on an assessment
around sequencing (“Some Remarks Relating to Next
Generation Genetic Analyzers”).

50 SPECIAL FEATURES | Where Would We Be without DNA Sequences?
Excursus funding policy Without any massive public and
private investment, the current international genome-based
research would simply not exist. After our interest in
genome research aroused in the mid-1980s, it was very
clear that substantial progress could only be reached with
strong funding. My paper “Technology Development and
Genomics” became the basis of a funding programme of the
German Ministry of Research. It was run over a number of
years and mobilized a total of about 150 million DM for the
scientific landscape in Germany. Part of the logic behind
the initial funding period and the following ones today
appears, however, questionable. The logic goes something
like this: first, we develop technology, we generate data, we
interpret the data, we develop new approaches for diagnosis
and new therapies, and finally, we market the final results.
Nice thought, but unrealistic. To date, most top countries act
differently. The best examples are the USA and China.
There is continuous, substantial funding for each link in
the chain, from technology development to the medical use.
Specifically, the technology sector in the United States may
have made billions of revenue. Unfortunately, early and
pioneering developments in Germany did not encounter
stable funders in this country. In a modification of the title
of this paper we might ask: Where could we be in Germany
if more DNA sequences had been made?!
From a nucleic acid chemist to a sequencing guy
After completing my graduate work at the former GMBF in
Braunschweig I joined a young and dynamic team working
on gene synthesis in Hamburg. It was mainly concerned
with the aforementioned DNA double strand of 33 bases in
length, which carries the genetic information for the
peptide hormone angiotensin II (3).
In 1980, with all our experience and developments from
Hamburg University in mind, Ronald Frank and I together
set up a new research group at the GBF. Within three years,
we had reached our five-year goal. We synthesized oligonucleotides
and then produced longer genes by chemical-enzymatical
procedures. Having produced DNA double strands,
our curiosity was aroused for optimal expression systems.
Expressed synthetic genes made us dream of artificial
proteins. Working with the genes of protease inhibitors and
genes of other rather small proteins, however, taught us that
even in these protein families there was too little information
available about the relationship of specific gene variants to
the respective biologically active proteins. It seemed
reasonable to work towards an acceptable algorithm. This
could be attempted at that time only via mass DNA sequencing
since it was (and still is) superior to peptide sequencing.
With the above chapter on the development of sequencing it
is clear that one could only compete on an international top
level, if one would accept the challenge of technological
innovation. We tried this by developing the “Shortmer”
sequencing primer technology, by ongoing developments in
robotics, bioinformatics and biochemistry that still give us,
as in the work of Gerhard Kauer (4) and Igor Deyneko, a
competitive advantage. As a consequence, since the
beginning of the nineties the various activities around
sequencing were the focus of the GBF/HZI group later
known as the Department of Genome Analysis. Fortunately,
we had an early chance to prove ourselves in international
sequencing projects such as those supported by the EU - the
yeast and Arabidopsis genome projects.
The ultimate project was certainly the public Human
Genome Programme (5), where we took part as one of 16
genome centres worldwide. Often referred to as a moon
landing project of bio-research this project caused a global
echo in the media and society like no other in the biological
sciences. A few years ago there were rumours from multiple
sources, that our international consortium was proposed as
a candidate for a Nobel Prize. Well, first a rumour is a
rumour, and secondly, Helmut Kohl has been rumoured to
be proposed several times for a Nobel Prize – he has not
been awarded the prize to this day. I found the Nobel
rumour interesting. I was in the USA when I learnt from a
British official about the Nobel story. My immediate
reaction was: “Brilliant! I would love to become 0.2 percent
of a Nobel laureate!” After all, a few hundred scientists were
involved in the project! However, unlike in physics, there
are rules for the life sciences that the prize may be awarded
only to a maximum of three people.
One may view genomics with its core element sequence
analysis as drudgery for research, development and
application in the life sciences and medicine. For these
areas it will be indispensable. As early as 2000 the journal
Science entered the sequencing of whole genomes in their
annals of interdisciplinary breakthroughs of the year (6).
Francis Collins, an advisor to President Obama, director of
NIH, and previously speaker of our Human Genome
Programme has recently put it this way: “Genome research
is really just beginning and it can be dizzying to live on an
exponential curve”. So it was and it is.

SPECIAL FEATURES | Where Would We Be without DNA Sequences?
51
Back to the headline Where would we be without DNA
sequences? This is what I ask in the title of this essay. The
simplest answer would be (correct, but a discussion killer):
without DNA sequences no life form would exist, including
ourselves. But research and development have enabled us to
undertake all kinds of travel in the universe of our interior
by working with nucleic acids sequences. As a fan of space
exploration I am interested in this inner universe much
more than in the universe around us. Perhaps a self-centered
thought, but certainly closer to discoveries that give
us more knowledge about ourselves and permits us to make
life a little better - if used responsibly. This is what I
secretly hope for myself. Against the background of
thousands of human genomes, which are currently being
analysed in the world, my own genome sequence should tell
some interesting secrets. I also expect this for the genome
sequences of my three colleagues in Braunschweig, whose
genomes are being analysed together with my sequence.
Profound knowledge of DNA sequences is the key to
knowing more about ourselves and our living environment,
today and tomorrow. As a nucleic acid person, what else can
you expect from your professional life?
Literature
(1) for various sequencing details look here:
http://en.wikipedia.org/wiki/DNA_sequencing#cite_note-6
(2) Christensen, Clayton M. (1997). The innovator‘s dilemma:
when new technologies cause great firms to fail. Harvard
Business Press. ISBN 9780875845852.
(3) Köster, H., Blöcker, H., & Frank R., Geussenhainer S.,
& Kaiser W. (1975) Total Synthesis of a Structural Gene for
the Human Peptide Hormone Angiotensin II. Hoppe-Seyler´s
Zeitschrift für physiologische Chemie 356 (2), 1585–1594.
(4) Kauer, G. & Blöcker, H. (2003) Applying signal theory to
the analysis of biomolecules. Bioinformatics 19, 2016-2021.
(5) International Human Genome Sequencing Consortium
(2001) Initial sequencing and analysis of the human genome.
Nature 409, 860-921.
(6) Pennisi, E. (2000) Breakthrough of the year. Genomics
Comes of Age. Science 290, 2220–2221.
Fig. 3. Four genomes under investigation: Michael Jarek,
Tschong-Hun Im, Maren Scharfe, Helmut Blöcker, all next to
hand-crafted DNA (farewell present to H.B.) Photo: HZI
Helmut Blöcker born in 1945, obtained his diploma in
chemistry at the Technical University of Braunschweig
(1972). He did his doctoral thesis (Dr. rer. nat.) at the University
of Hamburg (1972-1975) and worked for 5 years as
a postdoctoral scientist in the same group. In 1980 he set
up the DNA Synthesis Group at the GBF. He was head of the
department of Genome Analysis at the GBF/HZI (1994-2010).

52 SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
Nuclear Magnetic Resonance Spectroscopy,
a Major Player in the Arsenal of Platform Technologies
available in the HZI
AUTHOR | Dr. Victor Wray | Research Group Biophysical Analysis | vwr@helmholtz-hzi.de
The analytical instruments platform is the oldest core facility in the HZI with well established modern instrumental techniques and
expertise for structural investigations of all types of biomolecule. It provides detailed structural data in numerous forms that are
crucial for understanding the underlying molecular mechanisms and interactions governing the cellular processes involved in hostpathogen
interactions.
The platform provides large instrumental facilities for determining the three dimensional structure of all types of natural product
and fi nds application in most, if not all, of the main areas of research within the “Infection and Immunity” programme of the HZI.
Recent reports in this series have featured the use of electron microscopy and X-ray crystallography. Electron microscopy can
visualise a wide range of super macromolecules, organelles and bacteria, as well as providing intimate details of the adherence to,
and invasion of, host cells by a wide range of pathogens. The complementary technique of X-ray crystallography provides structural
details at the atomic level of proteins and their molecular complexes with ligands and other proteins. It has provided considerable
detail of both the mechanisms of bacterial adhesion, invasion and propagation, as well as human receptor signalling. Here we will
consider, in some detail, the use of nuclear magnetic resonance spectroscopy for studies of small molecular weight natural products
in solution. It plays a major role in the discovery and development of novel anti-infectives, as well as affording complementary
information to that gained by X-ray crystallography.
Introduction Early in the development of the institute it
was recognised that instrumentation capable of investigating
biological structures was necessary for an understanding
of basic natural processes. Enormous advances have
been witnessed over the last four decades in various instrumental
areas with the rapid development of fundamental
physical techniques and the integration of the potential of
computer technologies. Of particular importance for structural
biology has been the rapid evolution of nuclear magnetic
resonance (NMR) spectroscopy, mass spectrometry
(MS), X-ray crystallography (X-ray) and electron microscopy
(EM). These techniques are grouped together as one of the
core facilities in the HZI and have the ability of providing
intricate details of the whole spectrum of biological
structures from the smallest of natural products (NMR and
MS), through macromolecules and their complexes (X-ray)
to the largest of structures that include organelles and bacteria
(EM) (Fig. 1). In a previous report 2008/9 the elegant
application of X-ray crystallography was shown to yield
intimate details of microbial pathogen interactions with
the host organism at atomic resolution. Such details allow
the development of new strategies and tools to broaden our
understanding of bacterial infections. Here we will concentrate
on the complementary technique of NMR spectroscopy
and its application to the investigation of biologically active
natural products.
The power of the nucleus In the late 1940s it was shown
that the nucleus of certain atoms absorb electromagnetic
radiation in a magnetic field and the frequency of the
radiation was dependent on the strength of the magnetic
field, the nucleus involved and, very importantly, on the
particular chemical environment of that nucleus. Of the
common atoms only 1H, 13C, 15N and 31P showed this
phenomenon and clearly had great potential for delving
into the structures of organic compounds, and hence most
natural products. Subsequently, the development of powerful,
stable magnetic fields led to the high resolution of the
signals arising from the same nuclei in different chemical
environments. In tandem, a decisive step forward was the
introduction of pulsed NMR spectroscopy, the development
of which was associated with the rapid advancements in
computer technology, allowing enormous gains in sensitivity.
However, the development of two-dimensional (2D)
NMR methodology in the 1970s, particularly by R. R. Ernst,
for which he received a Nobel Prize in 1991, heralded the
start of a new era in NMR spectroscopy. The use of these
techniques now allowed the ready identification of bonding
patterns in molecules via either the through-bond interactions
of similar nuclei (homonuclear 2D NMR) or dissimilar
nuclei (heteronuclear 2D NMR), as well as conformational,
sequential and folding properties of units in a molecule by
the observation of through-space interactions.

SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
53
Small Natural Products
Macromolecules:
Proteins
NMR Spectroscopy
Molecular Complexes:
Protein-ligand
Protein-Protein
Mass Spectrometry
X-ray Crystallography
Super Macromolecules:
Organelles
Bacteria
Electron Microscopy
Fig. 1. Analytical Instruments Platform Photos: HZI
Since the early 1970s NMR instrumentation in our institute
has closely followed these technical breakthroughs as it
has become the most powerful technique for the routine
structure elucidation at the atomic level of biologically
active natural products in solution. For pharmacologically
relevant substances with molecular weights under 2000 Da
it has the advantage over X-ray of not requiring crystals of
the material and allowing rapid access to structural details.
As a consequence, over the years NMR spectroscopy has
been used to solve the structures of an extremely large
variety of natural products emanating from a diverse
number of research groups both within the HZI and from
external sources.
The quest for novel anti-infectives: Structures of
biologically active substances The emergence of new and
reoccurring pathogens continues to pose a serious threat to
human health even in developed countries. As a consequence
the search for, identification, characterisation and
clinical development of novel anti-infectives from natural
sources is, and will continue to be, a major goal of the
Infection and Immunity programme of the HZI. The
structure of such compounds is a pre-requisite for their
development and is most easily achieved by a combination
of NMR spectroscopy and mass spectrometry.
In contrast to chemical synthesis there is generally no prior
knowledge of the structure of a compound isolated from
plants, bacteria or fungi, all potential sources of new active
compounds. Consequently, after isolation, a small amount of
material is dissolved in an organic solvent and a simple
proton ( 1 H) spectrum is recorded (Fig. 2). Such a spectrum
already contains a wealth of information regarding the
nature of units present in the molecule. The position of
signals in the spectrum reflects the local environment of
the observed proton, allowing distinction of aromatic, sugar
and aliphatic groups. While the signal integral indicates the
number of protons present, their splittings provide details
of the number and disposition of protons in the immediate
vicinity. Additional information is afforded by a carbon ( 13 C)
spectrum which again shows a dispersion of signals that
allow the number and types of carbon atom to be readily
determined.

54 SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
Aromatic region Sugar region Aliphatic region
1
H spectrum
2D NMR experiments Yield
molecular fragments
13
C spectrum
Fig. 2. 1D NMR spectra of an unknown compound in methanol-d4 Graphic: HZI
A
NMR Data
B
Me
HO
MeO
OR
O
OH
C
MeO
HO
HO
MeO
O
Me
HO
MeO
O
O
OH
OMe
Fig. 3. Analytical procedure (A), 2D 1 H- 1 H spectrum showing sugar unit (B) and complete structure (C) Graphic: HZI

SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
55
The use of 2D NMR spectroscopy now allows the definition
of fragments in the molecule (pieces of the jigsaw, Fig. 3).
Thus homonuclear 2D 1H-1H spectra shows off-diagonal
signals corresponding to through-bond interactions
between hydrogen atoms two and three bonds apart. The
identification of such interactions allows the tracing of
series of these interactions that identify the various
fragments in the molecule. Further use of heteronuclear 2D
1
H- 13 C spectra confirms the types of fragment present and
allows the pieces of the jigsaw to be pieced together to give
a final three-dimensional structure. The exact conformational
details of this structure are then available from
inspection of the magnitude of the splittings in the 1D 1 H
spectrum and from through-space interactions of the
protons in the different fragments. Finally, confirmatory
data regarding the molecular weight is available from mass
spectrometry and other data from ultra-violet and infra-red
spectroscopies, as well as chemical information.
This approach has been widely applied over the last three
decades to elucidate the structure of numerous natural
products and it continues to be the method of choice. The
improvements in instrumentation particularly in terms of
sensitivity and signal resolution provided by modern
superconducting magnet systems now allows small
amounts of material to be investigated (0.2 to 1 mg) without
the need for chemical derivation. As is evident from
previous features in these reports, NMR structure elucidation
has played a major role in the discovery and evaluation
of new anti-infectives, particularly in unravelling the
structures of the complex number of biologically active
compounds produced by the myxobacteria. This is well
illustrated by the large number of basic structures of
secondary metabolites from the single bacterium Sorangium
cellulosum, Fig. 4. To date over 600 unique myxobacterial
metabolites have been identified and their structures
elucidated. Over the years a very promising number of
small molecules with anti-tumour, antiviral (proteasome
inhibitors), anti-infective (interferon regulators) and
anti-bacterial (antibiotics, cytolysin inhibitors, biofilm
inhibitors) activities have been discovered and characterised.
The most important of these was the discovery of
epothilone, a compound that has led to a treatment of
aggressive metastatic or locally advanced breast cancer.
NMR also provides the main means of establishing the
structure of synthetic derivatives of natural products
prepared for structure-activity studies, a pre-requisite for
drug development, as well as a priori chemical syntheses for
groups active in chemical biology and medicinal chemistry.
OH
OH
O O O OMe O
OH
MeO
OH
O OH OH
O
HO
HO
N
S
OH
O
OH OH
O
O
S
N
OH
O
O
OH
O
O
COOH
N
H
N
O
O
OMe
O
N
MeO OH
O
O
O
O
O
O
MeO
O
HO
O
OH HO
O
O
OH OMe
O
OMe
HO
S
N
H
O
O
H
OH
OH
O
HO
O
MeO
OH
HO
O
O
O
O
OH O
OH
OH
H
O
OH
O
O
H
N
N
O
O
HO
O
O
OH
O
O
OH
O
HO
O
HN
O
N
Me
O
O
N
H
O O
NH
OH
HN
HO
NH
H
N
O
OH
HN
O
OH
O
HO
O
HO
H
H H
Cl H
O O
Cl
O O
O
NH
OH
OH
O O
O
HO
HO
O
O
Cl
O
O
N
OH
OH
OH
COOH
COOH
OH
O
HO
H
O
N
O
H
OMe
O
O
OH
O
O
O
H
N
O
HOOC
OH
O
O
N
O
O
O
OH
N
O
OMe
O
OH
O
OH
OH
O
O
O
O
O -
O B
O
Na +
O
OH
O
O
OH O
O
O
O
O O
O
HO
O
OH
OMe
OH
O
O
OH
OH
OH
OH OH OH
OH
S
Me
N
O
OH
O
O OH O
O
OH
OH
HO
O
OH
OH
O
O
O
OMe
O
O
N
O
OH
HO
O
OH
OMe
O
H
N
H
OH
OH
O
HO
OH
OH
OMe
O
O
O
O
OMe
O
OH
OH
OMe
OH
COOH
MeO
MeO
HO
O
OMe
O
O
O
OH
MeO
H OMe
N
O
HO
COOH
HO
O
OH
NH2
O
H H
OH O
O
H
N
O
N
H
N
O
S
OH
HOOC
O
OH
OH
O
H3C CH 3
H 3CO
CH3
O
H3C
OH O
OH
O
O
OH
O
O
O
O
HO
O
OMe
OMe
Me
OMe Me
O O
Cl
O
OH
OH
Cl
O
O
NH
O
N
H
H
N
O
S
N
O
N N
N N
MeS O
HO OH
Fig. 4. Secondary metabolites from the myxobacterium Sorangium cellulosum Graphic: HZI, Gerhard Höfle

56
SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
In the past significant collaborations with universities and
other institutes have also been forged that have involved
investigations of natural products from bacteria, fungi,
plants and marine sources.
NH
O
HO
OH
O
O
O
OH
O
O
OH
N
H
Fig. 5. Biosynthetic precursors of apicularen B Graphic: HZI
O
Apicularen B
1,2- 13 C-acetate
1- 13 C-glycine
13
C-methyl-methionine
Biosynthetic pathways and monitoring of biological
systems Information of the biosynthesis of secondary
metabolites can also be furnished by monitoring the
products after “feeding” the producer bacterium with
13C-enriched precursor molecules. Under these conditions
the precursors are incorporated into the product that shows
enhanced signals in the 13 C NMR spectrum only for those
carbons derived from the enriched material. An example is
shown in Fig. 5 for apicularen B biosynthesised from
acetate, glycine and methionine.
The use of NMR spectroscopy for monitoring changes in
biological systems was recognised very early and was used
to investigate non-invasively the cytoplasm-vacuole
differential acidity and phosphate metabolites in plant cell
suspension cultures of Nicotiana tabacum and potato by
in vivo 31 P NMR spectroscopy. More recently the pathways
involved in the biodegradation of substituted aromatics
have been followed by adding specific enzymes from
particular bacteria in situ to key intermediates and
identifying the resulting compounds by 1H NMR spectroscopy.
This provides direct information for the reconstruction
of the metabolic networks found in aromatic degradation
and permits a rational exploitation of the capabilities of
the microorganisms involved.
FROM THE GENOME TO FUNCTIONAL GENOMICS
Functional genomics
Genome
Proteome
Cellular Function & Regulation
1D Sructure
Proteomics
2D gel elecrophoresis
MALDI / ESI MS
Protein sequencing
Post-translational
modification
Protein Production
Recombinant
Methods
Cloning
Expression
Protein Fermentation
Solid State
peptide
synthesis
Protein 3D Structure
Structural Methods
X-ray Crystallography
NMR spectroscopy
Flexible non-crystalline
proteines
Fig. 6. Instrumental techniques used in functional genomics Graphic: HZI

SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
57
Structural biology: Application of NMR in protein studies
The impetus to protein research generated by the decoding
of the both human and subsequent genomes at the beginning
of the century has been a result of the enormous
progress made in many areas of instrumentation. This was
reflected in the 2002 Nobel prizes in chemistry awarded to
Fenn and Tanaka for the development of mass spectrometric
analyses of biological macromolecules and to Wüthrich for
his development of NMR spectroscopy for determining the
three-dimensional structure of biological macromolecules
in solution. Soft desorption ionisation MS methods (ESI and
MALDI MS, figure 6) provide the one-dimensional sequence
of amino acids of proteins and indentify post-translational
modifications. This information is a prerequisite for the
determination of their three-dimensional structures and
subsequent correlations with cellular function and
regulation (figure 6). The importance of X-ray crystallography
for structural investigations of crystalline proteins has
been stressed in an earlier feature of these reports. Here an
alternative approach is considered that exemplifies the use
of NMR spectroscopy to elucidate the solution structure of
small flexible proteins, molecules that do not form crystals
and are not readily produced by recombinant methods.
A
B
Ion channel
activity
Interaction
with CD4
The HIV genome encodes a number of enzymes and
structural proteins as well as six accessory proteins. Of
these the HIV-1 specific virus protein U (Vpu) is an 81
amino acid class I integral membrane phosphoprotein that
induces degradation of the virus receptor CD4 in the
endoplasmic reticulum and enhances the release of virus
particles from infected cells. Although this flexible protein
does not form crystals, sufficient quantities of the protein
and related fragments were available from solid state
peptide synthesis that allowed studies to be undertaken by
circular dichroism and NMR spectroscopy. Both methods
indicated that the structure of the molecule was dependent
on the solution conditions and only in a hydrophobic
membrane-like environment did the molecule assume a
stable secondary structure, a phenomenon characteristic of
many small proteins in solution. Detailed 2D 1 H NMR
studies of the 50 amino acid cytoplasmic C-terminal domain
in its most structured state possessed a well defined
helix-interconnection-helix-turn structure in which the two
serine residues undergoing post-translational phosphorylation
were positioned in the interconnection between the
helices. The position of these structures and the helix of the
N-terminal anchor domain in a bilayer membrane-like
environment were established in oriented phospholipid
Fig. 7. Model of the membrane-bound structure of monomeric
Vpu (A) and its dynamic forms (B) Graphic: HZI
bilayers using 15 N solid-state NMR spectroscopy. These data
allowed a model of the monomeric structure of the molecule
to be constructed, figure 7, in which the first helix (α1) of
the N-terminal domain forms a trans-membrane helix
attached to the first helix (α2) of the cytoplasmic domain
which binds parallel to the membrane surface. This is then
attached to the final helix (α3), which shows only weak
interactions with the membrane. Such a structure is
compatible with the ion channel activity of the N-terminus
and the interaction of the C-terminus with the cytoplasmic
domain of CD4, as well as the requirement of a phosphorylated
Vpu for this function. Similar studies provided
details of the structure of a second HIV-1 accessory protein,
Vpr, and more recently indicated its unusual interaction
with a host protein, cyclophilin A.

58 SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
Large proteins and the compatibility with X-ray
The question now arises as to whether NMR spectroscopy
can also provide structural details for larger proteins and
are the structures determined in solution compatible with
those in the solid phase from X-ray crystallography. In
general, the answer to both questions is yes. Laboratories
such as the HZI Protein Sample Production Facility can now
produce 15 N, 13 C-doubly-labelled protein using recombinant
methods suitable for multi-dimensional heteronuclear NMR
investigations in solution. A salient example is the elucidation
of tryparedoxin, a virulence determinant in trypanosomatids,
the causative agent of trypanosmiasis (sleeping
sickness and Chagas disease) and leishmaniasis. The global
folding of the structure determined by three-dimensional
NMR spectroscopy was very similar to that of the crystal
structure although regions near the active site were less
well defined in solution (figure 8, A and B). Clearly, if the
structure was the only aim of the work, X-ray crystallography
would have been the method of choice. However, NMR
allowed the binding of inhibitory substrate analogues to be
readily investigated and disclosed regions on the protein
surface that are functionally relevant (figure 8, C and D),
which set a framework for the design of more rigid and
active ligands.
Fig. 8. Comparison of NMR and X-ray structures of tryparedoxin (A & B), surface model showing ligand-binding positions (C & D)

SPECIAL FEATURES | Nuclear Magnetic Resonance Spectroscopy, a Major Player in the Arsenal of Platform Technologies
59
Future perspectives There will be a continued intensive
use of the platform technologies in the foreseeable future
that is underscored by recent major investments in new
instrumentation. These will play a significant role in the
research undertaken in the new Drug Discovery Centre that
is currently in the planning stage. All aspects of structure
elucidation of old and new natural products of all types will
continue to be a key area in infection research as envisaged
in the current research initiative of the HZI. The continued
advances in technology, particularly the imminent availability
of ultra-high magnetic fields (≥ 1 GHz), opens up the
possibility of studying even larger systems such as
biomacromolecular assemblies of highest complexity and
biomedical significance by solution NMR techniques that
will complement those studied by X-ray. Clearly this is a
further important area of research that will make significant
contributions to modern biology and biomedicine by
increasing our knowledge of the structures of macromolecules
and their interactions within the cellular context.
Literature
1. Frank, R. (2007) The chemical pipeline – A research programme
and infra-structure for the discovery and evaluation
of new anti-infectives. Research Report 2006-2007, 32-43.
2. Reichenbach, H. (2007) Natural products: An indispensible
source of new drugs. Research Report 2006-2007, 54-59.
3. Wray, V., Schiel O., Berlin, J. & Witte, L. (1985) Phosphorus-31
nuclear magnetic resonance investigation of the in
vivo regulation of intracellular pH in cell suspension
culture of Nicotiana tabacum: The effect of oxygen supply,
nitrogen, and external pH change. Archive of Biochemistry
and Biophysics 236, 731-740.
4. Pieper, D. H., Pollmann, K., Nikodem, P. Gonzalez, B. &
Wray, V. (2002) Monitoring key reactions in degradation of
chloroaromatics by in situ 1 H nuclear magnetic resonance:
Solution structures of metabolites formed from cis-dienelactone,
Journal of Bacteriology. 184, 1466-1470.
5. Wray, V., Kinder, R., Federau, T., Henklein, P., Bechinger,
B. & Schubert, U. (1999) Solution structure and orientation
of the transmembrane anchor domain of the HIV-1-encoded
virus protein U by high-resolution and solid-state NMR spectroscopy.
Biochemistry 38, 5272-5282.
6. Krumme, D., Budde, H., Hecht, H-J., Mange, U., Ohlenschläger,
O. Ross, A., Wissing, J., Wray, V. & Flohé, L. (2003) NMR
studies of the interaction of tryparedoxin with redox-inactive
substrate homologues. Biochemistry 42, 14720-14728.
Victor Wray born in 1945, obtained his B.Sc. (1966) and
Ph.D. (1969) in Chemistry at the University of Hull. Postdoctoral
scientist in the Department of Organic Chemistry at
Imperial College, London (1969-1973). Joined the Physical
Measurements Group of the Centre for Molecular Biological
Research, a forerunner of the GBF and HZI (1973). Head of
the NMR spectroscopy group since 1987 and head of the Biophysical
Analytics Research Group (including the Analytical
Instruments Platform) since 2003.
7. Solbak, S. M. Ø., Reksten, T. R., Wray, V., Bruns, K., Horvli,
O., Raae, A. J., Henklein, P., Henklein, P., Röder, R., Mitzner,
D., Schubert, U., Fossen, T. (2010) The intriguing cyclophilin
A-HIV-1 Vpr interaction: Prolyl cis/trans isomerisation catalysis
and specific binding. BMC Structural Biology 10, 31.

60 SPECIAL FEATURES | International Cooperation of Helmholtz Centre for Infection Research with India and China
International Cooperation of Helmholtz Centre for
Infection Research with India and China: A Success Story
AUTHOR | Prof. Dr. G. Singh Chhatwal | Department of Medical Microbiology | gsc@helmholtz-hzi.de
Infectious diseases, which are responsible for a quarter of all deaths, are a serious global health problem. One reason is that the
infectious agents do not require passport, visa or fl ight ticket to travel from one country to another. A few years ago it took the
SARS virus just 24 hours to travel from Hong Kong to Toronto. To add to this mobility problem, pathogenic microorganisms differ
considerably in their antigen repertoire in different geographical regions. It is well known that strains belonging to the same
species have different types in China, India, Europe or North America. For an effective control of infectious diseases a global effort
is required, not only to predict and manage the epidemics, but also to design and develop region-specifi c intervention strategies.
Helmholtz Association, therefore, aims to strengthen international cooperation in the area of health research. Motivated by this
initiative, HZI has strengthened its international cooperation in the last few years, illustrated by the following examples.
Indo-German Science Centre for Infectious Diseases (IG-
SCID): a role model for international cooperation
In 2006 HZI, Medical University Hannover (MHH), and Indian
Council of Medical Research (ICMR) presented a vision
to establish a Virtual Centre for Indo-German cooperation
in the area of infectious diseases, especially the development
of new diagnostics, vaccines and antiinfectives. The
rationale behind this vision was strong clinical research in
the field in India and state-of-the-art modern technologies
and experimental animal facilities at HZI and MHH. This
high degree of complementarity made the concept into a winwin
situation. This vision became reality in 2006, when Prof.
N.K. Ganguly, Director General of ICMR, and Prof. Jürgen
Mlynek, President of Helmholtz Association, signed a Memorandum
of Understanding in the presence of the Prime Minister
of India, Dr. Manmohan Singh, and the Chancellor of
Germany, Dr. Angela Merkel, in Hannover. After this signing
the events took place fast and both Helmholtz Association
and ICMR ensured the financing of the centre. In April 2007
the Centre was inaugurated in New Delhi in the presence of
dignitaries from Helmholtz Association, HZI, MHH, ICMR
and a number of scientists working in the area of infectious
diseases. An office of IG-SCID was set up at the headquarters
of ICMR in New Delhi. Six months after inauguration IG-
SCID was expanded to include Jawaharlal Nehru University
(JNU) and a Memorandum of Understanding in this regard
was signed by Dr. Rajendra Prasad, Rector JNU, Dr. N.K. Ganguly,
and Prof. Rudi Balling (Director HZI) in the presence of
the Honorable Ministers Dr. Annette Schavan and Mr. Kapil
Sibal in New Delhi.
A centre with such elite partners was deemed to be a
success, but it has now become clear that this Centre has
become a role model for international cooperation. A unique
concept of this Centre has contributed towards this achievement.
The Centre funds joint research projects based on
excellent science and high relevance for both countries. The
projects are peer-reviewed by a joint steering committee
in a rapid review process. Besides funding the projects,
the Centre sponsors joint workshops on particular areas of
infectious diseases in India and Germany. The Centre puts
great emphasis on the training of young investigators from
both countries. German investigators who travelled to India
benefited from on-hand experience with clinical infection
research as well as epidemiological surveys and Indian
investigators gained expertise in Braunschweig and Hannover
in modern technologies and new animal experimental
techniques. The Centre recently started a new fellowship
programme where scientists from India and Germany can
undertake exchange visits and do research for a period of
up to 3 months. Since both German partners come from
Niedersachsen, its activities were presented during the
visit of the Honourable Chief Minister, Mr. David McAllister,
in New Delhi in September 2010. The Centre is a useful
addition to the joint activities of Niedersachsen with India
in science and technology.
There is a desire from both sides to continue this cooperation
and to look for possibilities to transform this Virtual
Centre into a physical one, most probably in India.
Fig. 1. Signing of MoU by Prof. Ganguly (left) and Prof. Mlynek
in April 2006 in Hannover Photo: HZI

SPECIAL FEATURES | International Cooperation of Helmholtz Centre for Infection Research with India and China
61
Fig. 2. Inauguration of IG-SCID in May 2007 in New Delhi
(from left: Prof. Balling, Prof. Chhatwal, Prof. Mlynek,
Prof. Ganguly and Prof. Hasnain) Photo: ICMR
Fig. 5. Sino-German meeting in Guangzhou, China coordinated
by Prof. Chhatwal from German side and Prof. George
Fu Gao, Vice President of Beijing Institutes of Life Sciences,
Chinese Academy of Sciences, from Chinese side in December
2010. Prof. Han Jianguo, Director of the Sino-German Center,
also participated in this meeting. Photo: HZI
Fig. 3. Expansion of IG-SCID to include Jawaharlal-Nehru University
as a partner during the visit of Dr. Annette Schavan,
German Federal Minister of Education and Research, in New
Delhi in October 2007 Photo: HZI
Cooperation with China: an important initiative
Infectious diseases represent a huge health hazard in China.
In the last decade the research activity in this field in China
has substantially increased. The results flowing in from
this research indicate that the strains in China differ not
only in their virulence repertoire but also cause different
clinical manifestations. To develop global control strategies
against infectious diseases, a strong cooperation with
China is extremely important. Therefore, HZI has started an
initiative with Chinese Academy of Sciences for intensive
collaboration. This initiative has resulted into two Sino-
German workshops in Goslar in 2009 and in Guangzhou in
2010. These activities were fully sponsored by the Chinese-
German Centre for Science Promotion in Beijing. This
centre clearly identified the need for such cooperation. From
German side, beside HZI, Justus Liebig University Giessen,
RWTH Aachen, TiHo Hannover and MHH participated in
workshops. From Chinese side, different institutes of Chinese
Academy of Sciences were partners. These workshops
resulted in a number of bilateral cooperations, and an application
for a bigger Sino-German project is being submitted.
We hope that the collaboration with China will be a useful
supplement to our other international cooperation.
Fig. 4. An event to present the scientific activities of IG-SCID
during the visit of Mr. David McAllister, Chief Minister of
Niedersachsen, in New Delhi in September 2010 (Prof. Heinz,
Mr. McAllister, Prof. Chhatwal) Photo: Henning Noske

FOCUS RESEARCH REVIEWS SPECIAL FEATURES
Photos from left to right: Lisa Schreiber performing magnetic cell sorting | The assistance to the head of HIPS: administrative manager
David Hofmann, personal assistant Birgitta Lelarge, and scientific assistant Dr. Markus Ehses (from left to right) | Dr. Leonor Gama-Norton
watches a crucial analysis step performed by the PhD student Marcin Cebula. Photos: HZI, Krämer (li) | HIPS/HZI (ce) | HZI, Scheibe (ri)

SCIENTIFIC REPORTS
FACTS AND FIGURES
64
Infection and Immunity
114 New Project Groups
121 PoFII Independent Research
125 Technological Platforms
132 The Helmholtz-Institute for Pharmaceutical
Research Saarland (HIPS)
138 TWINCORE, Centre for Experimental and
Clinical Infection Research GmbH
148 Publications 2010-2011

64 SCIENTIFIC REPORTS | Infection and Immunity
INFECTION AND IMMUNITY
PROGRAMME SPEAKER | Prof. Dr. Dirk Heinz | Department of Molecular Structural Biology |
dih@helmholtz-hzi.de
Infections remain the third most frequent cause of deaths worldwide and hence pose a serious threat to human societies in all
hemispheres. Despite the discovery and development of antibiotics, vaccines and improved hygiene conditions, we are confronted
with the fact that many infectious diseases are re-emerging. The increasing prevalence of antimicrobial resistance added to
globalization, climate change and growing aging populations in industrialized countries poses a continuous challenge in combating
and controlling infectious diseases of humans and domesticated animals. The infl uence of these anthropogenic factors is bestillustrated
by epidemic outbreaks of previously unknown diseases, such as novel zoonotic infections like HIV, SARS, and avian or
swine infl uenza. Moreover, infectious diseases such as tuberculosis, that used to steadily decline in the past, are currently reemerging
as global threats.
In addition to the increasing occurrence of epidemic viral diseases, industrial countries are facing novel challenges in the combating
and containment of bacterial infections. With the advances in modern medicine, the number of immunosuppressed patients highly
susceptible to opportunistic infections (e.g. transplantation patients or those under intensive care) is growing, and a high susceptibility
to infection is observed in the ageing population as a result of an age-related decline in the body’s immune defences. Furthermore,
resistance against virtually all antibiotics presently on the market poses an immense burden on health systems. Novel strategies
for diagnosis, prevention and therapy of infectious diseases are therefore essential for controlling these threats to public health.
The HZI, its subsidiary institute, the Helmholtz Institute for Pharmaceutical Research Saarland (HIPS), and its associated institute
TWINCORE (Centre for Experimental and Clinical Infection Research) together tackle a number of the most important questions in
infection research by building a strong basis in fundamental research and applying this knowledge to develop new strategies for
prevention and therapy of microbial infections for translation into public health benefits. They have set up the Helmholtz programme
“Infection and Immunity”, which comprises fi ve areas of infectious disease research refl ected in the topics
• Microbial Pathogenesis
• Host-Pathogen Interactions
• Infl ammation and Immunity
• Strategies for Prevention and Therapy
• Pharmaceutical Research
The former topic “Translational Infection Research” has become a cross-topic activity since it is highly relevant to all fi ve topics.
The complete infrastructure and expertise necessary to plan, prepare and conduct clinical trials has been developed together with
the Hannover Medical School (MHH). Currently a clinical trial centre for early clinical trials is being set up as a collaborative effort
by the Fraunhofer Institute for Toxicology and Experimental Medicine (ITEM), MHH and HZI.

SCIENTIFIC REPORTS | Infection and Immunity
65
Microbial Pathogenesis tackles the fundamental question of how bacterial pathogens cause disease. Adherence to host cells,
invasion, intracellular survival, dissemination and immune evasion are strategies used by microorganisms to establish infections in
a host. Understanding these complex processes is achieved by investigating the mechanisms of pathogenicity, the role of specifi c
virulence factors and pathogen diversity. Bacterial pathogens under investigation in this topic include Streptococci as causative
agent for respiratory and invasive diseases, Yersiniae and Listeriae as both enteropathogens and model organisms, and Mycobacterium
tuberculosis, the causative agent of tuberculosis. State-of-the-art technologies including structural biology, electron
microscopy and proteomics are utilized for the in-depth investigation of the molecular processes involved.
Studies in Host-Pathogen Interactions focus on understanding the interplay between the infected host and bacterial or viral
pathogens, where genetic and cellular factors controlling the infection process are studied in complex model systems like the
mouse. This also includes the complexity of microbial communities within the host involved in the formation of persistent biofi lms
on medical implants or in cystic fi brosis lungs that are refractory to antimicrobial treatment. Other research activities deal with
the transmission of zoonotic pathogens like infl uenza and prion protein diseases. Large-scale epidemiological studies, such as the
newly developed National Cohort, that help to further understand the complexity of host factors responsible for susceptibility or
resistance against pathogens are also part of this topic.
Inflammation and Immunity addresses the basic processes involved in eliciting innate and adaptive immune responses against
infectious agents. A strong emphasis is placed on understanding the mechanism and regulation of the interferon (IFN) system as
an important constituent of the early immune response and mediator of infl ammation. T cells as key players in the adaptive immunity
are studied with respect to their delicate balance between host protection, tolerance and autoimmunity. New technologies
developed in this research area include the development of novel cell lines and humanized mice as models for infection and host
response. Systems biology approaches are used to model molecular pathways involved in immunity.
The central aim of Strategies for Prevention and Therapy is the development of innovative tools and strategies for the prevention,
management and control of infections and infection-related diseases. One major research focus is the development of new
antigen delivery systems and adjuvants, in particular those amenable for the development of mucosal vaccines. This is linked to
activities where preclinical validation models based on humanized mice for screening, selection and prioritization of candidate interventions
are established. New drug targets relevant for hepatitis C virus (HCV) replication and propagation are investigated using
a cell-based HCV model. Furthermore, virus and host associated factors affecting the quality and quantity of anti-viral cytokine
responses and the protective mechanism of IFN are being studied.
The topic Pharmaceutical Research aims at integrating anti-infective research to bridge the gap between basic biological and
biochemical research and clinical applications, and consequently initiate an early interaction with the pharmaceutical industry. The
central goal is to identify and develop natural products and chemically synthesized compounds as bioactive lead structures, which
can be further developed using medicinal chemistry, total synthesis and biosynthetic approaches. Finally, further studies involve
optimisation of formulation and drug delivery technology to facilitate transport of promising compounds to their targets.

66 SCIENTIFIC REPORTS | Infection and Immunity
With the programme “Infection and Immunity”, the HZI addresses current and future fundamental problems of infection research
not only through the existing areas of research and expertise but also by implementation of a comprehensive set of novel and/
or complimentary initiatives to the current lines of research. These are concentrated in the provision of a pipeline for the development
of novel anti-infectives and their rapid translation into clinical applications.
Currently, expertise and infrastructure in functional genomics and proteomics are further strengthened to provide new approaches
for unravelling the mechanisms and pathways involved in host-pathogen interactions during infection. These will be exploited for
the identifi cation and characterization of targets as a prerequisite for anti-infective intervention. The discovery and development of
anti-infectives is being intensifi ed and signifi cantly expanded by exploiting the enormous genetic potential of microorganisms for
yielding novel bioactive compounds. This will accelerate the development of therapeutic tools with the ultimate goal of translation
into the clinic. Within the newly developed German Centre for Infection Research (Deutsches Zentrum für Infektionsforschung
(DZIF)) it is planned that the HZI will mainly contribute its strengths in natural product research for the development of novel antiinfectives
and its expertise in basic research on bacterial and viral infections. Together with the MHH, the German Collection of
Microorganisms and Cell Cultures (DSMZ) and the Technische Universität Braunschweig, the HZI will signifi cantly expand its activities
in promoting and developing translational infection research in the Braunschweig-Hannover area and beyond to foster effi cient
exploitation of basic research results in the DZIF.
Diarrhea/
Gastroenteritis
2.6 Mill.
AIDS/HIV
2.8 Mill.
Tuberculosis
1.8 Mill. Malaria
1.3 Mill.
Lower Respiratory
Infections
3.9 Mill.
Measles
700.000
Meningitis
170.000
Pertussis
290.000
Hepatitis
600.000
Tetanus
210.000
Most important ifectious diseases No. of deaths - cases/year Year
Lower Respiratory Infections 3.900.000 2002
HIV/AIDS 2.400.000 -3.300.000 2001-2009
Diarrhea / Gastroenteritis 1.800.000 -2.600.000 2002-2009
Tuberculosis 1.600.000 -3.000.000 2001-2009
Malaria 1.300.000 -2.700.000 2002-2009
Measles 700.000 2001
Hepatitis 520.000 -700.000 2005
Pertussis 290.000 2002
Tetanus 210.000 2002
Meningitis 170.000 2002
Infectious diseaes worldwide: Death cases per year Source: WHO (2001-2009)

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
67
01 Microbial Pathogenesis
TOPIC SPEAKER | Prof. Dr. G. Singh Chhatwal | Department of Medical Microbiology | gsc@helmholtz-hzi.de
In spite of the availability of a large number of different antibiotic and antiviral agents, the disease burden caused by infections is
continuously increasing and markedly impairs the achievements of medical care. Due to advanced medical practises chronic persistent
infections are becoming more and more apparent and constitute a major challenge for the medical profession. Furthermore,
the increasing emergence of antimicrobial resistant bacterial isolates is a major concern and multi- and pan-resistant pathogens
are frequently identifi ed especially in the clinical setting. The decreasing therapeutic options make it necessary to develop new
treatment strategies. To meet these challenges, a detailed understanding of the mechanisms of pathogenicity is of utmost importance.
Adherence to and invasion of the host cells, intracellular survival, dissemination, immune evasion – especially evasion of
intracellular defences – and persistence are just a few of the strategies used by microorganisms to establish an infection in the
host. In addition, the carrier stage of the pathogen, which has been neglected in the past, has become an important research
theme because of its role in latent infections. Another area of concern is the diversity of many infectious agents. Many bacterial
and viral species have hundreds of different serotypes with strong antigen variation which impedes development of effective
therapies. Therefore, a multi-disciplinary approach has to be applied in order to fully understand the pathogenic mechanisms of
microorganisms.
The main objective of this topic is the in-depth study of the biology of pathogenic microorganisms and their interaction with the
host cell. The topic deals with the following research themes:
Study of the host cell-pathogen interactions at the molecular and cellular level
In the course of their co-evolution with the host, bacterial pathogens have developed sophisticated strategies to exploit the host
cell and immune system for their own benefi t. Intracellular survival, dissemination in the host, and pathogen persistence require
a complex series of interactions within the host cell. A prime target is the cytoskeleton that is involved in numerous cellular
functions and processes that depend on the intrinsic dynamics of its constituents, in particular the actin system. A large number
of actin binding proteins have been described which regulate the dynamic reorganisation of the cytoskeleton. This research area
deals with the elucidation of exact signalling pathways and principles of the control of actin assembly. Whereas the subversion of
the actin system by a pathogen has been a topic of interest for a long time, the contribution of the dynamic microtubule system
to bacterial pathogenicity is emerging as an important new area of infection research and has been included in this topic. Listeria
monocytogenes and Yersinia enterocolitica are the focus of this research area.
Identification of novel virulence factors from pathogenic bacteria
Pathogenic microorganisms produce a variety of virulence factors with diverse functions in processes such as adherence, invasion,
intracellular survival, evasion of the host immune response and bacterial communication. Their functional characterisation not only
contributes towards our understanding of pathogenicity, but also helps in identifying promising candidates for the development
of vaccines, diagnostics and novel therapeutics. Fibronectin, collagen and plasminogen are important constituents of the extracellular
matrix shown to be directly involved in host-pathogen interactions. Fibronectin binding proteins of pathogenic bacteria
are important factors in adherence, invasion and also intracellular survival. Collagen binding to bacteria plays a role in certain
auto immune manifestations and the activation of plasminogen induced by bacterial proteins is crucial for tissue invasion. The main
focus of this research area is the virulence factors of different streptococci and Yersinia enterocolitica.

68 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
Analysis of bacterial virulence using proteomics
In this theme, the model organism L. monocytogenes is used to unravel the spatial and temporal dynamics of host-pathogen
interactions at the protein level. In particular effector mediated host cell signalling is studied using a qualitative phosphokinome
analysis. Since the progression from first P. aeruginosa acquisition until sustained colonisation of the lung of CF patients correlates
directly with general life expectancy, a highly sensitive immunoproteome workfl ow for the identifi cation of diagnostic and prognostic
antigenic markers is being established together with the outpatient clinic of the Hannover Medical School. Furthermore, the
project investigates signal transduction processes specifi c both for the bacterial pathogen P. aeruginosa and for lung epithelial
cells derived from CF patients.
Structural analysis of proteins involved in host-pathogen interactions
The elucidation of host-pathogen interactions at the atomic level provides mechanistic insights for the development of new
approaches to specifi cally interfere with infection processes. The 3D-structures of microbial virulence factors and, if known, their
complexes with host cell interaction partners and receptors are determined at high resolution, using the well-established techniques
of X-ray crystallography and NMR spectroscopy. The main emphasis is placed on virulence factors that contribute to microbial
adherence and invasion, remodelling of the host cell cytoskeleton, components and effectors of type III secretion systems, key
gene regulators of microbial virulence and amyloids involved in neurodegenerative diseases. We have also included in this topic
the protein production in eukaryotic cells for structural analysis.
The above-mentioned research areas are being handled at project level by a multi-disciplinary team of scientists that belong to
different departments and research groups at HZI and bring with them a variety of different expertise. The highlights of the results
obtained at project level are given in the following project reports.
Genome map of a highly infectious clinical Mycobacterium
tuberculosis strain from Latin America. Genes coding various
functional categories are depicted in different colours.
(see page 80)

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 69
01.1 Structural Analysis of Virulence Factors
PROJECT LEADER | Prof. Dr. Dirk Heinz | Department of Molecular Structural Biology | dih@helmholtz-hzi.de
PROJECT MEMBERS | Davide Ferraris | Thomas Heidler | Dr. Christian Kambach | Dr. Björn Klink |
Dr. Jörn Krauße | Dr. Anja Menzel | Nick Quade | Dr. Stefan Schmelz | Ulrich Wiesand
Pathogenic bacteria use an arsenal of virulence factors to
breach natural host barriers, circumvent or block defense
mechanisms or reprogramme host molecular processes
for their own benefit. As most of these bacterial factors are
missing in the host cell, they represent potential targets for
novel antibiotics. The aim of this project is to gain a precise
understanding of pathogen-host interactions during infection
by X-ray crystallographic structural analysis of microbial
virulence proteins in complex with their host target proteins.
Listeria InlB activates the human receptor tyrosine kinase
Met through dimerization L. monocytogenes is a harmful
food pathogen which can cause listeriosis, a systemic disease
with high mortality rates in neonates, the elderly and other
immuno-compromized individuals. The unique capability of
the bacterium to breach several important host-cell barriers
critically depends on two invasins, internalin A and B (InlB),
located at the bacterial surface. InlB interacts with the receptor
tyrosine kinase Met, leading to bacterial uptake. Met
is the receptor for hepatocyte growth factor (HGF), making
it a key player in cell growth, cell migration, wound healing
and cancer metastasis. Growth factor-mediated receptor
activation typically involves dimerization of the receptor by
the ligand, in this case as InlB2/Met2. In 2007, we determined
the crystal structure of the InlB/Met complex. Like in
the InlB structure, the complex displays a head-to-tail InlB
dimer interface leading to a Met/InlB/InlB/Met-arrangement
compatible with Met function, tyrosine kinase cross-phosphorylation
and receptor activation. To confirm the putative
role of InlB-mediated dimerization for Met activation, we
crosslinked InlB by two symmetrical inter-InlB disulfide
bridges like in the InlB dimers (Fig. 1). Strikingly, this covalent
InlB dimer leads to strong Met activation outperforming
even HGF. Conversely, weakening the InlB dimer interface
by introducing opposing positively charged amino acids
completely abrogates Met signalling.
A new class of bacterial guanine nucleotide exchange
factors for human GTPases Actin cytoskeleton remodelling
is essential for cell division, motility or cell-cell communication.
It is tightly regulated by multiple signalling pathways
employing many different actin-binding proteins. Most if not
all pathways converge on small Rho family GTPases which
act as switches inducing different actin-filament structures.
GTPase signalling requires control by guanine nucleotide exchange
factors (GEFs). Rho-GTPases are key targets of pathogens
modifying cell signalling or actin remodelling for their
own needs. Pathogens frequently bypass or activate Rho-
GTPase pathways by producing virulence factors mimicking
Rho-interacting proteins. This allows them to induce tight
adherence to or entry into host cells. Shigella flexneri IpgB1
and IpgB2 and Map from pathogenic E. coli are prototypic
members of an intriguing novel family of bacterial effectors.
These were originally classified as GTPase mimics, despite
the lack of sequence homology or GTP binding. They induce
filopodia (Map) or lamellipodia and stress fibers indicative
of Rac1 and RhoA activity for IpgB1 and IpgB2, respectively.
We solved the crystal structures of IpgB2 and its complex
with human RhoA (Fig. 2), unambiguously identifying IpgB2
as a bacterial RhoA-GEF not as a functional mimic. Structures
of the complex in various nucleotide bound states revealed
the molecular mechanism of GDP release, an essential
prerequisite for GTP binding.
Fig. 2. Structure of the complex between IpgB2 (blue) and
RhoA (brown). GDP is shown in orange and the GDP-coordinating
Mg 2+ is a yellow sphere. Red spheres are water molecules
coordinating Mg 2+ . The switch I region of RhoA is shown as a
yellow loop, switch II in green. The „catalytic loop“ of IpgB2 is
shown in red.
Fig. 1. Structure of the InlB dimer (blue) covalently cross-linked
by the introduction of two disulfide bridges (yellow sticks,
with electron density).
Ferraris, D. M., Gherardi, E., Di, Y., Heinz, D. W. & Niemann, H. N. (2010). Ligand-mediated
dimerization of the Met receptor tyrosine kinase by the bacterial invasion protein
InlB. Journal of Molecular Biology 395, 522-532.
Klink, B. U., Barden, S., Heidler, T. V., Borchers, C., Ladwein, M., Stradal, T. E., Rottner,
K. & Heinz, D. W. (2010). Structure of Shigella IpgB2 in complex with human RhoA: Implications
for the mechanism of bacterial GEF-mimicry. Journal of Biological Chemistry
285, 17197-17208.

70 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.2 Virulence Factors of Streptococci and Pneumococci
PROJECT LEADER | Prof. Dr. G. Singh Chhatwal | Department of Medical Microbiology | gsc@helmholtz-hzi.de
PROJECT MEMBERS | Silva Amelung | Dr. René Bergmann | Dr. Simone Bergmann | Dr. Marcus Fulde |
Angela Hitzmann | Melanie Lüttge | Dr. Andreas Nerlich | Dr. Patric Nitsche-Schmitz | Priv.-Doz. Dr. Manfred Rohde |
Dr. Vivek Sagar | Dr. Susanne Talay
Streptococcus pyogenes and S. pneumoniae (pneumococci) are
well known human pathogenic bacteria capable of causing a
wide spectrum of diseases. Group C and group G streptococci,
such as S. dysgalactiae ssp equisimilis (SDSE), S. anginosus or
S. canis, which have been considered as animal pathogenic,
are emerging as human pathogenic bacteria. For example
SDSE has recently been related to rheumatic heart disease.
In addition, oral streptococci which can cause dental caries
are also capable of causing serious invasive diseases in humans.
In spite of the availability of antibiotics the morbidity
and mortality due to streptococcal infections remains very
high. It is necessary to elucidate the pathogenicity mechanisms
in order to develop novel control strategies. This is the
major objective of the project. The highlights are:
Neglected streptococcal diseases Our results showed
that oral streptococci possess a pathogenicity mechanism
that has until now been described only for S. pyogenes and
S. pneumoniae. This mechanism involves binding and activation
of plasminogen by oral streptococci leading to tissue
destruction and dissemination in the host. The major bacterial
factors involved in this process are M-proteins, which
are expressed by S. pyogenes, and, as we described recently,
also by SDSE and S. canis.
The correlation of SDSE with rheumatic heart disease is a
cause of concern. SDSE strains from South India can bind
collagen through PARF peptide leading to autoimmune
reaction. High prevalence of SDSE indicates that this
neglected streptococcal species must be taken seriously
and should be eradicated.
Interaction of streptococci with endothelial cells
Prerequisite for streptococcal invasive disease is the dissemination
of bacteria via blood circulation. To evade the immune
system the streptococci invade endothelial cells through a
phagocytosis-like process. We could show that a streptococcal
protein SpyCep, which can degrade IL-8, is involved in the
immune evasion process leading to invasive infection.
Invasion of SpyCep-expressing S. pyogenes (left) and SpyCepcoated
latex beads (right) into human endothelial cells
Photo: HZI, Rohde
Pneumococci are the major organism involved in human
pneumonia. Using a human genome microarray we could
show that pneumococci can influence the gene expression
profile of lung endothelium. In addition, we could identify
phosphoglyceratkinase, a cytoplasmic enzyme of Pneumococcus
that is found on bacterial surface and can also bind
plasminogen thereby facilitating tissue invasion.
Activated plasmin(ogen) on the surface of S. canis can destroy
fibrin, leading to bacterial dissemination in the host Photo: HZI, Rohde
The role of group C and group G streptococci in human
diseases is not well investigated. Our results showed that
in South India, where such infections are very common, 80
percent of the diseases were caused by SDSE. Surprisingly,
S. anginosus was a causative organism in 20 percent of the
infections. We have recently developed a test which can
identify S. anginosus with a high degree of specificity.
In summary, the elucidation of the mechanisms involving
interaction of streptococci with human endothelial cells and
plasminogen offers new control strategies for invasive streptococcal
diseases. The study on neglected streptococcal
species allows us to understand the common pathogenicity
mechanisms among different streptococci that may lead to
novel diagnostic and therapeutic approaches.
Bergmann, R, Dinkla, K, Nitsche-Schmitz, DP, Graham, RM, Lüttge, M, Sanderson-
Smith, M, Nerlich, A, Rohde, M, Chhatwal, GS (2010) Biological functions of GCS3, a
novel plasminogen binding protein of Streptococcus dysgalactiae ssp. equisimilis.
International Journal of Medical Microbiology. doi:10,1016/j.ijmm.2010.06.007
Itzek A, Gillen CM, Fulde M, Friedrichs C, Rodloff AC, Chhatwal GS, Nitsche-Schmitz DP
(2010) Contribution of plasminogen activation towards the pathogenic potential of oral
streptococci. PLoS One 5: e13826

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 71
01.3 Molecular Mechanisms of Intracellular Trafficking,
Survival and Persistence of Streptococci
PROJECT LEADER | Priv-Doz. Dr. Manfred Rohde | Dept. Medical Microbiology | mro@helmholtz-hzi.de
PROJECT MEMBERS | Katja Branitzki-Heinemann
Streptococcus pyogenes, the group A Streptococcus (GAS), is
the major cause of human streptococcal infections that range
from uncomplicated to severe and life-threatening infections
like necrotizing fasciitis. Streptococci are also able to cause
recurrent infections such as erysipel and tonsillitis. This phenomenon
is described as carrier stage of streptococcal infections.
It allows GAS not only to persist but also to resist prolonged
antibiotic treatment. The carrier stage of streptococci has
been more or less neglected in the past. Only scant information
is available on the mechanisms and the factors involved.
With the help of SfbIGaro coated gold-nanoparticles we could
show that fibronectin binding does not result in integrin
clustering (D) as seen for GfbA. Only GfbApro mediated
fibronectin binding results in integrin clustering (H). Thus,
we demonstrate for the first time a biological function of the
aromatic domain of a fibronectin binding protein of a pathogenic
bacterium. In addition, first evidence is coming up that
streptococci invading via membrane ruffling always seem to
fuse with lysosomes.
Invasion and survival mechanisms of streptococci
Fibronectin binding proteins of streptococci have been
demonstrated to play an important role in adherence and
invasion. The streptococcal fibronectin-binding protein (SfbI)
from group A streptococci mediates adherence and invasion
through invaginations (A) and is also involved in the
intracellular survival of streptococci by formation of a novel
compartment termed caveosome. SfbI utilizes fibronectin as
a bridging molecule to bind to α 5
ß 1
integrins. By co-opting
this caveolae-mediated pathway SfbI-carrying streptococci
bypass the degradation mechanism of the host cell since no
fusion with lysosomes is detectable.
GfbA-expressing Group G streptococci exhibit GfbA on their
surface which also binds fibronectin and interacts with
α 5
ß 1
integrins. But GfbA-mediated invasion leads to the
formation of membrane ruffles (E), rearrangements of the
host-cell cytoskeleton, despite the fact that similar amounts
of fibronectin are bound compared to the SfbI-carrying
strains. GfbA-expressing streptococci follow the classical
endocytic pathway: fusion of lysosomes and phagosomes to
form phagolysosomes. By heterologous surface expression of
GfbA in the non-pathogenic S. gordonii it was demonstrated
that GfbA alone is responsible for the morphological distinct
invasion mechanism compared to the SfbI-mediated mechanism.
Sequencing of the GfbA gene demonstrated that only
the C-terminal part shows a high similarity with SfbI and the
N-terminal part, including the aromatic domain of GfbA and
SfbI, shows significant differences.
Expression of chimaric proteins from SfbI and GfbA
Based on the sequence data the N-terminal aromatic domain
was assumed to be responsible for the different invasion mechanism.
Therefore, a GfbA protein was constructed without
the aromatic domain (GfbApro) and the aromatic domain in
SfbI was replaced by the aromatic domain of GfbA (SfbGaro).
Both proteins were expressed on the surface of S. gordonii.
GfbApro mediated invasion showed the typical caveolaemediated
invasion pattern (F) without fusion with lysosomes
(G). On the other hand SfbIGaro was now accompanied by
membrane ruffling (B) with lysosomal fusion events (C).
A)SfbI-expressing streptococci invade host cells via the formation
of invaginations. E) GfbA-expressing streptococci invade via
membrane ruffles. B) heterologous expression of SfbI containing
the aromatic domain of GfbA (SfbIGaro) on S. gordonii
leads to invasion via membrane ruffles and fusion with lysosomes
(C); streptococci in red, lysosomes (green) are labeled for
the Lamp-1 protein. F) deletion of the aromatic domain in GfbA
(GfbApro) results in an invasion via invaginations and no fusion
with lysosomes (G). D) binding of SfbIGaro with fibronektin
onto integrins results in no integrin-clustering since only single
gold-nanoparticles are detectable (D, arrows), whereas binding
of fibronectin to GfbApro leads to integrin-clustering, large
gold-nanoparticles aggregation (H, arrows). Photos: HZI, Rohde
Nerlich, A., Rohde, M., Talay, S., Genth, H., Just, I., Chhatwal, G.S. (2009) Invasion of
endothelial cells by tissue invasive M3 type Group A streptococci requires Src kinase
and activation of Rac1 by a PI3-kinase independent mechanism. J. Biol.Chem., 284,
20319-20328
Rohde, M., Graham, R.M., Branitzki-Heinemann, K., Borchers, P., Preuss, C., Schleicher,
I., Zähner, D., Talay, S.R., Fulde, M., Dinkla, K., Chhatwal, G.S. (2011) Differences in
the aromatic domain of homologous streptococcal fibronectin-binding proteins trigger
different cell invasion mechanisms and survival rates. Cell. Microbiol. 13, 450-468

72 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.4 Analysis of Protein Networks Induced by Early
Host-pathogen Interactions
PROJECT LEADER | Dr. Lothar Jänsch | Research Group Cellular Proteomics | lja@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Uwe Kärst | Dr. Sebastian König | Dr. Manfred Nimtz | Dr. Tobias Reinl |
Dr. Josef Wissing | Evelin Berger | Susanne Freund | Christoph Gernert | Alexander Iphöfer | Thorsten Johl |
Kirstin Jurrat | Zofi a Magnowska | Maxi Scheiter
The Research Group Cellular Proteomics focuses on the
analysis of fundamental events during signal-transduction
processes which are important for infectious processes in
humans and the host’s adaptive immune responses towards
pathogens. Modules for cell biological, biochemical, mass
spectrometric and bioinformatic approaches have been developed
and combined for this purpose. Our aim: a quantitative
and time-resolved analysis of protein expression, localisation,
interaction and their post-translational modifications
(PTMs) in primary and immortalised human cells.
Methods Quantitative chemical proteome analyses facilitate
the identification of cellular “target proteins” and the
understanding of signalling networks. Therefore, we optimise
small molecules used in therapeutics and use them
after immobilisation as “baits” for interacting proteins. In
combination with chromatographic and mass spectrometric
methods these tools enable the analysis of transient structural
modifications on signalling components. In the case
of phosphorylations this refers to protein kinases, i.e. their
activities and their molecular interactions with substrate
molecules. iTRAQ-technology is used for quantitative
peptide sequencing (Orbitrap Velos) and iTRAQassist for
statistical evaluations, both matching the quality criteria
for clinical studies.
Signalling pathways depending on phosphorylations
during invasion by Listeria monocytogenes The bacterial
pathogen L. monocytogenes causes severe illnesses and
prenatal infections. The virulence factors InlA and InlB
induce the invasion of host cells by interacting with the
adhesin E-Cadherin (InlA) and the receptor tyrosine kinase
c-Met (InlB). The signalling pathways involved are controlled
through protein phosphorylations by kinases. A direct
analysis of time-resolved phosphorylation events on kinases
was demonstrated for the InlB-activated c-Met pathway for
the first time worldwide. This work led to the identification
of signalling components involved in listerial invasion that
were not described so far for the c-Met pathways activated
by the physiological ligand HGF. Functional investigations
of these kinases thus analyse their contribution to the
invasion as well as to motogenic and mitogenic processes.
Chemical proteomics at the HZI already can access about
70% of the total kinome from humans and mice and in addition
has devised novel tools, which are now complementing
information from the ubiquitin-signal network.
“Kinase tree” of all known human kinases; identified human
kinases are labelled red, the corresponding orthologs from
mouse in green. Graphic: HZI
Characterisation of signal transduction pathways in activated
NK and T cells T lymphocytes are essential for regulating
the immune system. Their actual cellular processes
and effector functions depend directly on the activation status
of such pathways. Quantitative phosphokinome analyses
with different regulatory T cells detected novel components
of the CD3/CD28-dependent activation pathways. As part
of a comparative analysis of CD3/CD28-activated signal
networks of effectory and regulatory T cells we analysed
the expression and phosphorylation status of about 150
kinases. We identified novel signalling components and
phosphorylation sites that are currently being investigated
to clarify their roles in the formation and function of suppressor
T cells in humans and mice. The methods developed
for analysing T cells are now also being applied for the first
time for the characterisation of activated NK-(natural killer)
cells that are part of the innate immune response.
Hemmen, K. Reinl, T.; Buttler, K. Behler, F., Dieken, H. Jaensch, L., Wilting, J., Weich,
H.A. (2010) High-resolution mass spectrometric analysis of the secretome from mouse
lung endothelial progenitor cells. Angiogenesis, in press
Reinl T., Nimtz, M., Hundertmark, C., Johl, T., Kéri, G., Wehland, J., Daub, H. and Jänsch,
L. (2009) Quantitative phosphokinome analysis of the Met pathway activated by the
invasin InlB from Listeria monocytogenes. Molecular Cell Proteomics 8(12):2778-95.
Hundertmark C., Fischer R., Reinl T., Klawonn F. and Jänsch L. (2009) MS-specific
noise model reveals the potential of iTRAQ in quantitative proteomics. Bioinformatics
25(8):1004-11.

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 73
01.5 Structural and Mechanistical Analysis of Functional
Amyloids
PROJECT LEADER | Prof. Dr. Christiane Ritter | Junior Research Group Macromolecular Infections |
cri07@helmholtz-hzi.de
PROJECT MEMBERS | Agnes Zimmer | Madhu Nagaraj | Johannes Spehr | Tobias Schubeis
Bacterial fibrillar adhesins with an amyloid-like fold have
recently been identified as a novel class of adhesins that
are expressed by many bacteria, both gram-positive and
gram-negative. A study on biofilms from different habitats established
that up to 40% of the bacteria present within these
communities carry amyloid-like fibrils on their surface. They
have been identified as virulence traits important for biofilm
formation, interaction with host proteins and promoting
survival in a wide range of conditions. Amyloid-like fibrils
are ordered aggregates that have a high content of specific
b-sheet secondary structure. Long associated with human
diseases such as Alzheimer’s disease, Parkinson’s disease
or Creutzfeld-Jacob disease the amyloid fold is now known to
be formed natively by a large set of diverse proteins without
toxic side-effects.
The major focus of our group lies on the determination of
the structural and mechanistical basis of these functions for
selected bacterial and fungal amyloids.
Tools for the structure determination of amyloid fibrils
One major bottleneck in amyloid research is the sparseness
of high-resolution structural information on naturally occurring
amyloids, because the size and non-crystalline nature of
the fibrils drastically restrict the use of established structural
techniques. We have, therefore, introduced a new, generally
applicable approach that derives the fold of amyloid
fibrils from a combination of quenched hydrogen exchange
measured by nuclear magnetic resonance (NMR), solid state
NMR and other spectroscopic techniques. Solid-state NMR
is a new method to determine high-resolution structures of
proteins. It is particularly interesting for fibrous proteins. We
are therefore in the process of establishing this technique at
the HZI, and we develop biochemical tools to generate protein
samples with selective isotope labelling patterns.
Curli: an amyloid coat that increases bacterial virulence
Curli is the major proteinaceous component of the extracellular
matrix produced by Enterobacteriaceae such as E. coli
and Salmonella typhimurium. Curli fibrils are involved in the
adhesion to biotic and abiotic surfaces and the promotion
of biofilm formation. They also interact with several host
proteins resulting in increased tissue penetration, inflammation
and sepsis. In vivo curli biogenesis is dependent
on the nucleation of the major curli component, CsgA, by
the homologous protein CsgB. In order to understand the
features responsible for the different functionalities of CsgA
and CsgB we analyse the structures, aggregation kinetics
and thermodynamic stabilities of the fibrils formed by both
extracellular space
Model of curli biogenesis. Curli expression is controlled by
the positive transcriptional regulator CsgD. CsgA is the major
structural subunit which forms a fibrillar structure upon nucleation
by the homologous protein CsgB. CsgG is a membran
chanel required for export of the structural subunits, and
CsgE and CsgF a chaperone-like molecules crucial for efficient
curli assembly. The insert depicts a model of a single CsgA
molecule within the fibrillar structure derived from quenched
hydrogen exchange NMR and sequence alignments.
proteins. In addition, we analyse the structure and function
of chaperones that are essential for curli biogenesis in vivo.
A mechanistical understanding of curli biogenesis will allow
us to identify new ways to interfere with fibril formation in
order to reduce bacterial virulence.
HET-s: a functional prion found in filamentous fungi
Only subsets of amyloids are self-propagating prions. To understand
the molecular determinants that govern the infectivity
of an amyloid we investigate the biophysical properties
of the functional prion protein HET-s from the filamentous
fungus Podospora anserina and of a recently identified homolog
from Fusarium graminearum. Despite limited sequence
identity the amyloid fibrils formed by both proteins can
cross-seed each other, i.e. they are able to breech the species
barrier. Using quenched hydrogen exchange NMR we could
explain this observation with a high structural similarity of
the amyloid fibrils.
Wasmer, C., Zimmer, A., Sabate, R., Soragni, A., Saupe, S.J., Ritter, C., and Meier, B.H.
(2010). Structural Similarity between the Prion Domain of HET-s and a Homologue
Can Explain Amyloid Cross-Seeding in Spite of Limited Sequence Identity. Journal of
Molecular Biology 402(2): 311-325.
Greenwald, J., Buhtz, C., Ritter, C., Kwiatkowski, W., Choe, S., Maddelein, M.L., Ness,
F., Cescau, S., Soragni, A., Leitz, D., Saupe, S.J., and Riek, R. (2010). The Mechanism of
prion inhibition by HET-S. Molecular Cell 38: 889-899.

74 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.6 Microtubule Dynamics and Bacterial Pathogenesis
PROJECT LEADER | Prof. Dr. Jürgen Wehland (†) | Research Group Cell Biology
PROJECT MEMBERS | Dr. Marco van Ham (mvh06@helmholtz-hzi.de) | Dr. Andreas Fischer | Dr. Marcin Ura |
Ramona Baier
Whereas the subversion of the actin system by pathogens
has been studied in great detail, much less is known about
the contribution of the dynamic microtubule system in
bacterial pathogenicity. Microtubules are essential for a
variety of cellular functions such as cell division, intracellular
organelle/vesicle transport and different forms
of cell motility. During the past few years we have been
focusing on a specific post-translational modification of
tubulin to study the contribution of microtubule sub-sets
to these various functions. A unique enzyme respon sible
for tubulin-tyrosination, the tubulin-tyrosine-ligase (TTL),
is well characterized due to substantial contributions from
our laboratory. The tyrosination cycle is highly conserved
among eukaryotes. To test the physiological importance of
the tubulin tyrosination cycle, we recently generated TTL
deficient mice. Analysis of these TTL null mice showed severe
disorganization of the brain, involving loss of cells and
abnormal neuronal extensions leading to perinatal death
demonstrating the vital role of this highly specific tubulin
modification.
TTL null mice Constitutive TTL null mice were generated
by conventional gene knockout and newborn TTL null mice
were undistinguishable from their wild type littermates.
However, TTL null mice displayed defective breathing
and ataxia and died within 24 hours after birth due to an
intricate combination of brain defects involving anomalies
in neuronal proliferation/differentiation balance and in
defective control of neurite outgrowth and differentiation.
On the cellular level, our results indicate that the tubulin
tyrosination cycle regulates microtubule interactions with
CAP-Gly domain-containing microtubule plus-end tracking
proteins, which provide explanations for the involvement of
TTL in the establishment of cell polarity.
Conditional TTL knockout in mice Since the constitutive
TTL knockout leads to perinatal death, the question arises
as to whether TTL deficiency per turbs other physiological
activities outside the brain and in addition affects adult
mice. To explore this, we recently have generated a conditional
TTL knockout mouse using the cre/lox system. Preliminary
data showed cell type specific deletion of the TTL
gene resulting in enriched levels of detyrosinated tubulin.
We are now in the phase of expanding TTL deficient mouse
lines that will be used to analyze the aforementioned brain
phenotype in more detail. Furthermore, we will analyse the
involvement of the TTL cycle in cells of different hematopoietic
lineages.
Perspective We are aiming at a detailed understanding of
the tubulin tyrosination cycle. This includes the elucidation
of the function of specific microtubule-binding proteins (e.g.
CLIPs) in the regulation of microtubule dynamics in the
background of TTL deficiency. This will be pursued both in
vivo and in vitro by the means of conditional TTL knockout
mice and using tissue culture systems focusing on cell
adhesion and polarity, respectively. In addition, a central
question of particular interest is in the cytoskeleton field:
how microtubules interact with the actin cytoskeleton and
how does the communication between the two cytoskeletal
systems work. Furthermore, of utmost interest are the questions,
(i) how bacterial and viral pathogens can exploit the
microtubule system for their own advantage especially by
usurping specific host cell pathways, and (ii) how pathogens
interfere with the crosstalk between both the actin and
microtubule system.
A migrating mouse embryonic fibroblast immunostained for the
microtubule network (red) and actin-rich structures in the lamellipodium
(green). Photo: HZI
Schwan, C., Stecher, B., Tzivelekidis, T., van Ham, M., Rohde, M., Hardt, W.D., Wehland,
J., Aktories, K. (2009) Clostridium difficile toxin CDT induces formation of microtubulebased
protrusions and increases adherence of bacteria. PLoS Pathogens 5, e1000626.
Erck, C., Peris, L., Andrieux, A., Meissirel, C., Gruber, A.D., Vernet, M., Schweitzer, A.,
Saoudi, Y., Pointu, H., Bosc, C., Salin, P.A., Job, D., Wehland, J. (2005) A vital role of
tubulin-tyrosine-ligase for neuronal organization. Proceedings of the National Academy
of Sciences USA 102, 7853-7858.

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 75
01.7 Function and Regulation of Yersinia Virulence Factors
PROJECT LEADER | Prof. Dr. Petra Dersch | Department of Molecular Infection Biology | pde08@helmholtz-hzi.de
PROJECT MEMBERS | Katja Böhme | Katharina Herbst | Dr. AnnKathrin Heroven | Dr. Annika Kochhut |
Wiebke Opitz | Dr. Fabio Pisano | Rebekka Steinmann | Tatjana Stolz | Tanja Thiermann | Frank Uliczka
Enteropathogenic Yersinia species cause a large range of
different gut-associated diseases, including enteritis, colitis,
mesenteric lymphadenitis which are commonly called yersiniosis.
In rare cases they can also induce certain autoimmune
diseases such as reactive arthritis. Yersiniae are widely
distributed and contaminated porc is the main source of
infection for men. They express a set of special surface structures
which allow them to bind and internalize into host cells.
These outer membrane proteins (adhesins and invasins)
promote efficient attachment to cell receptors which results
in the induction of certain signal transduction pathways
within the cell. Recruitment and activation of certain signal
molecules to bacteria bound cell receptors induce rearrangements
of the actin cytoskeleton and lead to the formation
of special membrane protrusions which migrate around the
bacteria and enclose it into a membrane-bound phagosome.
Cell invasion allows transcytosis of the bacteria through the
gut epithelium into the underlying lymphatic tissues and
deeper organs. We focus on the characterization of the function
and expression of Yersinia invasion factors to gain more
information about how enteric bacteria colonize host tissues
and become resistant against the host immune system.
Molecular analysis of the Yersinia-induced signal transduction
in epithelial cells In order to identify signalling pathways
which are essential for the internalization of Yersinia
pseudotuberculosis, we studied the cell uptake promoted by
the Yersinia invasion factors InvA and YadA which interact
directly or indirectly via extracellular matrix proteins
with β 1
-integrin receptors of the host cell. By colocalization
studies and kinase activation assays, we could demonstrate
that the protein kinase B (Akt), the phospholipase C-γ and
variants of the protein kinase C are recruited to bacteriabound
membranes and are activated in a time-dependent
manner. Application of inhibitors and use of knock-out cell
lines and RNA interference demonstrated that activation of
these factors occurs after activation of the integrin-bound
focal adhesion kinase (FAK), c-Src and the PI3 kinase and
are required for the bacterial entry process. Furthermore,
we obtained evidence that, besides the small GTPase
Rac-1, also other GTPases of the Rho family and different
actin-associated proteins, such as N-WASP and the Arp2/3
complex, are implicated in the uptake process. Furthermore,
we characterized two newly identified adhesins of Y. pseudotuberculosis
with high homology to the invasin protein, and
found that both promote tight binding to human enterocytes.
First analyses in the mouse infection model revealed that
loss of the adhesins prolonged the life time of infected mice
significantly.
Temperature-dependent expression of Yersinia virulence
factors Another important goal is to understand the
molecular control mechanism of Yersinia virulence genes
during the infection process. Temperature is one of the most
crucial factors sensed by the pathogens to adjust expression
of their virulence factors after entry from a cold external
environment into a warm-blooded host. We found that the
regulator protein RovA controlling expression of the invasin
protein undergoes a reversible conformational change upon
a temperature shift from 30°C to 37°C. This reduces the
DNA-binding affinity of the regulator and renders it more
susceptible for proteolysis by the bacterial protease Lon. In
addition, other post-transcriptional regulatory mechanisms
were analysed. These control systems are used to adjust
virulence gene expression during the infection process to
availability of nutrients and stress superimposed by the host
immune system.
Herbst, K., Bujara, M., Heroven, A.K., Opitz, W., Weichert, M., Zimmermann, A., &
Dersch, P. (2009) Intrinsic thermal sensing controls proteolysis of Yersinia virulence
regulator RovA PLoS Pathogens 5(5):e1000435.
Adherent Y. enterocolitica on human epithelial cells.
Photo: HZI, Rohde
Uliczka, F., Kornprobst, T., Eitel, J., Schneider, D., & Dersch P. (2009) Cell invasion of
Yersinia pseudotuberculosis by invasin and YadA requires protein kinase C, PLC-γ 1 and
Akt kinase Cellular Microbiology 11, 1782–1801.
Heroven, A.K., Böhme, K., Rohde, M., & Dersch, P. (2008) A Csr regulatory system, including
small non-coding RNAs, regulates the global virulence regulator RovA of Yersinia
pseudotuberculosis through RovM Molecular Microbiology 68,1179-1195.

76 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.8 Structural Characterisation of Pathogen Defence
Factors
PROJECT LEADER | Dr. Konrad Büssow | Research Group Recombinant Protein Expression |
kbu07@helmholtz-hzi.de
PROJECT MEMBERS | Sonja Wilke | Sarah Tokarski
Many different proteins are involved in the defence against
pathogens. The elucidation of the three dimensional structure
of proteins via X-ray structural analysis provides important
information about the way in which these function.
Structural investigations of many human proteins have not
proved possible to date, due to the fact that they were unable
to be produced in a pure state in sufficient quantities.
This is the prerequisite in order for protein crystals to be
cultivated and X-ray diffraction data recorded.
The majority of proteins for use in X-ray structural analysis
are produced in bacteria. The bacteria are genetically
modified to enable them to produce the respective target
protein in large quantities. This procedure is both rapid and
economical. However, many human proteins involved in
the defence against pathogens are unable to be produced in
this manner. The necessary processing stages required to
transfer these proteins to their biologically active form are
not implemented.
Proteins from animal cell cultures Cultivated animal cells
are an alternative to bacteria in the production of proteins
for X-ray structural analysis. In the majority of cases insect
cells are infected with genetically modified baculoviruses.
However, mammalian cell lines are also highly useful, in
particular in the case of proteins discharged from the cells
in their natural environment that are found in extracellular
fluid or on the exterior of cells.
These discharged proteins, for example antibodies or
cytokines, are often stabilized via disulphide bridges and
carry carbohydrate chains on their surface. The hamster cell
line CHO-Lec 3.2.8.1 is suitable for producing these for X-ray
structural analysis purposes. Mutations here mean that the
carbohydrate chains are small and uniform, with the consequence
that the proteins produced are easily crystallized.
Accelerated cell line generation A disadvantage here is
the long time, approximately one year, that is required for
the creation of a genetically modified CHO-Lec cell line using
standard methods. Introducing a new procedure we have
now been able to significantly reduce this time. This procedure
is based on a fluorescent reporter gene, GFP, which
enables the cloning of genetically modified CHO-Lec cells
with particularly good production properties with fluorescence
activated cell sorting (FACS).
Recombinase-mediated cassette exchange technology
(RMCE) enables a further acceleration of the process of pro-
Cloning of a stable GFP cell line through fluorescence-activated
cell sorting Graphic: HZI
ducing cell lines. RMCE uses site-directed recombination
to replace a marker gene such as GFP with another gene of
choice. This process takes just a few weeks to be completed.
We have combined the cloning of a GFP cell line via cell
sorting with RMCE and have already been able to successfully
demonstrate the cassette exchange in CHO-Lec cells.
We have established production cell lines for a number
of lysosomal associated membrane proteins (LAMP) with
this technique. These cell lines secret engineered LAMP
proteins that lack the membrane spanning region. LAMPs
play an important role in the lysosomes and are required
for elimination of pathogens by phagocytosis. A number of
LAMPs were produced in larger amounts and crystallized.
The spatial structure of a LAMP was determined for the
first time. However, the quality of the structure still has to
be improved for a detailed understanding of the molecular
details of these important proteins.
Wilke,S., Krausze,J., Gossen,M., Gröbe,L., Jager,V., Gherardi,E., van den,H.J., & Bussow,K.
(2010) Glycoprotein production for structure analysis with stable, glycosylation mutant
CHO cell lines established by fluorescence-activated cell sorting. Protein Science 19,
1264-1271.

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 77
01.9 Mycobacterial Phagosomes and Immunity
PROJECT LEADER | Dr. Maximiliano G. Gutierrez | Junior Research Group Phagosome Biology |
mgg08@helmholtz-hzi.de
PROJECT MEMBERS | Achim Gronow | Bahram Kasmapour | Gang Pei | Dr. Cristina Vazquez
M. tuberculosis owes its success to its ability to survive
within macrophage phagosomes. M. tuberculosis blocks
phagosome maturation, specifically the stage of phagosomelysosome
fusion, and as a consequence grows within macrophages,
contributing to disease. The mechanisms whereby
M. tuberculosis arrests the phagosome maturation are still
elusive. The machinery by which mycobacteria manipulate
intracellular defenses is likely multifactorial and dynamic.
Many intracellular targets have been invoked to explain the
block in bactericidal performance of the infected macrophage
such as rab proteins. From these studies, it becomes clear
that interference of intracellular trafficking is a key event in
the survival strategy of this pathogen.
We have already shown that NF-kB activation is an event
required for the killing of intracellular mycobacteria by
macrophages (Gutierrez et al., 2009). More important, block
of NF-kB activation leads to inhibition of mycobacterial
phagosomes fusion with lysosomes. In this study, a whole
genome microarray analysis also identified some interesting
candidates on which the projects in our laboratory are based.
The most promising candidates are Rab34, Rab20 and sortilin.
Since the role of these proteins in phagosome maturation
is not known, the first part of the project consisted in the
characterization and elucidation of protein function in the
context of phagosome maturation. This knowledge will allow
us to understand the intracellular behavior of these proteins
during mycobacterial infection of macrophages.
Rab GTPases and phagosome maturation For Rab34, we
have performed the in-deep characterization of a polyclonal
antibody we generated and determined that the main localization
of Rab34 in macrophages is the Golgi complex. We
also found, using live cell imaging, western blot of purified
phagosomes and electron microscopy, that Rab34 is associated
to phagosomes transiently. As this rab protein likely
has a role in phago-lysosomal fusion, we tested the effect
of Rab34 mutants in the acquisition of lysosomal markers
by phagosomes. Our data indicate that expression of the
constitutively active mutant increased the fusion between
phagosomes and lysosomes.
In the case of Rab20, we cloned Rab20 in pEGFP-C1 and generated
a negative mutant (unable to bind GTP) for studies in
macrophages. Preliminary studies revealed that Rab20 was
associated to the Golgi and large vesicular structures that
remain to be characterized. Our preliminary data point out
for a role during macropinosome formation. Interestingly, we
also have found associated to latex bead phagosomes during
early times of internalization. In the context of mycobacterial
infection, Rab20-GFP was heavily recruited into mycobacterial
phagosomes (Figure 1). We are currently testing whether
this recruitment could have an impact in phagosome maturation.
Sorting of lysosomal enzymes to phagosomes We found
that sortilin mediates the direct trafficking of selected
lysosomal enzymes from Golgi to the latex bead phagosome.
This represents a novel pathway by which selected proteins
are transported to the phagosome bypassing lysosomes
during maturation of phagosomes (Wähe et al., 2010). We
plan to expand these studies to mycobacterial trafficking and
killing using a sortilin KO mouse (in collaboration with Dr.
A. Nykjaer, Aarhus, Denmark).We are also investigating the
role of sortilin in mycobacterial phagosome maturation using
different mutants of sortilin that are not able to signal for
adaptor proteins, endocytosis or retromer interaction.
Fabrino, D. L., Bleck, C. K., Anes, E., Hasilik, A., Melo, R. C., Niederweis, M., Griffiths, G.,
& Gutierrez, M. G. (2009) Porins facilitate nitric oxide-mediated killing of mycobacteria.
Microbes and Infection 11, 868-875.
Fig. 1. RAW 264.7 macrophage expressing Rab20-GFP (green)
and heavily infected with mycobacteria (red). Some of the bacteria
are localized in large vacuoles positive for Rab20-GFP.
Photo: HZI, Gang Pei
Gutierrez, M. G. & Griffiths, G. (2009) Phagosome-cytoskeleton interactions. In: Intracellular
Niches of Pathogens A Pathogens Guide through the Host Cell, (Schaible, U.; Haas,
A.) Wiley-VCH, Weinheim, Germany.
Wähe, A., Kasmapour, B. , Schmaderer, C., Liebl, D., Sandhoff, K., Nykjaer, A., Griffiths,
G., & Gutierrez, M. G. (2010) Sortilin mediates the transport of Acid Sphingomyelinase
and Prosaposin from the Golgi to phagosomes. Journal of Cell Science 123, 2502-11 (Cover).

78 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.10 Molecular Mechanisms of Host-cell Pathogen
Interactions
PROJECT LEADER | Prof. Dr. Klemens Rottner | Former Research Group Cytoskeleton Dynamics |
krottner@uni-bonn.de
PROJECT MEMBERS | Dr. Jennifer Block | Dr. Stefan Köstler | Markus Ladwein | Margit Oelkers |
Dr. Malgorzata Szczodrak
This project aims at characterising the precise molecular
mechanisms driving actin reorganisation during different
types of motility processes and the interaction of different
pathogens with their hosts.
Actin polymerisation is catalysed by protein complexes
enhancing actin filament nucleation, such as the Arp2/3-
complex or formins. Prominent important actin regulators
include activators of Arp2/3-complex, e.g. WASP and
WAVE family members, which can operate downstream
of the Rho-GTPases Cdc42 and Rac1. Novel members of
this Arp2/3-complex activators include proteins of the
WHAMM/JMY subfamily and WASH (Rottner et al., 2010),
the latter of which we could recently identify to contribute
to Salmonella invasion (Hänisch et al., 2010).
Another prominent Arp2/3-complex binding protein that
had previously been implicated in dynamic actin reorganization
events such as lamellipodium protrusion or different
types of host-pathogen interaction is the Src-substrate
cortactin. We have generated cells and mice harbouring a
conditional mutation of the cortactin gene. To our surprise,
genetic removal of cortactin in fibroblasts did not abolish
Arp2/3-complex activation and actin polymerization
stimulated by constitutively active Rac1. However, cortactin
deletion abrogated signal transduction to activation of
Rac or Cdc42 small GTPases, for instance downstream of
growth factor activation, thus affecting migration and actin
reorganization more indirectly than previously thought (Lai
et al., 2009).
These results are consistent with more recent data obtained
by employing a novel actin assembly assay that we could
recently develop. We demonstrated that cortactin is incapable
of driving Arp2/3-complex-dependent actin assembly
at ectopic sites in living cytoplasm (Oelkers et al., submitted).
They also confirmed our previous data on the relative
turnover of cortactin and Arp2/3-complex in the lamellipodium,
which had already indicated that the majority of the
cortactin molecules associating with the structure cannot
engage in Arp2/3-complex activation at the lamellipodium
tip (Lai et al., 2008).
Fig. 1. Cortactin is essential for InlB-mediated Listeria invasion.
Quantification of Listeria entry in the presence (Control) and
absence (Cortactin KO) of cortactin, as indicated. Listeria
genetically deficient for both Internalin (A) and InlB (grey) were
used as non-specific invasion control. Graphic: HZI
Finally, in more recent efforts, we explored the role of
cortactin in different types of host-pathogen interaction.
Interestingly, cortactin appears essential for InlB-mediated
Listeria invasion (Fig. 1), but neither for entry triggered by
Shigella flexneri nor for actin pedestal formation induced
at the cell surface by different types of pathogenic E. coli
(Oelkers/Lai et al., unpublished).
Rottner, K., Hänisch, J., & Campellone, K. (2010) WASH, WHAMM and JMY: Regulation
of Arp2/3 complex and beyond. Trends in Cell Biology 20(11), 650-61.
Hänisch, J., Ehinger, J., Ladwein, M., Rohde, M., Derivery, E., Bosse, T., Steffen, A.,
Bumann, D., Misselwitz, B., Hardt, W.-D., Gautreau, A., Stradal, T.E.B., & Rottner, K.
(2010) Molecular dissection of Salmonella-induced membrane ruffling versus invasion.
Cellular Microbiology 12(1), 84-98.
Lai, F.P.L., Szczodrak, M., Oelkers, J.M., Ladwein, M., Acconcia, F., Benesch, S., Auinger,
S., Faix, J., Small, J.V., Polo, S., Stradal, T.E.B., & Rottner, K. (2009) Cortactin promotes
migration and platelet-derived growth factor-induced actin reorganization by signaling
to Rho-GTPases. Molecular Biology of the Cell 20(14), 3209-23.
Lai, F.P.L., Szczodrak, M., Block, J., Faix, J., Breitsprecher, D., Mannherz, H.G., Stradal,
T.E.B., Dunn, G.A., Small, J.V., & Rottner, K. (2008) Arp2/3-complex interactions and
actin network turnover in lamellipodia. EMBO Journal 27(7), 982-92.

SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis 79
01.11 Signalling to Actin Dynamics
PROJECT LEADER | Prof. Dr. Theresia E.B. Stradal | Former Research Group Signalling and Motility |
theresia.stradal@uni-muenster.de
PROJECT MEMBERS | Stefan Arens | Jan Hänisch | Kathrin Schloen | Kai Schlüter
The manipulation of actin cytoskeletal turnover is a
common theme in bacterial pathogenesis. Bacteria do this
to establish a niche for replication, to evade the immune
system by hiding inside non-phagocytic cells or by prohibiting
phagocytosis, to name only some. Manipulation of
actin dynamics is achieved directly by modifying actin, or
indirectly via regulators controlling actin dynamics such as
members of the Rho family of GTPases. This preference can
be explained by the crucial function of Rho-protein regulated
and actin dependent processes for innate and adaptive
immunity. Rho-GTPases serve as molecular switches that
- while cycling - turn actin remodelling on and off. Bacterial
cytotoxins affect all stages of this cycle, thereby activating
or inactivating Rho GTPases. For instance, reversible
shut-down comes from bacterial effectors with GAP-like
activities (e.g. SptP from Salmonella) and activation can
be mediated by factors with GEF-activity (e.g. SopE from
Salmonella; [1]). A novel family of bacterial virulence
factors termed WxxxE-family exerts bacterial GEF-activity,
similar to SopE, with which they also share structural
similarity. We were recently able to pinpoint residues
of the interaction surface between the Shigella flexneri
WxxxE-factor IpgB2 and human RhoA that are crucial for
bacterial GEF activity [2]. We currently identify interaction
networks around bacterial and cellular GEFs, GAPs
and their regulated GTPases.
One example of a more direct manipulation of actin
assembly constitutes pathogenic E. coli of the types EPEC
(enteropathogenic E. coli) and EHEC (enterohaemorrhagic E.
coli): Both of them firmly attach to the host cell surface and
induce actin polymerisation directly beneath the bacterium.
EPEC induce actin assembly by mimicking receptor
tyrosine kinase signalling, a mechanism that is also used
by pox viruses in a strikingly similar fashion [3]. EHECinduced-actin
assembly requires translocation of two bacterial
proteins: the translocated intimin receptor Tir, which
serves as attachment anchor traversing the host cell plasma
membrane, and EspF U
/TccP, activator of N-WASP, and thus
the host cell actin polymerization machinery is essential for
this process. Since Tir does not directly bind to EspF U
, additional
factors must bridge these components. In order to
elucidate this pathway, we analysed the composition of EspFu-based
protein complexes using mass spectrometry. We
could identify host cell IRSp53 as direct interactor of both
Tir and EspF U
. Moreover, IRSp53 co-localized with EspF U
in
actin pedestals, and loss of IRSp53-function abrogated actin
assembly into pedestals. Thus, we identified IRSp53 as the
missing factor linking bacterial Tir and EspF U
[4]. Current
work is focussed on elucidating the precise molecular
interactions between host cell and bacterial factors driving
pedestal formation, which will also include analyses of the
binding surfaces at atomic resolution (collaboration SB).
Murine embryonic fibroblasts expressing a constitutively active mutant of human RhoA, wildtype IpgB2 or its attenuated mutant Q116A
(adapted from Klink et al.,2010). Note that the virulence factor IpgB2 phenotypically induces RhoA activity . Photos: HZI
Hänisch, J., Ehinger, J., Ladwein, M., Rohde, M., Derivery, E., Bosse, T., Steffen, A.,
Bumann, D., Misselwitz, B., Hardt, W.-D., Gautreau, A., Stradal, T.E.B., & Rottner, K.
(2010) Molecular dissection of Salmonella-induced membrane ruffling versus invasion.
Cellular Microbiology 12(1), 84-98.
Klink,B.U., Barden,S., Heidler,T.V., Borchers,C., Ladwein,M., Stradal,T.B.*., Rottner,K., &
Heinz,D.W.*. (2010) Structure of Shigella IpgB2 in complex with human RhoA: implications
for the mechanism of bacterial guanine nucleotide exchange factor mimicry. Journal
of Biological Chemistry 285, 17197-17208.
Rottner,K. & Stradal,T.E.*. (2009) Poxviruses taking a ride on actin: new users of known
hardware. Cell Host and Microbe 6, 497-499.

80 SCIENTIFIC REPORTS | Infection and Immunity | Microbial Pathogenesis
01.12 Generation and Exploitation of DNA Sequence Data
PROJECT LEADER | Dr. Helmut Blöcker | Department of Genome Analysis | bloecker@helmholtz-hzi.de
PROJECT MEMBERS | Cyril Alozieuwa | Dr. Sabin Bhuju | Dr. Igor Deyneko | Ayssar Elamin | Michael Jarek |
Yulia Kalybaeva | Dr. Gabriele Nordsiek | Ibrahim Sabra | Maren Scharfe | Prof. Dr. Mahavir Singh | Dr. Matthias Stehr |
Hanaa Wanas
Sequence analysis projects Sequencing and analysis of
primary DNA is one of the principal basic techniques in
modern biological research. Our work includes the comparative
sequence analysis of clinical isolates from pathogenic
organisms such as M. tuberculosis with special emphasis on
genes involved in virulence, persistence, antibiotic resistance
and host preference.
We analysed selected regions from the horse, pig and cattle
genomes, most of which are suspected to be disease-related.
Deep annotation of a number of bacterial genomes is complete.
Recently, we sequenced three more of the big myxobacterial
genomes.
The implementation of two so-called next generation DNA
sequencers (Illumina/Solexa) enables us to considerably
expand the genomic analyses, quantitatively as well as qualitatively
(Fig. 1.). Accordingly, several expression analyses
have been run on sequencers with samples from cow, badger,
mouse and man. To allow for RNAi screening with complex
libraries, especially negative selection screening, we established
deep sequencing and successfully analysed a rather
large amount of samples. A further focus was on ChIP-Seq
which is about interaction of proteins (for example transcription
factors) with DNA. We mostly investigated disease-relevant
human samples.
Mycogenomics Our goal is the development of new methods
for tuberculosis diagnosis and treatment. The focus is
primarily on the identification of clinical multiple-resistant
Mycobacterium tuberculosis strains and on the characterization
of virulence-associated proteins.
Tuberculosis kills approximately two million human beings
worldwide per year. A big problem is the growing issues of
multiple-resistant bacteria (MDR) or extremely resistant pathogens
(XDR) which can only be treated with so called Second-
Line Drugs. We develop new diagnostic tools for MDR-TB.
In the EU project Fast XDR-Detect we develop a colour-based
assay for a 96-well plate format to detect MDR tuberculosis.
For the development of this assay, the genomes of numerous
clinical Mycobacterium tuberculosis strains were sequenced
to identify resistance-associated mutations (Fig. 2, see page
68). The identified mutations are used as molecular markers
for the assay.
In the EU project FASTEST-TB we are looking for new mycobacterial
antigens (diagnostic protein markers) for rapid
tests in a 96-well plate format. So far, we have found about
Fig. 1. Distribution of DNA clusters to be sequenced. About two
years of technology development increased the yield of megabases
per sequencer to 1,500 percent. Photos: HZI
100 new candidate proteins that are currently evaluated.
The enzyme Antigen85A synthesizes the most abundant glycolipid
in the cell wall, TDM (trehalose 6,6’-Dimycolate; Cord
factor). Antigen85A is an important component in the cell
wall but can also be used as a vaccine. We have produced
the recombinant protein for a detailed study of the immunological
properties of the enzyme. We were first to report the
development of a non-radioactive assay, which was used to
determine the enzyme´s catalytic parameters. The new assay
is also suitable for high-throughput experiments and for
the screening of substance libraries in order to explore new
lead structures.
Novel bioinformatics technology We are exploring applications
of signal theory for the function-oriented analysis of
biomolecules. Our intent is to reveal similarities, homologies
and analogies based on considerations of their physico-chemical
properties and to confirm these findings by wet lab data.
The software “FeatureScan” is publicly available (http://genome.helmholtz-hzi/featurescan).
Our system is able to spot
property-dependent similarities where letter code-based systems
(A, C, T, G) fail. Thus, comparing promoters from man
and chimpanzee, our novel technology revealed functional
regions which otherwise would have been undiscovered.
Deyneko,I.V., Kalybaeva,Y.M., Kel,A.E., & Blöcker,H. (2010) Human-chimpanzee promoter
comparisons: Property-conserved evolution? Genomics 96, 129-133.
Von Groll A., Martin A., Stehr M., Singh M., Portaels F., da Silva P.E. & Palomino JC.
(2010) Fitness of Mycobacterium tuberculosis strains of the W-Beijing and Non-W-
Beijing genotype. PLoS One 5, e10191.
Adhikary,T., Kaddatz,K., Finkernagel,F., Schonbauer,A., Meissner,W., Scharfe,M.,
Jarek,M., Blöcker,H., Muller-Brusselbach,S., & Müller,R. (2011) Genomewide Analyses
Define Different Modes of Transcriptional Regulation by Peroxisome Proliferator-Activated
Receptor-beta/delta (PPARbeta/delta). PLoS ONE 6, e16344.

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81
02 Host Pathogen Interactions
TOPIC SPEAKER | Prof. Dr. Klaus Schughart | Department of Infection Genetics | kls@helmholtz-hzi.de
The severity of an infectious disease is determined by the intricate interactions between pathogen and host. Many studies have
been performed to identify specifi c virulence factors in individual pathogenic microbe species but our knowledge about important
factors in the host is still very limited. For example, genetic and environmental factors are crucially involved in the host response
and determine favorable or unfavorable disease outcomes. Also very little is known about bacterial communities in the host and
how they may infl uence the activities of pathogenic microbes or the host response. Furthermore, pathogens that are able to cross
the species barrier represent a constant threat to the human population. Here, not much is known about the molecular mechanisms
of host adaptation.
Therefore, the objective of this topic is to obtain a better understanding about the complexity of the interactions between the host
and and infectious pathogens. The different research projects aim to identify host factors which infl uence its susceptibility and
resistance to infections, to unravel mechanisms of cross-species transmission, and to characterize microbial communities and
how they may modulate the host response to pathogens. To achieve these goals, the topic will perform research projects in the
following areas.
Microbial communities in the host and their relevance for infectious diseases
In the oral cavity approximately 500 bacterial species have been identifi ed so far. Many of these bacteria are able to form biofi l m s ,
so called dental plaque. In order to form biofi lms, bacterial species must communicate with each other. We are, therefore, studying
the molecular basis of bacterial communication pathways and investigate possible ways to disrupt them.
In cystic fi brosis (CF) patients, Pseudomonas aeruginosa is the most dominant bacterial pathogen causing chronic lung disease.
However, P. aeruginosa isolates from patients revealed that they do not represent a single variant but rather different phenotypes.
And the presence of different phenotypes has been correlated with poor prognosis in CF patients. To better understand the underlying
biological and molecular mechanisms of phenotype changes, we will study the diversity of bacterial communities and isolates
in CF patients.
Biofi lms play an important role in the establishment of pathogenic bacterial colonistaion in the host. Very little is known so far
about pathogenic communities in biofi lms and the relationship between the biodiversity and pathogenicity in presumed sterile
environments of the host. We are thus determining the biodiversity in biofi lms from patient material like pacemakers, bone and
dental implants. With the knowledge which bacterial communities live at the different niches in the human body new approaches to
control or prevent biofilms and to optimize their biodiversity with regards to the control of pathogenic bacteria are being developed.
The nasal cavity and the gastro-intestinal tract harbor a huge number of bacterial organisms which may be advantageous or, in
some cases, pathogenic to the host. We are, therefore, aiming to defi ne the conditions which favor or exclude the colonization with
pathogenic bacteria. A special focus is the colonization of the nose with multi-resistant Staphylococcus aureus and its interaction
with other community members.

82 SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactives
The host response to viral and bacterial pathogens in experimental models of infection
Group A streptococci are prevalent human pathogens capable of causing a variety of diseases ranging from simple pharyngitis
to very severe infections, such as necrotizing fascitiis and streptococcal toxic shock syndrome. We have previously shown that
strains of mice from various genetic backgrounds differ markedly in their susceptibility to Streptococcus pyogenes. Using these
resistant and susceptible mouse strains, we will identify the immunological and molecular mechanisms controlling the resistance
or susceptibility to S. pyogenes and design new strategies to enhance resistance in susceptible hosts. Staphylococcus aureus is
one of the most important causes of life-threatening bacterial infections in Western countries.
The incidence of S. aureus infections has dramatically increased in the last decade, occurring in both, community-acquired and
hospital-acquired cases. Despite the substantial progress made in understanding the mechanism employed by S. aureus to persist
within the host, few or none studies have addressed the contribution of host factors to the establishment of S. aureus infection.
Research in this area has been hampered by the lack of feasible animal models that recapitulate the different aspects of S. aureus
infection. We have established an experimental mouse model that sustains a long-term S. aureus infection and, therefore, enables
us to dissect the mechanisms underlying the divergent functionality of the host immune response during the different phases of
infection.
Infl uenza A virus represents one of the most important threats to human health and economics. Thus far, very little is known about
host resistance or susceptibility to infl uenza infections. At the HZI, we have established an infection model for infl uenza A H1N1
virus in the mouse and identifi ed signifi cant strain differences in the LD 50
response. We perform comparative studies on the pathogenicity,
gene expression patterns in the lung, viral load, and immune response in infected mice from different strains. This information
will be used to map quantitative trait loci (QTL) that contribute to infection susceptibility. Potential candidate genes will
then be tested by phenotyping combinations of single gene mouse mutants (knock-out, gene trap, classical or ENU-induced) in
various genetic backgrounds.
Molecular specificity of interspecies and zoonotic pathogen transmissions
The mammalian prion protein, PrP C , is able to switch its conformation from a monomeric soluble form into an aggregated transmissible
prion state, PrP Sc . Once a prion is introduced into a susceptible host, it triggers a PrP conversion cascade which leads to
prion disease. Individual prion strains may lead to distinct prion disease pathologies in one host species. Interspecies transmission
will be strongly infl uenced by the extent to which a particular PrPSc conformation is compatible with the primary PrP amino acid
sequences of both species. The zoonotic transmission of BSE to humans, and the resistance of humans towards sheep scrapie
are well known examples of this process. However, the underlying molecular mechanism of conformational conversion and prion
propagation is not at all understood. A major obstacle in the research on prions was the limited availability of samples for prion
3D structure investigations. At the HZI we have established a novel technology to overcome this bottleneck. In particular NMR
spectro scopic techniques and other biophysical techniques are used to unravel the structural origin of prion transmission barriers

SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactives
83
and prion adaptations to new hosts. The 3D structures of mammalian prion associated PrP will be revealed and should provide
detailed insight into biophysical mechanisms of the PrP C to PrP Sc conformation conversion.On a broader scope, we have a general
interest in the molecular structural investigation of specifi c host:pathogen protein interactions that modulate the interspecies or
zoonotic transmissibility of viruses and bacteria.
Epidemiological studies in humans to determine environmental and genetic risk factors
The causes of human diseases are multi-factorial and consist of environmental, life style, and genetic risk factors. To better assess
these factors, we will study the incidence of infectious diseases in humans and determine their genetic and environmental risk
factors. To this end, the HZI and other centres of the Helmholtz Association as well as many university partners have initiated a
nationwide population-based prospective cohort study, the National (Helmholtz) Cohort. This cohort will contain about 200,000
healthy volunteers from all over Germany. We will administer a questionnaire on acute and chronic infections, vaccination status,
and exposure to pet and farm animals. In addition, swabs from the nasal cavity, saliva and feces will be collected and analysed
for their microbial communities. The participants of the cohort will be followed for 10-20 years, and we will seek to determine the
risk factors which contribute to infectious diseases and to secondary diseases that may be directly or indirectly associated with
infections.

84
SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions
02.1 Pathogenesis of Chronic Pseudomonas aeruginosa
Infections
PROJECT LEADER | Prof. Dr. Susanne Häußler | Research Group Chronic Pseudomonas Infections |
sus@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Andreas Dötsch | Dr. Mathias Müsken | Juliane Schmidt | Sebastian Schulz | Dr. Piotr Bielecki
The diagnostic and therapeutic strategies that have served
us well in the treatment of acute bacterial diseases have
not yielded favourable outcomes when applied to chronic
infections where bacteria grow in matrix-enclosed sessile
biofilm communities. Although every single cell is able to
induce a stress response with a characteristic change in
the gene expression pattern, living in populations provides
a species with additional mechanisms of survival, the
most obvious being heterogeneity and cooperation.
Diversity facilitates survival Pseudomonas aeruginosa is
the most dominant bacterial pathogen causing chronic
lung infection in cystic fibrosis (CF) patients. Although
most patients are colonized with only one or few P. aeruginosa
clones, the isolation of various morphotypes is a
very characteristic microbiological finding. This diversity
seems to play a key role in the persistence of chronic lung
infections. Our research focuses on the elucidation of the
molecular mechanisms responsible for this diversity and
the characterization of particularly well adapted P. aeruginosa
biofilm phenotypes.
Small Colony Variants We have previously demonstrated
that an adherent subgroup of so-called small colony
variants (SCVs) is selected in the CF lung. The switch to
an autoaggregative biofilm-forming SCV phenotype of
P. aeruginosa seems to be linked to the expression of the
“Chaperone Usher Pathway” (CupA) gene cluster. This gene
cluster encodes for bacterial fimbriae and is regulated via
the modulation of the newly identified bacterial signal
molecule, cyclic di-GMP, in an antagonizing way. With the
aim to identify those mutations that are responsible for
the switch to a biofilm forming P. aeruginosa phenotype
as well as the acquisition of antibiotic resistances, we will
apply a whole genome sequencing approach. Therefore, the
genomes of pools of clinical P. aeruginosa strains will be
sequenced using next generation sequencing technologies
and we will search for consistent nucleotide exchanges
in sets of strains which express a similar phenotype. The
next step will be to verify whether the respective mutation
is responsible for the biofilm or the resistance phenotype.
The knowledge about the genotypes that are specifically
selected at different stages of the disease should significantly
advance our knowledge on the evolution and
adaptation mechanisms of P. aeruginosa to its habitat. It
might aid in the development of new antimicrobial treatment
regimes for the clinical management of chronically
infected CF patients.
Cooperation is supported by communication Apart from
two homoserine lactones, P. aeruginosa produces a third
intercellular signal that is referred to as the Pseudomonas
quinolone signal (PQS). It is involved in cell densitydependent
virulence factor regulation – also known as
quorum sensing (QS) – and the establishment of biofilms.
However, the molecular mechanism underlying downstream
PQS signalling is largely unknown. PqsE, encoded
on the last gene of the PQS biosynthetic operon, seems to
play a key role in the translation of the presence of the
PQS signal and bacterial behaviour at the single cell level.
The elucidation of the function of this important enzyme is
a major focus of the group.
Susanne Häußler preparing for a new experiment in the lab.
Photo: Twincore/HZI
Pommerenke C, Müsken M, Becker T, Dötsch A, Klawonn F, Häussler S. (2010). Genotype-
Phenotype correlation in Pseudomonas aeruginosa. PLoS Pathogens. 6(8): e1001074.
Müsken M, Di Fiore S, Römling U, Häussler S. (2010) 96-well plate based optical method for
the quantitative and qualitative evaluation of Pseudomonas aeruginosa biofilm formation
and its application for susceptibility testing. Nature Protocols 5(8):1460-9. Epub 2010 Jul 29.
Dötsch, A., F. Klawonn, M. Jarek, M. Scharfe, H. Blöcker, S. Häussler. (2010). Evolutionary
conservation of essential and highly expressed genes in Pseudomonas aeruginosa. BMC
Genomics. 11(1):234.
Müsken M, Di Fiore S, Dötsch A, Fischer R, Häussler S. (2010) Genetic determinants of
Pseudomnoas aeruginosa Biofilm establishment. Microbiology. 156:431-41
Häussler S. (2010) Multicellular signalling and grwoth of Pseudomonas aeruginosa. International
Journal of Medical Microbiology. 300(8):544-8.
Häussler S, Parsek MR. Biofilms 2009: (2010) new perspectives at the heart of surfaceassociated
microbial communities. Journal of Bacteriology. 192(12):2941-9.

SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions 85
02.2 Unravelling Mechanisms of Host Defence against
Gram-positive Pathogens in the Mouse Model
PROJECT LEADER | Priv.-Doz. Dr. Eva Medina | Research Group Infection Immunology | eme@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Oliver Goldmann | Dr. Jens Abel | Christina Ziegler | Daniela Bruhn | Alva Rosendahl
Staphylococcus aureus and Streptococcus pyogenes are major
human pathogens. The focus of the Infection Immunology
Research group is to unravel the immune mechanisms
involved in host defense against these pathogens.
The mouse model of genetic predisposition to severe
Staphylococcus aureus infection A prominent feature of
S. aureus infections is their inter-individual phenotypic variability,
ranging from mild to lethal clinical manifestations.
Common genetically determined variations in the immune
system may contribute to the observed heterogeneity in
the host response to this pathogen. We have examined the
immune mechanisms underlying the host genetic predisposition
to severe S. aureus infection in the murine system.
Our results confirmed a significant inter-strain variation in
resistance to this pathogen. Thus, whereas C57BL/6 mice
were the most resistant and survived infection, A/J, DBA/2
and BALB/c mice were highly susceptible and succumbed
to infection shortly after bacterial inoculation. Susceptibility
of mice to S. aureus was associated with an inability to
limit bacterial growth in the kidneys and development of
pathology (Figure 1).
Depletion of neutrophils rendered resistant C57BL/6 mice
completely susceptible to S. aureus indicating that neutrophils
are essential for the observed resistance. We also
found that a delay in neutrophil recruitment to the site of
infection may underlie the increased susceptibility of A/J
mice to S. aureus. Therefore, the superior resistance of
C57BL/6 mice is likely to be the result of a fast recruitment
of effector cells to the site of infection that bring straight
away under control the infectious pathogen.
Role of TLR/MyD88 signalling in the pathophysiology of
experimental Streptococcus pyogenes infection
Streptococcus pyogenes represents one of the most frequent
causes of mild infection of the upper respiratory tract and
the skin with the potential to cause life-threatening invasive
diseases such as necrotizing fasciitis and septicemia. Based
on our previous in vitro studies, the myeloid differentiation
factor 88 (MyD88) was found to be involved in the response
of innate immune cells to S. pyogenes. To determine the
relevance of these in vitro observations to the pathogenesis
of S. pyogenes in vivo, we have investigated the impact of
MyD88 deficiency on host resistance to S. pyogenes in a
mouse model of infection. Our results show that MyD88 -/-
mice harbored significantly more bacteria in the organs and
succumbed to infection much earlier than MyD88 +/+ animals.
Absence of MyD88 resulted in diminished production of inflammatory
cytokines such as IL-12, IFN-γ, and TNF-α as well
as chemoattractants such as MIP-2 and KC and hampered
recruitment of effector cells involved in bacterial clearance
Bacterial loads in resistant C57BL/6 (mouse 2) and susceptible
A/J mice (mouse 3) infected for 24 h with the bioluminescent
S. aureus strain SH1000 ALC2906. Uninfected C57BL/6
(mouse 1) and A/J (mouse 4) are included for comparison.
(macrophages and neutrophils) to the infection site. Furthermore,
MyD88 -/- but not C57BL/6 mice exhibited a massive
infiltration of eosinophils in infected organs (Figure 2) which
can be explained by an impaired production of the regulatory
chemokines, MIG/CXCL9 and IP-10/CXCL10, shown to
be able to inhibit transmigration of eosinophils. Our results
indicate that MyD88 signalling targets effector cells to the
site of streptococcal infection and prevents extravasation
of cells that can induce tissue damage. Therefore, MyD88
signalling may be important for shaping the quality of the
inflammatory response elicited during infection to ensure
optimal effector-functions.
H&E stained lung tissue sections obtained from S. pyogenesinfected
MyD88 -/- mice. Left, low magnification photograph
showing a massive cellular infiltration. Right, a higher
magnification photograph showing that the infiltrating cells
are eosinophils.
von Köckritz-Blickwede M., Rohde M., Oehmcke S., Miller L.S., Cheung A.L., Herwald H.,
Foster S., and Medina E. (2008). Immunological mechanisms underlying the genetic predisposition
to severe Staphylococcus aureus infection in the mouse model. American Journal of
Pathology. 173:1657-68.
von Köckritz-Blickwede M., Konrad S., Foster S., Gessner J.E., and Medina E. (2009). Protective
role of complement C5a in an experimental model of Staphylococcus aureus bacteremia.
Journal of Innate Immunity. 2:87 – 92.
Loof T., Goldmann O., Gessner A., Herwald H., and Medina E. (2010). Aberrant Inflammatory
Response to Streptococcus pyogenes in Mice Lacking Myeloid Differentiation Factor 88.
American Journal of Pathology. 176: 754-63.

86
SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions
02.3 Microbial Communication
PROJECT LEADER | Prof. Dr. Irene Wagner-Döbler | Research Group Microbial Comunication | iwd@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Helena Sztajer | Dr. Brigitte Kunze | Dr. Michael Reck | Andre Lemme | Xiaoli Xue |
Ina Buchholz | Jürgen Tomasch | Thomas Riedel | Diana Patzelt | Szymon Safranski | Hui Wang
Bacteria synthesize signalling molecules, so-called autoinducers,
which are excreted into the medium and control
important physiological traits in a concentration dependent
way. Cells sense autoinducer concentrations by specific
receptors in the membrane or within the cytoplasm. Above
a threshold concentration, the so-called quorum, the signalling
molecules induce, for example, genetic competence
in the caries bacterium Streptococcus mutans, resulting in
increased genetic exchange and biofilm formation. Therefore,
this type of cell-cell communication is termed quorum
sensing. It plays an important role for infections and symbioses.
Understanding the mechanisms of quorum sensing
opens up new possibilities for influencing infections and
developing alternatives for antibiotics.
Interkingdom signalling: The caries bacterium Streptococcus
mutans inhibits hyphae formation of the human
pathogenic fungus Candida albicans Streptococcus
mutans lives in the polymicrobial biofilm on our teeth and
is strongly associated with caries. Among hundreds of other
species of bacteria also fungi occur in the oral cavity, e.g.
the human pathogen Candida albicans, which can cause bad
infections in immuno-compromised patients, after antibiotic
therapy or in the elderly. When studying signalling
molecules of S. mutans we discovered a fatty acid compound,
SDSF (Streptococcus diffusible signal factor). It suppresses
the formation of hyphae of the fungus, which are crucial
for virulence and biofilm formation. SDSF is structurally
related to farnesol, the quorum sensing signal of C. albicans,
so this is a case of quorum sensing mimicry across huge
phylogenetic distance, e.g. between the kingdom of bacteria
and the kingdom of fungi.
Biofilm inhibitors We have identified small chemical
molecules which interfere with microbial communication.
Carolacton, which is produced by the soil bacterium Sorangium
cellulosum, is particularly interesting because it has
no antibacterial or cytotoxic activity, but inhibits selectively
biofilm formation of Streptococcus mutans. Carolacton
causes membrane damage and cell death and thus disturbs
biofilm formation. Microarray investigations suggest that
several of the two component systems of S. mutans, which
are its main sensory systems for environmental adaptation
and quorum sensing, are impaired by carolacton. Details of
this mechanism and possible applications of carolacton for
caries prevention are currently studied in a systems biology
joint project.
Quorum sensing in the Roseobacter group Bacteria of
the Roseobacter group are important for biogeochemical
cycles in the ocean, because they comprise up to 25 % of all
bacterial cells in marine habitats. Dinoroseobacter shibae,
one of the model organisms in the Roseobacter Transregio,
lives in symbiosis with unicellular marine algae. D. shibae
provides the algae with the essential vitamins B12 (cobalamin)
and B1 (thiamine), and receives sugar compounds
for its heterotrophic metabolism in turn. Currently, we are
studying the role of quorum sensing for this symbiosis, as
well as the contribution of photosynthesis to the metabolism
of the bacterium, which in the light gains energy by
photophosphorylation, allowing it to burn less carbon and
thus “save” CO 2
.
Dinoroseobacter shibae on its host algae, the marine
dinoflagellate Prorocentrum lima. The bacteria have been
specifically stained by CARD-FISH (catalyzed reporter
deposition fluorescent in situ hybridisation). The remaining
bacterial community is invisible under these conditions. The
orange spot appearing in the left dinoflagellate is the socalled
pyrenoid, a region of the plastid which contains very
high concentrations of the rubisco enzyme. Photo: HZI
Xue, X., Tomasch, J., Sztajer, H., and I. Wagner-Döbler (2010). The delta subunit of RNA
polymerase, RpoE, is a global modulator of Streptococcus mutans environmental adaptation.
Journal of Bacteriology. 192:5081-92.
Kunze B, Reck M, Dötsch A, Lemme A, Schummer D, Irschik H, Steinmetz H, Wagner-Döbler
I. (2010) Damage of Streptococcus mutans biofilms by carolacton, a secondary metabolite
from the myxobacterium Sorangium cellulosum. BMC Microbiology. 10:199.
Wagner-Döbler, I., Ballhausen B, Baumgart M, Brinkhoff T, Buchholz I, Bunk B, Cypionka
H, Daniel R, Drepper T, Gerdts G, Hahnke S, Han C, Jahn D, Kalhoefer D, Kiss H, Klenk H-P,
Kyrpides N, Liebl W, Liesegang H, Meincke L, Pati A, Petersen J, Piekarski T, Pommerenke
C, Pradella S, Pukall R, Rabus R, Stackebrandt E, Thole S, Thompson L, Tielen P, Tomasch
J, von Jan M, Wanphrut N, Wichels A, Zech H, Simon M. (2010) The complete genome
sequence of the algal symbiont Dinoroseobacter shibae – a hitchhiker´s guide to life in the
sea. ISME Journal. 4(1):61-77.

SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions 87
02.4 Systems Genetics of Infection and Immunity
PROJECT LEADER | Prof. Dr. Klaus Schughart | Department of Infection Genetics | kls@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Rudi Alberts | Dr. Mahmoud Bahgat | Leonie Dengler | Dr. Feng He | Dr. Bastian Hatesuer |
Dr. Berenike Henneberg | Dr. Heike Kollmus | Dr. Tatiana Nedelko | Dr. Bastian Pasche | Dr. Claudia Pommerenke |
Dr. Nuno Viegas | Dr. Esther Wilk | Silke Bergmann | Barkha Bhatnagar | Paulina Blazéjewska | Hairong Chen | Haiya Wu
Our research group is identifying genetic factors which
contribute to the susceptibility or resistance of the host
to influenza A infections. For these studies, we are using
mouse genetic reference populations consisting of mouse families
with different genetic backgrounds as well as mouse
mutants which carry deletions in specific gene loci.
Every year, about 10,000 to 30,000 people die from influenza
infections in Germany. The most severe pandemic in
recent human history, in 1918, has claimed about 30-50
Million deaths world-wide. The influenza virus is able to
change its antigenic properties very rapidly. In addition, the
viral genome is segmented and these segments can be exchanged
between different virus subtypes in other species.
Such re-assorted viruses may then adapt to the human host
and cause new pandemics.
It has long been suspected that genetic factors may play an
important role for resistance or susceptibility to influenza infections
in humans. However, such studies are very difficult
to perform in humans and are often blurred by environmental
factors, e.g. other serious health conditions, nutrition or
medication. Therefore, we have established an experimental
model in which we infect mice with different mouse-adapted
influenza A viruses. In particular, we are studying mouse
families derived from laboratory-inbred mouse strains, families
in which two or more parental genomes are segregating
and families in which parts of the genome were derived from
wild mice and introduced into the genomic background of
a laboratory strain. In addition, we are analysing the role of
individual genes for the host defense to influenza infection
by studying mouse lines in which a single gene locus has
been mutated by genetic engineering.
So far, we demonstrated a very strong influence of the genetic
background on host susceptibility in mice. One mouse
family, the DBA/2J inbred strain, was highly susceptible to
the PR8 H1N1 virus whereas another family, the C57BL/6J
inbred strain, was very resistant. In the susceptible mouse
strain the influenza virus replicated much faster and the
susceptible strain also exhibited a very strong innate immune
response. Our results suggest that the high viral load
in the lungs and the hyper-inflammatory response of the
immune system are responsible for the fatal outcome of the
infection in the susceptible strain. In addition, we showed in
mouse mutants that the Rag2 and the Irf7 genes are important
for the host to survive an infection with influenza virus.
We are now performing genome-wide gene expression
analyses over the course of an infection to better understand
(A) Weight loss and late death in mice carrying a mutation in
the Rag2 gene (A). The Rag2 knock-out mutation results in
the absence of T and B cells of the adaptive immune response.
Mice survive the early phase of infection because the innate
immune system can control virus replication to a certain extent,
but infected mice succumb later because the virus cannot
be finally cleared from their lungs. (B) Histological sections
of mouse lung tissue infected with influenza A virus H1N1.
The brown staining detects the presence of the influenza A
virus Nuclear Protein in Rag2 mutant mice at 10 days after
infection whereas wild type mice (C57BL/6J) have cleared the
virus (reproduced from Wu et al., 2010).
the host-pathogen interactions at the molecular level and to
relate changes in gene expression with cellular responses of
the immune system and pathologies in infected lungs. Our
research activities are integrated into several national and
international networks. We co-ordinate GeNeSys (German
Network for Systems Genetics) and SYSGENET (European
network for systems genetics to understand human diseases),
and we participate in FluResearchNet (German influenza
research network), Infrafrontier (European network to
establish mouse phenotyping and archiving infrastructures),
CASIMIR (European network for mouse database integration)
and the CTC (world-wide Complex Trait Consortium).
P. Blazejewska, L. Koscinski, N. Viegas; D. Anhlan; S. Ludwig; K. Schughart, (2011). Pathogenicity
of different PR8 influenza A virus variants in mice is determined by both viral and
host factors. Virology Journal, 412(1):36-45
M. Bahgat, P. Błazejewska, and K. Schughart. Inhibition of lung serine proteases in mice: a
new approach to control influenza infection. Virology Journal, 8:27
B. Srivastava, P. Blazejewska, M. Heßmann, D. Bruder, R. Geffers, S. Mauel, A. D. Gruber, K.
Schughart. (2009) Host genetic background strongly influences the response to influenza A
virus infections. PLoSONE, 4, e4857.

88
SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions
02.5 The Structural Basis of Mammalian Prion Transmission
Barriers
PROJECT LEADER | Dr. Thorsten Lührs | Junior Research Group of Structure-Based Infection Biology |
tlu07@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Felix Deluweit | Dr. Vandana Gupta
The mammalian prion protein, PrP C , is able to switch its conformation
from a monomeric soluble form into an aggregated
transmissible prion state, PrP Sc . Once a prion is introduced
into a susceptible host, it triggers a PrP conversion cascade,
which leads to prion disease (Fig. 1A). Interspecies transmission
barriers have been observed to correlate with the amino
acid sequence of PrP. On the other hand, multiple prion
strains of the same PrP amino acid sequence have been
identified that cause characteristic pathologies in one host
species. We aim to investigate prion 3D structures in order to
understand the transmission barrier between mice and hamsters
at atomic resolution, and to understand the structural
basis of prion strains in relation to prion host range. Therefore,
we have established a novel technology for the in vitro
generation of specific aggregated prion conformations. We
employ a combination of solution NMR, solid state NMR, and
other biophysical or chemical techniques in order to obtain
meaningful structural information of these aggregates.
Fig. 1: Progress on Prion in vitro Amplification. (A) The
benign cellular prion protein, PrPC, is an abundant
mammalian protein, which may undergo conformational
conversion and aggregation into infectious prions, PrPSc. (B)
We have developed a novel method, that employs specific
shear forces (left) for the efficient in vitro amplification
of prions. A device, the Shear Induced Fragmentation /
Aggregation (ShIF/A) array, has been developed by our group
to efficiently optimize many conversion reaction in parallel.
(C) Using ShIF/A we can convert recombinant PrPC into
patially proteinase K (PK) resistant PrPSc aggregates, using
natural hamtser Sc237 prions as seeds (top). In unseeded
reactions no PK resistance is observed (bottom).For optimal
prion amplification, the rotation frequency (Hz) of the rotor
has to be controlled within narrow boundaries.
Transmission barriers Between different mammalian species
usually exists a so-called ‘species barrier’: mice do not
develop prion disease after challenge with hamster prions.
Transgenic mice that express additionally hamster PrP can
be infected with hamster prions and then selectively incorporate
hamster PrP into prion particles. The exclusive expression
of a mouse/hamster chimeric PrP, termed MH2M,
makes mice susceptible to both hamster and mouse prions,
and the resulting prions transmit disease to both wild-type
hamsters and wild-type mice. Thus, the species barrier is in
part determined by the primary amino acid sequence of the
prion protein, and the substitution of only a few amino acid
residues can completely alter prion transmission patterns.
This notion has gained significant importance because
human variant Creutzfeld-Jacob Disease has been causally
linked to the exposure to BSE prions of cattle.
Prion sample preparation and structural analysis A key
technique in achieving our research goals is the conversion
of isotopically labelled ( 2 H, 13 C and 15 N) recombinant PrP
into PrP Sc . In order to obtain homogeneous particle preparations,
we have invented a novel technique in 2010 (Fig.1B/C).
Such aggregates are further characterized by asymmetric
flow-field-flow fractionation (AF4), which has recently been
introduced into amyloid research. This chromatography-like
technique is capable of separating particles by size without
the need for a stationary matrix. At the same time the absolute
hydrodynamic properties of individual particle fractions
are determined by multi-angle and dynamic light scattering.
This technique is also routinely used to characterise the
oligomerisation status of other proteins. The core techniques
for the protein structure determinations are NMR in solution
and in solids, but also other biophysical techniques like
fluorescence, FTIR and CD-spectroscopy. The strength of
this approach lies in its robustness towards subtle structural
heterogeneity of the investigated aggregates. Thus, with this
a “consensus structure” can be obtained to provide essential
structural insights.

SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions 89
02.6 Biofilm Communities
PROJECT LEADER | Dr. Wolf-Rainer Abraham | Research Group Chemical Microbiology | wab@helmholtz-hzi.de
PROJECT MEMBERS | Andreía B. Estrela | Ahmed S. Gebreil | Dr. Marcela G. Heck | Rolf Kramer |
Dr. Heinrich Lünsdorf | Maira Peres de Cavalho | Esther Surges
This project understands microbial communities in biofilms
as functional units and pursues the goal of finding new
ways to their control by investigation of the diversity of microbes
and their interactions. The focus is on the fact that in
nature bacteria mostly live not as pure strains but in communities.
It is of special interest how pathogenic bacteria
live in their hosts and how they deal with non- or facultative
pathogenic bacteria in biofilms in the human body.
Formation of dental biofilms One of the main complications
with implants in the hospital is the infection which is
mostly accompanied by biofilm formation on the implant.
Such biofilms are difficult to treat since the bacteria in
biofilms exhibit special protective mechanisms against antibiotics
and the immune system. Problems arise in particular
in the oral cavity which harbours by nature a multitude
of different bacteria. We wanted to know which bacteria
settle first on implants in the oral cavity and, therefore, we
examined two weeks old biofilms of tooth implants from
different patients. We always found complex communities
of bacteria which strongly varied from patient to patient. By
statistic analysis we could, however, show that there are
some characteristic species of bacteria which were found
at certain sites of the implant again and again and thus
are characteristic for these sites. Interestingly enough, we
found at the implants of the same patient more frequently
Streptococci than on the natural teeth, however, no amassment
of pathogenic bacteria was observed.
Metabolic activities in biofilms The knowledge which
bacteria in which biofilms occur at which places is, however,
only the first step in order to understand the complex
phenomenon biofilm. We must also examine the role of the
individual bacteria in the biofilm community. One approach
is the question about substrate use. In order to clarify how
quickly and how strongly a substrate is incorporated by
bacteria we use substrates which are labelled with stable
isotopes, thus are not radioactive, and follow their incorporation
into the individual types of bacteria. In this way,
we can determine kinetics and model the flow of material
in the community. Since we are not only interested in the
interactions of the bacteria among themselves but also in
host-bacteria interactions we also began to accomplish such
studies in cell cultures and animal models. As the materials
are not radioactive no special safety precautions are necessary.
With such models we can show that the same bacteria
have different metabolic pathways in different organs, and
in one organ e.g. glucose is directly taken up while in a
tumour bacteria nourish themselves preferentially of its
metabolites.
Biofilm community recorded by confocal laser microscopy.
The cells were stained with Nile Red and slightly hydrophobic
bacteria cells (red) can be discerned from highly hydrophobic
ones (green) Photo: Ahmed Gebreil/ Maximilliano Gutierrez
Natural products to control biofilms The aim of all of
these investigations is a better understanding of biofilm
communities with the goal of modulating them and of
displacing pathogenic bacteria. We use well-known natural
substances but we also look for new ones in mushrooms. A
first investigation of fungal isolates revealed that many of
them produce a number of quite diverse natural products
which are able to dissolve biofilms of pathogenic bacteria
without, however, being antibiotic. These investigations are
still at the beginning; however we are already able to state
that mushrooms are a rich source for substances which can
modulate biofilms.
A. B. Estrela, M. G. Heck, W.-R. Abraham (2009) Novel approaches to control biofilm infections.
Curr. Med. Chem., 16, 1512-1530.
A. B. Estrela, W.-R. Abraham (2010) Combining biofilm controlling compounds and antibiotics
as a promising new way to control biofilm infections. Pharmaceuticals, 3, 1374-1393.
W. Heuer, M. Stiesch, W. R. Abraham (2010) Microbial diversity of supra- and
subgingival biofilms on freshly colonized titanium implant abutments in the human mouth.
European Journal of Clinical and Microbiological Infection Diseases,
http://dx.doi.org/10.1007/s10096-010-1068-y

90
SCIENTIFIC REPORTS | Infection and Immunity | Host Pathogen Interactions
02.7 Metabolic Diversity
PROJECT LEADER | Prof. Dr. Dietmar Pieper | Research Group Microbial Interactions and Processes |
dpi@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Ramiro Vilchez | Dr. Melissa Wos | Macarena Marin | Daiana Morales | Amelia Silva
Microorganisms in their natural habitats typically live in
complex communities where the composition and functioning
is strongly influenced by the environmental conditions.
The understanding of complex community functioning with
the goal of their rational manipulation and exploitation
is not only important in environmental habitats such as
marine or terrestrial ecosystems but also in other habitats
like the human body which is also colonised by bacterial
communities in scales that outnumber human cells. While
these host-associated microbes may be beneficial for human
health, the human body can also be a reservoir for opportunistic
pathogens.
Nasal communities As an example, the human anterior
nares are the major source and risk factor for invasive
infections by Staphylococcus aureus, an increasingly multiresistant
pathogen causing a large spectrum of infectious
diseases. However, very little is known about the composition
of the nasal microbiota and possible community interactions.
We have now elucidated the community composition
of 40 individuals using advanced culture-independent
approaches and have shown the presence of previously not
expected dominant community members such as Finegoldia
magna, Dolosigranulum pigrum, Anaerococcus sp. or
uncultured actinomycetes. We also evaluated interactions
between S. aureus and other microbial species present in
the anterior nares and, as an example, observed a statistically
significant converse association between S. aureus and
F. magna, indicating that these species have a substantially
different distribution and abundance pattern reflecting
their inability to share the same niche. The previously
undiscovered co-colonization patterns and natural variations
in species composition provide insights for future
intervention strategies for the control of health care- and
community-associated S. aureus infections.
Intestinal communities: The structure of the human gut
microbial community (microbiome) is determined by host
genetics and environmental factors, where alterations in
its structure have been associated with the onset of many
different diseases. Establishing a defined human gut
microbiome within inbred rodent models provides a means
to study microbiome-related pathologies. We now compared
the efficacy, quality and stability of the human microbiome
established within germ-free (GF) rats, GF mice, and antibiotic-treated
mice. Remarkable differences were observed
between the different rodent models. While the majority of
abundant human-donor bacteria were established in the GF
rats, only a subset was present in the GF mice. Specifically
members of Clostridia cluster IV, which comprise important
butyrate producers contributing to processes important for
colonic health, failed to colonise the mouse gut. Our results
indicate that the genetic background of the recipient system
strongly influences the efficacy and quality of the populating
human gut microbiota and clarifies the suitability of
the models.
Environmental communities and new metabolic pathways
We have also improved our analytical tools to study microbial
communities in contaminated environments whereby
using a novel validated catabolic gene array dedicated to
achieve fast monitoring of catabolic gene diversity and
abundance and thus the catabolic landscape in combination
with metagenomic screening approaches, structure (on the
DNA level) and function (by analyzing mRNA) could reliably
be described. These culture-independent analyses were
complemented by the elucidation of novel metabolic routes
and enzymes.
Scheithauer BK, Wos-Oxley ML, Ferslev B, Jablonowski H, Pieper DH. (2009). Characterization
of the complex bacterial communities colonizing biliary stents reveals a host-dependent
diversity. ISME Journal 3:797-807.
Wos-Oxley M, Plumeier I, von Eiff C, Taudien S, Platzer M, Vilchez-Vargas R, Becker K and
Pieper DH. (2010). A poke into the diversity and associations within human anterior nare
microbial communities. ISME Journal 4:839-851.
Brennerova MV, Josefiova J, Brenner V, Pieper DH, Junca H. (2009). Metagenomics reveals
diversity and abundance of meta-cleavage pathways in microbial communities from soil
highly contaminated with jet fuel under air sparging bioremediation. Environmental
Microbiology 11:2216-2227.
Vilchez-Vargas R, Junca H, Pieper DH. (2010). Metabolic networks, microbial ecology and
‚omics‘ technologies: towards understanding in situ biodegradation processes. Environmental
Microbiology 12:3089-3110.

SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity
91
03 Inflammation and Immunity
TOPIC SPEAKER | Prof. Dr. Hansjörg Hauser | Department of Gene Regulation and Differentiation |
hha@helmholtz-hzi.de
The hosts’ immune response against invading pathogens can lead to massive infl ammatory reactions which might result in
immunopathology, thereby causing severe problems to the host. Thus, understanding the mechanisms how pathogens’ actions on
the one hand induce protective immune responses and on the other hand cause tissue damage is the key for the development of
protective and therapeutic measures.
The topic “Infl ammation and Immunity” deals with pathogen-induced reactions, including innate immune responses, infl ammatory
responses, and induction of long-term immunity. While these aspects are studied with the aim of understanding basic principles,
the topic also aims at defi ning novel drug targets and therapeutic intervention strategies.
Induction of interferons and their activities
The recognition of pathogen-associated molecular patterns by various receptors leads to signaling events, which finally result in the
induction of interferon (IFN) genes. Upon secretion, IFNs have diverse activities on their target cells. The topic aims at identifying
the involved molecular players, understanding their interaction with viral proteins and describing the dynamics of IFN-induced
signal transduction in different cell types.
Consequences of inflammation
In most cases, infl ammatory events have benefi cial outcomes to the organism. Post-infl ammatory actions involve apoptosis and
autophagy, which are of central importance to eliminate infected or damaged cells. In this topic, the involved basic mechanisms
are studied as such, but also with regard to pathogen-mediated modulation of these processes. In cases of chronic infl ammatory
reactions, e.g. in rheumatoid arthritis (RA), the dysregulated immune responses have negative outcomes. An important translational
aspect within this topic concerns the suppression of chronic infl ammation in murine models of RA by inhibition of a central
signalling molecule in proinfl ammatory T cells.
T cells in autoimmunity and tolerance
Effector T cells play a critical role for the elimination of different types of pathogens from the infected host. However, if directed
against self tissues, they also can drive autoimmune reactions, thereby causing severe damage to the host. To prevent such selfdestructive
immune reactions, the immune system has evolved a peculiar T cell subpopulation, the reglatory T cells, which play an
important role in the maintenance of self tolerance. Within this topic, the generation of diverse T cell subsets and their functional
properties are studied in detail in infection and infl ammatory mouse models.
Systems Biology: Theoretical models for complex immune responses
In the past, researchers have been mainly focussing on the elucidation of linear pathways of biological events. With the development
of high-throughput techniques and with the acquisition of complex data sets it has become more and more diffi cult to translate
this huge amount of information into comprehensive concepts. In this topic, we use mathematic modeling approaches to
describe dynamic and complex immunological processes such as IFN signalling and B and T cell homeostasis.

92 SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity
03.1 Development and Functional Properties of
Foxp3+ Regulatory T Cells
PROJECT LEADER | Prof. Dr. Jochen Hühn | Department of Experimental Immunology | jhu08@helmholtz-hzi.de
PROJECT MEMBERS | Pedro Almeida | Devesha Bhagwat | Sascha Cording | Dr. Stefan Flöß | Larissa Fournier |
Yi-Ju Huang | Dr. Katjana Klages | Dr. Julia Polansky | Lisa Schreiber | Dr. René Teich | Aras Toker
Regulatory T cells (Tregs) play a key role for the control
of numerous immune reactions and the maintenance of
immunological tolerance. CD4 + Tregs are characterized
by the expression of the transcription factor Foxp3 which
is central for both the development of Tregs and their suppressive
characteristics. The majority of Foxp3 + Tregs is
already formed during the development of T cells within
the thymus and shows features of a stable T cell line. We
could recently show that epigenetic modifications within
an evolutionarily conserved region of the foxp3 gene are
decisive for the stability of Foxp3 expression (Huehn et al.,
2009). These epigenetic modifications are already initiated
during the thymic maturation of Tregs. The epigenetic
modifications of this evolutionarily conserved region are
essential for its activation, allowing the binding of specific
transcription factors (Polansky et al., 2010). Thus, these
epigenetic modifications can be regarded as a kind of molecular
switch.
Foxp3 + Tregs are not exclusively generated in the thymus
but also in the periphery, where antigen recognition under
tolerogenic conditions leads to de novo induction of Foxp3 +
Tregs from conventional, naïve T cells. In particular, we
observed a very efficient de novo induction in mucosaassociated
lymphoid tissues with the lymph node stroma
cells playing a key role. Currently, we are studying the
mechanisms leading to the ‘tolerization’ of stroma cells
within mucosa-associated lymphoid tissues. Moreover, we
are investigating whether Foxp3+ Tregs which were de novo
induced within mucosa-associated lymphoid tissues have a
superior suppressive capacity to prevent the development of
chronic-inflammatory diseases of the gut.
Foxp3 + Tregs also can very efficiently suppress wanted
immune responses; in a large number of tumour patients
and in various in vivo tumour models a preferential accumulation
of Foxp3 + Tregs within the tumour tissue has
been observed. Currently we are studying to which extend
this accumulation can be explained by specific recruitment,
local expansion or de novo induction of Foxp3 + Tregs within
the tumour tissue. In any case, the large number of Foxp3 +
Tregs within tumour tissues very efficiently prevents the
generation of tumour-specific immune responses. With
the aid of a transgenic mouse that allows the selective
depletion of Foxp3 + Tregs we were able to demonstrate that
the transient depletion of Foxp3 + Tregs under therapeutic
Foxp3 + cells within the spleen: The cells were stained with
fluorescently labelled antibodies. B cells, T cells und Foxp3 +
cells are stained in blue, red and green, respectively.
Picture: Dr. Eberl, Pasteur Institute, Paris, France.
conditions leads to a reactivation of the tumour-specific
immune response. The most efficient regression of tumour
growth was observed when we vaccinated against
tumour-associated antigens simultaneously to the transient
depletion of Foxp3 + Tregs. In future projects we aim to find
ways to selectively eliminate Foxp3 + Tregs or to transiently
inhibit their suppressive capacity. These studies are also of
major interest for the development of vaccination strategies
against chronic infectious diseases.
All in all, we expect that our investigations will result in
both, a better understanding in the molecular mechanisms
of Treg development and function in vivo. This will provide
us new strategies for the induction and modulation of Tregs
in the scope of a therapeutic approach.
Klages, K., C.T. Mayer, K. Lahl, C. Loddenkemper, M.W. Teng, S.F. Ngiow, M.J. Smyth, A.
Hamann, J. Huehn*, and T. Sparwasser*. (2010) Selective depletion of Foxp3+ regulatory
T cells improves effective therapeutic vaccination against established melanoma. Cancer
Research 70:7788-7799. *equally contributed
Polansky, J.K., L. Schreiber, C. Thelemann, L. Ludwig, M. Krüger, R. Baumgrass, S. Cording,
S. Floess, A. Hamann, and J. Huehn. (2010) Methylation matters: Binding of Ets-1
to the demethylated Foxp3 gene contributes to the stabilization of Foxp3 expression in
regulatory T cells. Journal of Molecular Medicine 88:1029-1040.
Huehn, J., J.K. Polansky, and A. Hamann. (2009) Epigenetic control of FOXP3 expression:
the key to a stable regulatory T-cell lineage? Nature Reviews in Immunology 9:83-89.

SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity 93
03.2 Mucosal Immunity and Inflammation
PROJECT LEADER | Priv.-Doz. Dr. Dunja Bruder | Research Group Immune Regulation | dbr@helmholtz-hzi.de
PROJECT MEMBERS | Andrea Autengruber | Harro Frauendorf | Dr. Marcus Gereke | Dr. Andreas Jeron |
Dr. Sabine Stegemann | Dr. Milena Tosiek | Julia Volckmar
We focus on the basic mechanisms underlying T cell-mediated
inflammation in mucosal tissues, mechanisms involved
in peripheral tolerance induction and the impact of infections
on the balance between tolerance and autoimmunity.
Further activities concentrate on pathogen-induced immune
modulation following bacterial and viral infections of the respiratory
tract as well as a therapeutic vaccine to reactivate
cellular immunity to chronic HCV infection.
Tolerance, autoimmunity and infection T cell reactivity
to self antigen in the lung is studied in a transgenic mouse
model based on the expression of influenza hemagglutinin
(HA) in alveolar type II epithelial cells (AECII). Self-antigen
recognition in the lung by CD4+ T cells results in chronic
obstructive pulmonary disease, followed by the induction
of Foxp3+ regulatory CD4+ T cells (Tregs) that suppress
inflammatory immune reactions. These cells release immunosuppressive
mediators such as TGF-ß and IL-10 which
directly contribute to Treg induction. Moreover, they also
express a variety of surface molecules complementary to
receptors on Tregs, suggesting a broader function of AECII in
peripheral tolerance induction. In addition, AECII from the
inflamed lung express elevated levels of antiviral genes.
Whereas HA antigen recognition by CD4+ T cells results in
the induction of Tregs, self-reactive CD8+ T cells from the
inflamed lung are neither activated, nor regulatory. However,
some of HA-specific CD8+ T cells represent memory cells.
Foxp3+ Tregs appear to be dispensable for regulation of CD8+
T cell-mediated lung inflammation since ablation of Tregs
does not result in gain of effector function in CD8+ T cells.
This suggests that tolerance mechanisms being active in CD8
and CD4 T cell-mediated lung inflammation may differ.
Studies in mice predisposed to the development of type
1diabetes revealed that infection with Listeria monocytogenes
leads to rapid onset of the disease. Infection impairs Treg
Fig. 1. Alveolus of a murine lung. blue: DAPI; red: tubulin
Photo: HZI&SySy GmbH, Christian Erck
Fig. 2. Identification of alveolar type II epithelial cells (AECII)
in the alveoli of a murine lung section by staining for surfactant
protein-C (SP-C) (in red) as a typical AECII marker.
Foto: MHH, Christian Hennig&Jana Bergmann
function dependent on listeriolysin O. Ongoing studies shall
help to clarify whether break down of self-tolerance is a
result of the infection itself.
Immune modulation in the context of respiratory infections
Pathogens such as Bordetella bronchiseptica modulate
host immune responses thereby facilitating persistence.
Together with C. Guzmán, we study the impact of chronic
B. bronchiseptica infection on molecular plasticity of Tregs.
We found that Tregs from chronically infected mice express
elevated levels of neuropilin-1, an activation-independent
marker for Tregs. Currently, we are analyzing the coherence
between the observed changes in Treg phenotype and suppressive
function.
Using a mouse model for influenza A virus / Streptococcus
pneumoniae superinfection we study the mechanisms
underlying the increased lethality of mice by S. pneumoniae
induced sepsis following primary infection with the
influenza virus. Increased susceptibility toward S. pneumoniae
infection is not the result of toll-like receptor (TLR)-7
induced lymphopenia. However, TLR-7 signaling contributes
to the fine tuning of antiviral immune responses. Using TLR-
7 deficient mice we could demonstrate that signalling by this
receptor that senses viral RNA influences the outcome of
secondary pneumococcal infection.
Chronic HCV infection represents the major cause for the
development of hepatocellular carcinoma. In frame of the
HGF financed alliance “Immunotherapy of Cancers” we are
aiming at develop a therapeutic vaccine against HCV. The approach
used is based on in vivo targeting of HCV antigen to
maturing Dendritic Cells (DC). We observed a very efficient
CD4+ and CD8+ T cell response as well as comparably high
antibody titers.

94 SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity
03.3 Immuneffectors: Molecules, Cells and Mechanisms
PROJECT LEADER | Dr. Siegfried Weiss | Research Group Molecular Immunology | siw@helmholtz-hzi.de
PROJECT MEMBERS | Imke Bargen | Nicole Dietrich | Dr. Sandra Düber | Dr. Nelson Gekara |
Dr. Jadwiga Jablonska | Katja Kochrübe | Dr. Sara Leschner | Dr. Stefan Lienenklaus | Dr. Bishnudeo Roy |
Swati Shukla | Evgeniya Solodova | Christian Stern | Dr. Nuno Viegas | Kathrin Wolf | Natalia Zietara
Our immune system consists of several lines of defense. One
of the first of such lines are type I interferons (IFN) with
their most important members IFN-α and IFN-β. Originally,
IFNs have antiviral activity but are also induced by all kinds
of other infections and play an important role under normal
physiological conditions. In our experiments, we could show
that in macrophages IFNs are also induced by ligands of the
toll like receptor 2 (TLR2). TLR2 ligands are found in the cell
wall of Gram-positive bacteria and mycoplasms. Interestingly,
to be able to trigger IFN production, the TLR2-ligand complex
has to be endocytosed by the macrophages. Inhibition of
endocytosis leaving the receptor-ligand complex at the cell
surface allows only production of pro-inflammatory cytokines
like TNF-α. In accordance, the ability of a cell to endocytose
surface TLRs decides whether the cell can produce IFNs
after TLR-ligand binding or not. Along this line, we could
show that mast cells are not able to endocytose such TLRs
or to phagocytose bacteria that might have engaged TLRs.
Therefore, mast cells are not able to produce IFN during an
infection by bacteria although they are perfectly able to do
so in case of a virus infection. Anatomically, mast cells are
positioned underneath the various epithelia. Most likely, they
represent the first immune cells to encounter pathogens that
invade the body. During bacterial infection production of IFN
is often detrimental. Thus, the lack of IFN production and
the restriction of mast cells to the secretion of pro-inflammatory
cytokines might represent a well-balanced evolutionary
adapted defense mechanism.
We noticed that mice that lacked the gene for IFN-β or of one
of the receptor chains (IFNAR) exhibited an altered angiogenesis
i.e. formation of blood vessels. This was especially
obvious in fast growing transplantable tumours: In IFN-β koor
IFNAR ko-mice the tumours grew much faster. This was
due to an enhanced angiogenesis resulting in more and more
mature blood vessels in tumours of the recombinant mice.
We could show that neutrophilic granulocytes were involved
in this process (Figure 1). Without a functional IFN system
the tumours contained more of such granulocytes and they
produced more pro-angiogenic factors. By isolating granulocytes
from tumours of the ko-mice we could show that
already small amounts of IFN-β down-regulate the production
of the pro-angiogenic factors. Thus, it becomes clear that
IFNs are a natural component of the tumour surveillance
system of our body.
Many bacteria like Salmonella typhimurium migrate to solid
tumours after systemic application and proliferate there.
Thus, they can be used as shuttle to express therapeutic genes
directly in the neoplastic tissue sparing the healthy organs.
Enhanced tumour angiogenesis and growth is mediated by
neutrophilic granulocytes unresponsive to type I IFN. Mice were
subcutaneously injected with B16F10 melanoma cells that were
mixed either with neutrophils obtained from tumor bearing
Ifnar1 -/- mice (IFNAR-/-+B16) or from tumour bearing wild type
mice (C57BL/6+B16). The tumours mixed with IFN-unresponsive
neutrophils grew much faster. Blood vessels in the tumors were
investigated by immune histology using anti-laminin to stain
endothelial cells (red) and anti-actin to stain smooth muscle cells
(green). The presence of high amounts of actin in the IFNAR-/-
+B16 tumours is indicative for the enhanced maturity of the
blood vessels and thus of a better blood supply of the tumour.
Photo: HZI
Essential for the specific expression of such substances is
the definition of bacterial promoters that restrict expression
to the tumour. To this end we have screened a so-called
promoter-trap-library with our Salmonella and could detect
several promoters with the appropriate specificity. Presently,
we characterize such promoters using bioinformatics as well
as experimental approaches. In parallel, we investigated
tumour-specific bacterial gene expression using micro expression
arrays. We were able to define more than 20 genes
that were specifically expressed in the tumour and not in the
spleen. First of all this will help to understand the physiological
status of the bacteria residing in the tumour. In addition,
this will also lead to additional control elements that will
enable us to express therapeutic genes specifically in the
cancerous tissue.

SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity 95
03.4 Interferons in Viral Defence and Immunity
PROJECT LEADER | Dr. Andrea Kröger | Department of Gene Regulation and Differentiation | akg@helmholtz-hzi.de
PROJECT MEMBERS | Katja Finsterbusch | Lucas Kemper | Dr. Mario Köster | Antje Ksienzyk |
Ramya Nandakumar | Berit Neumann | Dr. Ulfert Rand
Interferons (IFNs) are central signalling molecules in the
defense against pathogens and have essential functions in
the interplay of immune cells. The IFN system is the first
line of defense against infections. The effect is based on the
transcriptional activation of multiple antiviral and immune
modulatory genes. The aim of the project is to characterize
the molecular and cellular context of host defense against
infections. The detailed understanding of the involved mechanisms
and the spatio- and temporal dynamics will open
new opportunities for the prevention of infectious diseases.
The IFN network The network of IFN production and action
is strictly regulated, and its deregulation can lead to a
pathologic condition in the host. Spatio-temporally resolved
single cell data on viral infections have revealed IFN induction
mechanisms to be a highly stochastic process while the
response to IFN is bimodal. Based on these data dynamics
and quantitative mathematical models describing the underlying
signaling circuits are developed. This should allow
the prediction of the outcome of an infection. To investigate
the coordinated functions of the IFN network in vivo we
generated reporter mice which allow the investigation of the
IFN signal distribution in the whole body. With these tools
we found that viruses with different pathogenicities lead to
IFN responses which differ in quality, quantity and temporal
activity.
Fail-safe pathway for antiviral activity Many viruses have
developed strategies to circumvent the IFN system and thereby
avoid the defense of the host. We identified an alternative
mechanism of antiviral defense which is IFN-independent.
The transcription factor IRF-1 plays an important role in
this IFN-independent mechanism. Mice which lack IRF-1
are more susceptible against viral infections. We identified
viperin as a gene which plays a major role in this effect.
On encountering viral infections the IRF-1 gene usually
regulates the IFN response, but if the virus has developed
strategies to circumvent this response there is an alternative
interferon independent mechanism of antiviral defense. The
impact of the IFN-independent mechanism in host defense
will be investigated in other infection models like HCV.
Signal progression during viral infection Time-lapse
microscopy of IFN induction (green) and response (red)
resolves spatio-temporal dynamics in IFN signal propagation.
Photo: HZI
IRF-1 induces innate and adaptive immune responses
Apart from the antiviral response the IFN system is also
essential for the induction and stimulation of immune
responses. Enhanced immune cell activation is critical for
anti-tumour responses. The expression of IRF-1 in a lung metastases
model inhibits the development of metastases. This
effect is mediated by the attraction and activation of natural
killer cells. Concurrently, the induction of IRF-1 results in
tumour specific CD8+ T cell response which protects against
secondary tumour development. The inflammatory tumour
microenvironment plays an important role in the outcome of
the immune response. Detailed analysis of the microenvironment
and the signalling cascades of the infiltrating immune
cells will elucidate the mechanism of immune response
induction in future.
Stirnweiss, A., Ksienzyk, A., Klages, K., Rand, U., Grashoff, M., Hauser, H., Kröger, A.
(2010) IFN regulatory factor-1 bypasses IFN-mediated antiviral effects through viperin
gene induction. Journal of Immunology 184(9), 5179-5185.
Pulverer, J.E., Rand, U., Lienenklaus, S., Kugel, D., Zietara, N., Kochs, G., Naumann, R.,
Weiss, S., Staeheli, P., Hauser, H., Köster, M. (2010) Temporal and spatial resolution of
type I and III interferon responses in vivo. Journal of Virology 84(17), 8626-8638.
Dietrich, N., Rohde, M., Geffers, R., Kröger, A., Hauser, H., Weiss, S., Gekara, N.O. (2010)
Mast cells elicit proinflammatory but not type I interferon responses upon activation of
TLRs by bacteria. Proceedings of National Academy of the Science USA 107(19):8748-8753.

96 SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity
03.5 Cell-death Mechanisms in Immunity
PROJECT LEADER | Prof. Dr. Ingo Schmitz | Research Group Systems-oriented Immunology and Inflammation
Research | isc09@helmholtz-hzi.de
PROJECT MEMBERS | Michaela Annemann | Frida Ewald | Ralf Höcker | Dr. Yvonne Rauter | Marc Schuster | Tanja Telieps
Types of cell death Multicellular organisms have evolved
multiple cell-death mechanisms which are activated upon
different stimuli and cell type-dependent to allow a finetuning
of cellular responses to environmental conditions.
Necrosis is characterized by swelling and rupture of the cells
so that intracellular contents leak out of the cell and initiate
an inflammatory immune response. During apoptosis, on
the other hand, cells shrink and the integrity of the plasma
membrane stays intact. Therefore, leakage of intracellular
components does not take place and activation of the immune
system is inhibited. Apoptosis plays an important role
in the interplay between pathogens and the host cell. In
this regard, immune cells may kill infected tissue cells by
the induction of apoptosis. On the other hand, viruses and
bacteria are able to inhibit host cell apoptosis to ensure their
own survival and further spreading. Deregulation of apoptosis
may lead to various diseases such as viral hepatitis (too
much apoptosis) or cancer (too little apoptosis).
Apoptotic signal transduction Two major apoptosis pathways
exist referred to as the extrinsic and intrinsic pathway,
respectively. The intrinsic pathway acts on the mitochondrium
and is regulated by proteins of the Bcl-2 family. The
extrinsic signalling cascade is initiated at the cell surface
by so-called death receptors with CD95 being a prototypical
family member. CD95 plays a crucial role in a number of
immunological processes such as T cell-dependent immune
responses but also in tumour development. Not much is
known about the apoptosis signalling mechanisms which
operate during acute and chronic infections in vivo. The aim
of our project is to analyse these signalling pathways using
different transgenic mice strains and apoptosis-modulating
substances (e.g. caspase inhibitors). One family of proteins
we may focus on are the FLICE-inhibitory proteins (FLIP),
important regulators of CD95 signaling. FLIP proteins have
been identified in mammalian cells (cellular FLIP = c-FLIP)
as well as in γ-herpesviruses (v-FLIP). We could recently
show that, though structurally very similar, viral and cellular
FLIP proteins act by different biochemical mechanisms. The
functional consequences and potential therapeutic windows
opened by these differences will be analysed in the future.
The intrinsic apoptosis pathway is activated by different stimuli
such as UV irradiation or growth factor deprivation. Subsequently,
pro-apoptotic BH3-only proteins are activated to inhibit the
anti-apoptotic Bcl-2 family members. This event allows Bax and
Bak to form pores in the outer mitochondrial membrane so that
apoptogenic factors such as cytochrome c are released. In the
cytosol, cytochrome c binds to Apaf-1 to form a multiprotein complex,
called the apoptosome, at which pro-caspase-9 is activated.
Caspase-9 then cleaves caspase-3 to start a proteolytical cascade
ultimately leading to cell death. The extrinsic apoptosis pathway
is activated by death receptors. Upon binding of trimeric ligands,
the receptor oligomerizes and recruits the adapter molecule
FADD and pro-caspase-8 to form a death inducing signaling complex
(DISC). Caspase-8 is activated at the DISC and can directly
activate caspase-3 or it cleaves the BH3-only protein Bid to tBid
leading to activation of the intrinsic pathway. c-FLIP proteins
can inhibit the activation of caspase-8 at the DISC.
Autophagy Autophagy is a lysosomal degradation pathway
that is important for cellular homeostasis, mammalian development
and immunity. On the one hand, autophagy is able to
eliminate intracellular pathogens. However, some pathogens
use the autophagic pathway to escape immune recognition.
Though the process of autophagy has been elucidated on
the molecular level in recent years, little is known about
signaling mechanisms regulating the effector functions of
these molecules. We have recently uncovered a novel regulatory
mechanism involving the stress kinase p38 and Atg5,
the latter being an essential component of the autophagic
pathway. We will address the role of autophagy regulation for
the intracellular survival of Staphylococcus aureus in mammalian
cells.
Ueffing N, Keil E, Freund C, Kühne R, Schulze-Osthoff K, Schmitz I (2008) “Structural
and mutational analyses of c-FLIPR, the only murine short FLIP isoform, reveal requirements
for DISC recruitment”. Cell Death and Differ 15(4):773-82.
Ueffing N, Schuster M, Keil E, Schulze-Osthoff K, Schmitz I (2008) “Upregulation of
c-FLIPshort by NFAT contributes to apoptosis resistance of short-term activated T cells”.
Blood 112(3):690-8.
Ueffing N, Singh KK, Christians A, Thorns C, Feller AC, Nagl F, Fend F, Heikaus S, Marx
A, Zotz RB, Brade J, Schulz WA, Schulze-Osthoff K, Schmitz I*, Schwerk C (2009) “A
SNP in the cellular FLICE-inhibitory protein (c-FLIP) gene determines protein isoform
production and is associated with risk of follicular lymphoma in humans”. Blood
114(3):572-9.
* corresponding author

SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity 97
03.6 Inflammation and Regeneration
PROJECT LEADER | Priv.-Doz. Dr. Gerhard Gross | Department of Gene Regulation and Differentiation |
ggr@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Thomas Böldicke | Markus Heine | Sandra Laggies | Dr. Virginia Seiffart |
Stefan Somplatzki | Lijing Sun | Dr. Herbert Weich | Dr. Manfred Wirth
Infection of tissues and trauma, shock and sepsis induce a
severe inflammatory response, the primary role of which is
to eliminate pathogens, promote tissue repair and return the
host to optimal normal function. In contrast to acute inflammation,
chronic inflammation leads to a progressive shift in
the type of cells which are present at the site of inflammation
and, in general, leads to the destruction of the tissue in or
near the inflammatory process often resisting regeneration.
We follow the hypothesis that inhibition of the inflammatory
signaling cascades will allow the design of new antiinflammatory
and therapeutic modalities. In this regard, we
investigate the role of caveolin-1 in inflammation in general
and in virus infection in particular. Caveolin-1 represents a
multifaceted membrane protein involved in cellular transport
and signaling. It interacts with the human influenza virus
matrix protein M2 and, moreover, modulates virus production.
Further investigation of M2/Caveolin-1 interaction and
its signal transduction pathways may, therefore, result in a
novel target for therapeutic intervention.
As another anti-inflammatory strategy we are developing
specific intrabodies which are able to retain the Toll-like
receptors 2 and 9 (TLR2 / TLR9) in the endoplasmic reticulum
(ER). We could already demonstrate that an adenovirus
vector-dependent, systemic expression of an anti-TLR2
intrabody has the capacity to provide resistance to a lethal
septic challenge with Bacillus subtilis. We will now focus
on the investigation whether or not TLR2- and the recently
constructed TLR9-intrabodies are able to interfere with a
chronic inflammatory disease such as rheumatoid arthritis.
Rheumatoid arthritis is a severe chronic, systemic, inflammatory,
autoimmune disorder that may affect many tissues
and organs but principally attacks and destroys synovial
joints. About 1% of the world‘s population is affected. This
disorder may lead to a substantial loss of mobility and, in
general, patients suffer from serious pain and are afflicted
by an increased mortality. Current therapies for rheumatoid
arthritis are often unsatisfactory and an ongoing search
for new therapeutic modalities is, therefore, required. We
recently showed that a novel systemic targeting strategy for a
central inflammatory signaling mediator (TGF-beta-activated
kinase-1; TAK1) using the RNAi technology dramatically alleviates
rheumatoid arthritis symptoms in a murine collageninduced
arthritis model. Down-regulation of TAK1 protects
the synovial joints from degradation and causes a significant
reduction of inflammatory Th1 and Th17 cells. TAK1 is,
there fore, a novel and attractive therapeutic target to interfere
with rheumatoid arthritis. At present, we aim to transfer
these results onto other chronic inflammatory disorders.
Angiogenesis and the activation of the blood vascular system
play important roles in sustaining a chronic inflammation. In
contrast, however, the role of the lymphatic vasculature in
chronic inflammation has remained unclear. We have established
novel potent isolation techniques which allowed for
the first time the isolation of human lymph endothelial cells
from normal skin and from diseased tissue (lymphangiomas).
In addition, we were successful in isolating mouse endothelial
progenitor cells from adult lungs which showed a bipotential
capacity to differentiate in vitro and in vivo into blood
endothelial cells and lymph endothelial cells. These primary
cells have been extensively compared and also subjected to
expression profiling to find genes as possible targets for the
therapy of inflammatory disorders.
In vitro differentiation of endothelial progenitor cells (EPCs)
from mouse lung: Bipotential capacity for the generation
of blood- and lymph-endothelial cells. Prox1, marker of
lymph-endothelial cells (green); marker of blood- and lymph
endothelial cell (red). Photo: HZI
Becker, J., Pavlakovic, H., Ludewig, F., Wilting, F., Weich, H.A., Albuquerque, R., Ambati,
J., & Wilting,J. (2010) Neuroblastoma progression correlates with downregulation of the
lymphangiogenesis inhibitor sVEGFR-2. Clinical Cancer Research 16, 1431-1441.
Charbord, P., Livne, E., Gross, G., Haupl, T., Neves, N. M., Marie, P., Bianco, P., & Jorgensen,
C. (2010) Human bone marrow mesenchymal stem cells: A systematic reappraisal
via the genostem experience. Stem Cell Review, in press
Courties, G., Seiffart, V., Presumey, J., Escriou, V., Scherman, D., Zwerina, J., Riuz, G.,
Zietara, N., Jablonska-Koch, J., Weiss, S., Hoffmann, A., Jorgensen, C., Apparailly, F., &
Gross, G. (2010) In vivo RNAi-mediated silencing of TAK1 decreases inflammatory Th1
and Th17 cells through targeting of myeloid cells. Blood, in press.

98 SCIENTIFIC REPORTS | Infection and Immunity | Infl ammation and Immunity
03.7 Cellular Models for Infection
PROJECT LEADER | Dr. Dagmar Wirth | Research Group Model Systems for Infection and Immunity |
dkl@helmholtz-hzi.de
PROJECT MEMBERS | Muhammad Badar | Lacramioara Botezatu | Milada Butueva | Leonor Gama-Norton |
Natascha Kruse | Daniel Maeda | Prof. Dr. Peter P. Müller | Dr. Kristina Nehlsen | Dr. Upneet Sandhu |
Stephanie Sievers
Within this project we develop cellular models as well as
transgenic mouse model systems based on previously developed
strategies for controlled transgene expression. These
systems are employed to address specific questions within
infection research.
Targeted integration of expression cassettes In the past,
we developed highly efficient tools to predictably express
transgenes in cells and mice. For this purpose we used
recombinase-mediated targeted integration of expression
cassettes into previously tagged chromosomal loci (hot-spots).
This enabled us to investigate and optimize expression
cassettes at defined chromosomal loci. We could show that
optimal expression features depend on the interplay of chromosomal
site and the regulatory elements of the incoming
vector. This allowed identifying optimal cassette design for a
panel of different integration sites.
Tightly controlled and tissue-specific transgene
expression in the mouse: a model for induced hepatitis
Based on the strategies for predictable transgene expression,
we have established a platform of mouse-embryonic stem
cells for targeted integration of transgene cassettes. This
strategy was employed to induce liver-specific expression of
viral antigens from HBV and HCV. We could show that induction
of antigens in the liver results in a massive activation of
CD8 T cells resulting in pronounced hepatitis. This mouse
model is currently being characterized to elucidate the interaction
of T cells with liver antigens.
A toxin/antitoxin system To achieve long term expression
of transgenes selection systems are usually used that allow
enriching transgene-expressing cells in presence of selection
drugs. Since this regimen is not feasible in in-vivo systems,
we have developed an alternative strategy to stabilize transgene
expression without selection. For this purpose a cell
system was developed in which toxin- induced cell death is
overcome by expression of an antitoxin. Upon coupling expression
of the anti-toxin to the transgene we could stabili ze
transgene expression over time in absence of selection drugs.
Prevention of microbial biofilms on medical implants
To prevent the formation of bacterial biofilms on implants
slow release coatings were developed with the aim to provide
bactericidal activity for a critical period after implantation.
Two different strategies were employed. A surface-modified
mesoporous silica layer containing drug deposits could
release an antibiotic over an extended period of time. Alternatively,
a coating derived from a biologically synthesized
secondary metabolite provided antibacterial in vitro activity
against P. aeruginosa for approximately one week.
Sandhu, U., Cebula, M., Behme, S., Riemer, P., Wodarczyk, C., Reimann, J., Schirmbeck,
R., Hauser, H. and Wirth, D. (2011) Strict control of transgene expression expression in a
mouse model for sensitive biological applications. Nucleic Acids Research (in press)
Gama-Norton, L., Herrmann, S., Schucht, R., Coroadinha, A., Löw, R., Bartholomae C,
Schmidt M, Alves, P., Baum, C., Schambach, A., Hauser, H., and Wirth, D. (2010) Retroviral
vector performance upon integration into defined chromosomal loci of modular
packaging cell lines. Human Gene Therapy 21(8):979-91.
Mouse models with induced liver-specific activation of antigens
A. Proof of the activated luciferase through bioluminescence
B. The activation of ovalbumin results in a damage of hepatocytes
and the liberation of specific enzymes (ALT)
Nehlsen, K., Herrmann, S., Zauers, J., Hauser, H. and Wirth, D. (2010) Toxin-antitoxin
based transgene expression in mammalian cells. Nucleic Acids Research 38(5), e32.
May, T., Butueva, M., Bantner, S., Markusic, D., Seppen, J., Weich, H., Hauser, H., and
Wirth, D (2010). Synthetic gene regulation circuits for control of cell expansion. Tissue
Engineering Part A. 6(2): 441-452
Nehlsen, K., Schucht, R., Gama-Norton, L., Kromer, W., Baer, A., Cayli, A., Hauser, H. and
Wirth, D. (2009) Recombinant protein expression by targeting pre-selected chromosomal
loci. BMC Biotechnology, 9, 100.
Badar M., K. Hemmen, M. Nimtz, H. Hauser, M. Stieve, M. Stiesch, T. Lenarz, U. Möllmann,
S. Vogt, M. Schnabelrauch, P.P. Mueller. (2011) Evaluation of madurahydroxylactone
as a slow release antibacterial implant coating. TOBEJ, in press

SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
99
04 Strategies for Prevention and Therapy
TOPIC SPEAKER | Prof. Carlos A. Guzmán, MD, PhD | Department of Vaccinology and Applied Microbiology |
cag@helmholtz-hzi.de
One third of all deaths occurring each year worldwide are directly caused by infectious agents. Microorganisms are also responsible
for at least 15% of new cancers, and they are involved in the pathogenesis of many chronic non-infectious diseases. Furthermore,
a recent report suggested that the incidence of lung cancer is signifi cantly increased in case of chronic tuberculosis. In addition,
the major public health problem represented by infections is rendered even more dramatic by the global emergence of multiple
drug-resistant strains. It is, therefore, critical to establish new approaches to fi ght microbial pathogens. Thus, the main objective
of this topic is to develop innovative tools and strategies for the prevention, management and control of infections and infectionassociated
diseases. This is achieved by identifying novel intervention targets and bioactive molecules, designing new immune
interventions, and establishing novel diagnostics and biomarkers.
In the project “Molecular Mechanisms of Hepatitis C Virus Infection and Replication” virus-host interactions crucial for hepatitis C
virus (HCV) replication are investigated. These efforts should help to both defi ne new drug targets conferring inhibition of HCV
replication and implement screening assays for the identifi cation of molecules preventing interactions crucially involved in HCV
replication. To this end, cell based assays are exploited to identify cellular co-factors that HCV usurps for its propagation. Cellbased
high-throughput screening systems are also being developed to identify HCV-specifi c inhibitors using HZI natural compound
libraries. In this context, a novel luciferase-based screening system to monitor HCV replication was developed which was adapted
to 384-well format in collaboration with the Department of Chemical Biology. This allowed the screening of large compound
libraries for small molecules that may interfere with HCV replication. At the same time, the direct infl uence of immunosuppressive
drugs on HCV propagation was assessed. This is of interest, because chronic HCV infection is one of the key indicators for liver
transplantation, and immunosuppression is mandatory after transplantation to prevent graft rejection. Thus, these studies can
help to eventually improve the management of HCV patients post transplantation. A particular focus of the work lays in the
characterization of determinants defi ning the narrow species-tropism of HCV which should support the development of immunocompetent
small animal models for this virus.
In the project “Senescence Surveillance in Chronic Hepatitis and Hepatocellular Carcinome” innovative mouse models are
developed to study the role of the cellular senescence programme for immune surveillance during chronic liver infl ammation and
liver cancer. This is linked with studies addressing the oncogenic potential of HBV and HCV structural proteins in hepatocarcinogenesis.
Beyond this, functional genomics driven approaches are exploited to study the molecular mechanisms of liver damage,
liver regeneration and liver cancer. To this end, microRNA based shRNA technology enabling stable or reversible RNAi-mediated
inhibition of endogenous genes in cultured cells or in experimental mouse models is used. Genome-scale retro- and lentiviral
shRNA libraries targeting the murine and the human genome are harnessed to identify new therapeutic targets in liver cancer and
liver regeneration.
The project “Immune Aging and Chronic Infection” aims to defi ne the effects of persistent infections in immune homeostasis and
immune senescence. In this context, there is a consensus that the adaptive immune response to emerging infections is compromised
in the elderly. Aging results in a loss of both naïve T-cells and T-cell clonal diversity, thereby increasing the susceptibility to
novel infections and decreasing the ability to generate protective immunity upon vaccination. However, the underlying mechanism

100 SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
has not been completely elucidated. Cytomegalovirus (CMV) is a ubiquitous pathogen, infecting the majority of the human
population worldwide. CMV establishes a lifelong latent infection which dominates the repertoire of the memory T-cell compartment.
Thus, the cellular and molecular mechanism of induction of the adaptive immune system during CMV infection is being
elucidated, as well as the broader biological implications in terms of evolution of T-cell repertoires and immune function against
CMV and other chronic infections. The knowledge emerging from this project is expected to facilitate the development of intervention
strategies specifi cally tailored for the elderly.
The project “Interface between Innate and Adaptive Immunity” is dissecting the role played by type I interferon (IFN-I) for limiting
viral spread, in periphery as well as at the level of the central nervous system. This is a critical issue since IFN-I is induced a few
hours after infection, warranting initial host survival, since an adequate virus specifi c adaptive immune response able to eradicate
the pathogen is activated after days. More specifi cally, the activities address how DNA (human CMV and vaccinia virus) or RNA
(HCV and vesicular stomatitis virus) encoded viruses induce IFN-I responses and which type of IFN-inhibiting factors they have
evolved. In this context anti-viral responses of human dendritic cell (DC) subsets such as conventional DC and plasmacytoid DC
are studied in vitro, whereas the role of murine DC subsets is assessed exploiting in in vivo models. Gene stimulation signatures of
virus-stimulated human DC subsets are of particular interest and may eventually yield new biomarkers. Interestingly, all examined
viruses have developed evasion strategies to inhibit either the induction of IFN-I responses or the IFN-I effector function. Dedicated
experiments are tackling the differential IFN-I induction in different anatomic niches, as well as the impact of virus induced IFN-I
on the induction of virus specifi c CD8+ T cell responses, virus neutralizing antibody responses, and NK cell activity. The emerging
knowledge will be exploited to improve pegylated IFN-α/Ribavirin therapy of chronic HCV infection and to establish new approaches
for immunotherapies with antiviral cytokines.
The “Antigen Delivery Systems and Vaccines” project is focused on the development of tools and strategies to optimize the
delivery of vaccine antigens, particularly by the mucosal route, and their subsequent exploitation for the generation of vaccines
against specifi c diseases. This is linked to a programme aimed at the establishment of cost-effi cient preclinical validation models
based on humanized mice for the screening, selection, and prioritization of candidate interventions. In the context of vaccination,
it is critical to optimize antigen design. This is particularly true when pathogens which have evolved immune escape mechanisms
are considered. In the case of Trypanosoma cruzi, despite strong immune responses elicited after natural infection or vaccination,
parasite survival suggests that these responses are inadequate. Studies performed with recombinant proteins spanning different
domains of cruzipain, a major cystein proteinase of T. cruzi, suggested that immunization with full-length cruzipain resulted in poor
recognition of the N-terminal catalytic domain. In contrast, “masking” of protective determinants can be reverted by using the
N-terminal domain as a tailored immunogen. This approach thus allows redirection of the host response, thereby providing
enhanced protection against acute and chronic phase infection. Other activities were focused on optimizing antigen delivery by the
mucosal route. In this context, the stimulation of specifi c T helper (Th) subsets is critical to promote effi cient immunity without
side effects. However, there is an incomplete knowledge on the effector mechanisms triggered by mucosal vaccination. Thus, it
was evaluated which Th subsets are stimulated following intranasal immunization. The results demonstrated that there is a
preferential induction of Th17 cells.

SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
101
The “Therapeutic Cellular Vaccines” project is geared towards the development of strategies to break the immune escape
mechanisms operating in persistent infections. This can be achieved by specifi c stimulation of professional antigen presenting
cells, especially dendritic cells (DC). However, a strong immune activation can also lead to immune pathogenic processes which
may be controlled by immunosuppressive cells, such as mesenchymal stromal cells. Thus, antigen presenting cells were modifi ed
using adenoviral vectors encoding antigens and immunomodulatory molecules to improve their antigen presentation capacities.
It was also investigated if the stimulatory capacity of DC can be enhanced by infection with Mycobacterium bovis BCG. To facilitate
translation of basic research into cell therapies, fl exible and robust good manufacturing praxis (GMP)-compliant processes for
the cultivation and modifi cation of different cell types, including DC, were established using a closed integrated bag system. The
use of dielectric barrier discharge (plasma) treatment in presence of suitable precursors allowed modifying the surface properties
of the bag-system, thereby enabling the cultivation of adherent cells (e.g. mesenchymal stem cells).
The “Molecular Diagnostics of Microbial Pathogens” project is focused on the development and application of high resolution
molecular diagnostic tools for the study of food- and waterborne bacterial pathogens. These tools aim at the identification of single
strains which are responsible for infections of individual patients or cause outbreaks of infectious diseases. They allow assessing
the presence as well as the virulence and degree of metabolic activity of pathogens present in human, animal, and environmental
samples. By implementing methods such as the Multi-Loci Variable Number of Tandem Repeats Analysis (MLVA), the project aims
at understanding the infection routes of bacterial pathogens from food and water as well as to obtain insights into the evolution
and epidemiology of pathogenic bacteria in clinical and natural environments. Validation of a MLVA method developed for Vibrio
parahaemolyticus with a set of Chilean isolates indicated that, despite pandemic occurrence, local evolution is relevant for infectivity.
Adaptation of the approved MLVA method for Legionella pneumophila to fi nished drinking water also allows detection and identifi -
cation of specifi c MLVA-genotypes without cultivation.
Fig. 2. Infection of mouse cells with retroviral HCV pseudoparticles. These pseudoparticles carry either the wild-type envelope proteins
(left side, upper left) or the envelope proteins adapted to the mouse CD81 (left side, upper right). Utilizing these pseudoparticles we inoculated
mouse cells that carry either all four human factors or the corresponding mouse factors (left side, bottom). The pseudo-particles
transfer an eGFP reporter gene so that infected cells can be identified on the basis of their eGFP fluorescence. The quantity of infected
cells is indicated (right side). (see next page)

p7
102 SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
04.1 Molecular Mechanisms of Infection and Replication
in the Hepatitis C Virus
PROJECT LEADER | Prof. Dr. Thomas Pietschmann | Department of Experimental Virology | Twincore |
tpi07@helmholtz-hzi.de
PROJECT MEMBERS | Dorothea Bankwitz | Dr. Julia Bitzegeio | Dr. Christiane Brohm | Dr. Sandra Ciesek |
Martina Friesland | Dr. Sibylle Haid | Kathrin Hüging | Dr. Eike Steinmann
The Hepatitis C Virus (HCV) infects only humans and
chimpanzees. Therefore, besides chimpanzees, which can
hardly be used for experimentation due to high costs and
ethical concerns, the only animal model currently available
are partially “humanised” mice. These animals lack an immune
system, permitting xenotransplantation with primary
human liver cells, which then sustain HCV propagation.
Although certainly very useful for drug development, this
model is, however, unsuitable for the development of vaccines,
since no immunological studies can be carried out
due to the immune defect. Given these circumstances, we
are examining why HCV does not propagate in murine
liver cells. By analysing the mechanisms that restrict HCV
propagation in mouse cells we learn how HCV makes use of
essential co-factors and which murine antiviral factors may
limit HCV replication in mice. Collectively, these studies
should help to define what hurdles have to be surmounted
in order to develop an immune-competent small-animal
model for HCV.
Entry of HCV into mouse cells cannot occur without human
cell-entry factors, since the proteins on the surface of the
virus are for the most part incompatible with the decisive
mouse proteins on the mouse liver cells. To enable HCV
to enter into the liver cells, the virus must interact with
four different molecules on the surface of the target cell.
These cellular proteins are SR-BI, CD81, Claudin-1 (CLDN1)
and Occludin (OCLN). Among these, primarily CD81 and
OCLN are used by the virus in a species-specific fashion as
murine orthologs sustain HCV cell entry only poorly. Our
latest results show that murine CLDN1 supports infection
with limited efficiency, indicating that CLDN1 also contributes
to HCV species specificity.
Adaptation of HCV to murine cell-entry factors We attempted
in this context to adapt the virus to the mouse. For
this purpose, a human liver cell-line was produced, which
does indeed possess human SR-BI, Claudin-1 and Occludin;
however, the human CD81 protein is missing. Subsequently,
we artificially introduced the mouse version of CD81, after
which the cells were again susceptible to infection; however,
the level of efficiency was approximately 100 times lower
when compared to the human CD81 (Fig. 1). Through serial
passage of the virus in these cells, a virus variant was selected
that was able to utilize the mouse version of CD81 as
well as the human protein (Fig. 2, see page before). This alteration
produced a disadvantage for the virus; the adapted
virus has now become considerably more susceptible to
neutralization by antibodies from patients. Presumably, this
human
cells with
mouse CD81
infectivity (TCID 50
/ml)
10 8
10 7
10 6
10 5
10 4
10 3
HCV
before
CD81
serial virus passage
infectivity (TCID 50
/ml)
10 8
10 7
10 6
10 5
10 4
10 3
after
CD81
sequencing
C E1 E2 NS2 NS3 NS4B NS5A NS5B
adapted virus
genome
Fig. 1. Adaptation of HCV to murine CD81. HCV was transferred
to human liver cells that carry murine CD81 instead of human
CD81. The cell-culture supernatant of these cells was serially
passaged to the human cells with mouse CD81. The success of
this adaptation was evaluated (below) by inoculation of cells
that carry either human or murine CD81. After the adaptation,
the viral genome was isolated and sequenced. Three mutations,
in the area of the envelope proteins and responsible for the
adaptation to CD81, were identified; they have been indicated
with a star in the virus genome (below right).
adaptation has exposed particularly sensitive areas on the
surface of the virus, thus rendering it more susceptible to
neutralization by antibodies. It is possible that this “opened”
structure, which facilitates conformational changes of viral
proteins required for cell entry, is decisive for the usage of
non-human entry factors.
Outlook Given these findings, it should be theoretically
possible to adapt the virus to further non-human cellular
replication factors. In this manner we can find out how HCV
makes use of the respective host proteins, and why this is
not possible in, for example, mouse cells. The obstacles that
inhibit development of an immune-competent small-animal
model can thus be identified and in the long run strategies
can be devised to ultimately overcome these.
Bitzegeio J, Bankwitz D, Hueging K, Haid S, Brohm C, Zeisel MB, Herrmann E, Iken M,
Ott M, Baumert TF & Pietschmann T. (2010) Adaptation of hepatitis C virus to mouse
CD81 permits infection of mouse cells in the absence of human entry factors. PLoS
Pathogens 6:e1000978.
Haid S, Windisch MP, Bartenschlager R & Pietschmann T. (2010) Mouse-specific residues
of claudin-1 limit hepatitis C virus genotype 2a infection in a human hepatocyte
cell line. Journal of Virology 4(2):964-75.
NS4A

SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy 103
04.2 Interaction between Innate and Adaptive Immunity
PROJECT LEADER | Prof. Dr. Ulrich Kalinke | Research Group Experimental Infection Research | Twincore |
ulrich.kalinke@twincore.de
PROJECT MEMBERS | Dr. Claudia Detje | Dr. Theresa Frenz | Elena Grabski | Sabrina Heindorf | Julia Heinrich |
Christian Mers | Claudia Soldner
Following viral infection, type I interferon responses that
secure the initial survival of the host are usually induced
within hours. It is approximately a week later that adaptive
immunity is activated to an extent that it is able to eradicate
the infecting pathogen. In earlier studies we showed that
upon viral infection a small number of highly specialised
immune cells, also addressed as plasmacytoid dendritic cells
(pDC), produce large quantities of protective type I interferon.
Practically all viruses examined more closely developed
countermeasures that inhibit the induction or the function
of such type I interferon responses. We are investigating how
different viruses induce type I interferon responses, which
strategies they have developed to undermine these responses
and how interferons secure the survival of the host.
Mechanisms of type I interferon induction and its protective
function The molecular mechanism of type I interferon
induction may vary depending on the inducing agent and the
analysed tissue of cell type. We study in the mouse model
how the entry of the vesicular stomatitis virus (VSV) into the
central nervous system via the olfactory system is inhibited
interferon-dependently (see also Figure 1). Furthermore, we
analyse the influence type I interferon responses have on
immune cell functions. In this context, we have found that a
vaccinia virus-derived attenuated pathogen variant, modified
vaccinia virus (MVA), which triggers strong type I interferon
responses, also induces a type I interferon-dependent expansion
of virus-specific T cells. In this context type I interferon
stimulation of both the antigen-presenting dendritic cells as
well as T cells plays a central role. Furthermore, we investigate
the influence type I interferon has on the induction of
long-lasting antibody responses. This work plays an important
role for the development of new vaccination strategies.
Virus-mediated activation of human pDC Some patients
with chronic hepatitis C virus (HCV) infection can be successfully
treated with an interferon-alpha/ribavirin combination
therapy. However, so far little is known about whether
the body’s own interferon system is sufficiently activated
in HCV infection. We investigate which HCV and/or host
encoded factors influence the strength and composition of
HCV-induced cytokine response of human pDC. This work is
carried out in vitro with primary human pDC isolated from
blood samples from healthy donors. In the future also experiments
with pDC from chronically HCV infected patients will
be carried out.
Perspective An improved understanding of the multiple
functions of type I interferon can help to optimize type I
interferon-based therapies of tumors, autoimmune diseases
and viral infections. Furthermore deeper knowledge of viral
mechanisms of immune stimulation and evasion will provide
new approaches for vaccine development.
After intranasal infection of mice, VSV infects olfactory nerves,
travels over the axons to the olfactory bulb and is arrested in
peripheral areas of the olfactory bulb by a type I interferon
dependent mechanism. The image shows the histological analysis
of the olfactory bulb of a mouse that was infected with
an eGFP-expressing VSV (upper panel). It is clearly visible that
the virus is stopped in the glomerular layer of the olfactory
bulb (lower panel). Photos: Twincore
Frenz, T., Waibler, Z., Hofmann, J., Hamdorf, M., Lantermann, M., Reizis, B., Tovey, M,G.,
Aichele, P., Sutter, G., & Kalinke, U. (2010) Concomitant IFNAR-triggering of T cells and
of DC is required to promote maximal MVA-induced T-Cell expansion. European Journal
of Immunology 40, 2769-2777.
Waibler, Z., Anzaghe, M., Frenz, T., Schwantes, A., Pöhlmann, C., Ludwig, H., Palomo-
Otero, M., Alcamí, A., Sutter, G., & Kalinke, U. (2009) Vaccinia virus-mediated inhibition
of type I interferon responses is a multifactorial process involving the soluble type I
interferon receptor B18 and intracellular components. Journal of Virology 83, 1563-1571.
Detje, C.N., Meyer, T., Schmidt, H., Kreuz, D., Rose, J.K., Bechmann, I., Prinz, M., & Kalinke,
U. (2009) Local type I IFN receptor signaling protects against virus spread within
the central nervous system. Journal of Immunology 182, 2297-2304.

104 SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
04.3 Antigen Delivery Systems and Vaccines
PROJECT LEADER | Prof. Carlos A. Guzmán, M.D., Ph.D. | Department of Vaccinology and Applied Microbiology |
cag@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Pablo D. Becker | Dr. Jennifer Debarry | Dr. Thomas Ebensen | Dr. Miriam Nörder |
Dr. Peggy Riese | Rimma Libanova | Kirsten Scholz | Dr. Kai Schulze | Sebastian Weissmann | Dr. Beata Zygmunt
The main aim of this project is the development of tools and
strategies to optimize the delivery of vaccine antigens, particularly
by the mucosal route, and their subsequent exploitation
for the generation of vaccines against specific diseases.
Improving antigen design Despite the strong immune
responses elicited after natural infection with Trypanosoma
cruzi or vaccination against it, parasite survival in the host
suggests that these responses are insufficient or inherently
inadequate. T. cruzi contains a major cystein proteinase,
cruzipain, which is an attractive candidate vaccine antigen.
We produced full-length recombinant cruzipain (rCz), as
well as truncated forms encompassing its N- and C-terminal
domains. Groups of mice were then immunized with the
recombinant proteins combined with CpG-ODN and Czspecific
antibody responses were measured. The highest
IgG titers were observed in mice vaccinated with either the
C-terminal domain or the full-length protein. In contrast, low
antibody titers were stimulated in mice immunized with the
N-terminal domain, thereby suggesting a parasite immune
escape mechanism by avoiding antibody production against
the catalytic N-terminal domain. Similarly, cellular responses
were poorly restimulated with the N-terminal domain in
mice immunized with full-length rCz, whereas most of the
response was evoked by the C-terminal domain. However, the
cellular responses were much stronger when the N-terminal
domain was used for immunization, suggesting that cellular
responses can be down-regulated by full-length Cz to avoid
recognition of the N-terminal domain. Masking of the essential
domain can be reverted by using the N-terminal domain as
a tailored immunogen to redirect host responses to provide
enhanced protection. In fact, mice immunized with the N-
terminal domain were able to better control infection, resulting
in significant lower parasite loads throughout the acute
phase and limited tissue injury during the chronic phase.
Optimizing vaccination strategies Mucosal vaccination
is an attractive strategy for antigen administration, since
this approach is not associated with pain or stress, has an
extremely easy and cost-efficient administration logistics
and does not require highly trained personnel. However,
although many features of the mucosal immune system
have been dissected, there is an incomplete knowledge on
the effector mechanisms triggered by mucosal vaccination.
In this context, the stimulation of specific T helper (Th)
subsets is critical to promote efficient immunity without
side effects. Thus, we investigated which Th subsets are
preferentially stimulated following intranasal immunization.
In comparison to parenteral immunization we detected
an up-regulation of CCR-6 on CD4+ cells which correlated
with an enhanced production of IL-17. This preferential
induction of Th17 immune response is independent from
the kind of adjuvant used.
(A) Antibody response in mice immunized with full-length rCz
or its domains. . (B) Parasite loads during the acute phase of
T. cruzi infection in vaccinated mice. (C) Micrographs of skeletal
muscle from control mice show severe inflammation (×40),
whereas animals immunized with rCz (D), the C-terminal (E)
and the N-terminal (F) domain show moderate inflammation
with edema (lower and upper arrows, ×10), moderate inflammation
and collagen deposition (upper and lower arrows, ×40)
and mild inflammation (×20), respectively; *P < 0.05, **P <
0.01, ***P < 0.001.
It is known that Th17 cells play a critical role in immune
responses against important respiratory pathogens. Thus,
our observation that a Th17 response pattern is specifically
promoted by intranasal immunization has important implications
in the context of optimizing rational vaccine design.
Cazorla, S. I., Frank, F. M. , Becker, P. D., Arnaiz, M., Mirkin G. A., Corral R. S., Guzmán
C. A*. and Malchiodi E. L.* (2010). Redirection of the immune response to the functional
catalytic domain of the cystein proteinase cruzipain improves protective immunity
against Trypanosoma cruzi infection. Journal of Infectious Diseases. 202:136-144.
*Corresponding authors.
Zygmunt, B. M., Rharbaoui F., Groebe L., and Guzman C. A. (2009). Intranasal immunization
promotes Th17 immune responses. Journal of Immunology. 183:6933-6938.

SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy 105
04.4 Chronic Infection and Cancer
PROJECT LEADER | Prof. Dr. Lars Zender | Junior Research Group Chronic Infection and Cancer |
lze08@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Tae-Won Kang | Dr. Michelle Stange | Dr. Torsten Wüstefeld | Dr. Tetyana Yevsa |
Daniel Dauch | Florian Heinzmann | Lisa Hönicke | Anja Hohmeyer | Nils Jedicke | Marina Pesic | Ramona Rudalska
Liver Cancer (Hepatocellular Carcinoma) represents one
of the most frequent and most deadly cancers worldwide.
The incidence of the disease is increasing, with more than
700,000 new cases per year, 50,000 of which occur in
Europe.
We are taking functional genomic approaches to study
molecular mechanisms of liver damage, liver regeneration
and liver cancer. As our major tool, we are harnessing the
naturally occurring process of RNA interference (RNAi).
Over the past couple of years we have developed and refined
microRNA based shRNA technology, which allows stable
or reversible RNAi-mediated inhibition of any endogenous
gene in cultured cells or in mouse models of liver regeneration
and liver cancer. The use of inducible RNAi allows us
to interfere with gene products within regenerating livers
or growing tumours in mice, either at a single gene level or
through pooled multi-gene screening. Genome-scale retroand
lentiviral shRNA libraries targeting the murine and
the human genome are used to conduct screens in order to
identify new therapeutic targets in regenerative medicine
and hepatobiliary oncology.
1. Translational Regenerative medicine and targeting
of liver stem cells The liver has tremendous potential
to regenerate following tissue damage caused by toxins
or infection. It is unique that differentiated hepatocytes,
which normally reside in the Go phase of the cell cycle, can
re-enter the cell cycle subsequent to liver damage and give
rise to new hepatocytes. However, when chronic liver damage
occurs, there is an exhaustion of the regenerative capacity
of hepatocytes and only partial compensation by a stem
cell compartment. The consequence is chronic liver failure,
which represents a major health problem worldwide.
We have established a unique system which allows conducting
in vivo RNAi screens to identify positive and negative
regulators of hepatocyte proliferation during chronic liver
damage. A recently completed in vivo RNAi screen identified
a dual specificity protein kinase as a promising therapeutic
target for hepatic regenerative medicine. Functional
characterization showed that knockdown of the target by
different shRNAs led to a more than 1000-fold increased
proliferative capacity of hepatocytes in a mouse model of
chronic liver damage and greatly improved survival. We
are currently working to translate the obtained genetic
information into new pharmacological strategies for hepatic
regenerative medicine.
2. Translational Oncology, mosaic cancer mouse modeling,
in vivo RNAi, identification of new, innovative
therapeutic targets in liver cancer Tumour genetics can
guide the application of particular therapeutic strategies
and can predict treatment outcome. By combining in vivo
RNAi technology with powerful mosaic liver cancer mouse
models, we have made substantial progress in pinpointing
new cancer genes and tumour suppressor networks in hepatocellular
carcinoma (HCC). In addition to probing tumour
suppressor genes in vivo one at a time, we also performed
in vivo screens to multiplex this approach. We recently performed
the first in vivo RNAi screen, which identified more
than ten new tumour suppressor genes in HCC.
Current projects in the laboratory are aiming to identify
genetic vulnerabilities in hepatobiliary malignancies. These
approaches are based on the concept of “synthetic lethality”,
which assumes that by virtue of their accumulated genetic
alterations, tumour cells may acquire vulnerabilities that
create opportunities for new therapeutic interventions.
Several high confidence targets have been identified and we
are currently trying to translate these into new treatment
strategies.
Dr. Torsten Wüstefeld (le) und Florian Heinzmann (ri) discussing
results from an experiment. On the left side: Dr. Tetyana
Yevsa, on the right side: Ramona Rudalska. Photo: HZI
Zender, L., Xue, W., Zuber, J., Semighini, C.P., Krasnitz, A., Ma, B., Zender, P., Kubicka, S.,
Luk, J.M., Schirmacher, P., McCombie, W.R., Wigler, M., Hicks, J., Hannon, G.J., Powers,
S. & Lowe, S.W. (2008) An oncogenomics-based in vivo RNAi screen identifies tumor
suppressors in liver cancer. Cell 135(5), 852-64.
Xue, W., Zender, L., Miething, C., Dickins, R.A., Hernando, E., Krizhanovsky, V.,
Cordon-Cardo, C. & Lowe, S.W. (2007) Senescence and tumour clearance is triggered by
p53 restoration in murine liver carcinomas. Nature 445(7128), 656-60.
Zender, L., Spector, M.S., Xue, W., Flemming, P., Cordon-Cardo, C., Silke, J., Fan, S.T.,
Luk, J.M., Wigler, M., Hannon, G.J., Mu, D., Lucito, R., Powers, S. & Lowe, S.W. (2007)
Identification and validation of oncogenes in liver cancer using an integrative oncogenomic
approach. Cell 125(7), 1253-67.

106 SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy
04.5 Therapeutic Cellular Vaccines
PROJECT LEADER | Dr. Werner Lindenmaier | Department of Gene Regulation and Differentiation |
wli@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Kurt E. J. Dittmar | Dr. Wilhelm Meyring | Dr. Oliver Schön | Dr. Nadia Zghoul |
Claudia Preuß | Ellen Kuppe
Cell-based immunotherapies have great potential for the
treatment of tumours and persistent infections. Activation
of the cellular immune response can eliminate the cancer
cells and the cells which are persistently infected. However,
tumour cells and persistent pathogens have evolved mechanisms
to escape the normal immune response. Therefore,
specific stimulation by professional antigen presenting cells,
especially dendritic cells (DC), is necessary to overcome
these obstacles. On the other hand, a strong induction of
the immune response, especially when directed against self
tumour antigens, bears the danger of inducing pathogenic
autoimmunity. In this case control of the overwhelming
response by immunesuppressive cells like mesenchymal
stromal cells is an attractive option.
Sterile docking for cellular therapies However, the biggest
obstacle for broader application of cellular therapies is the
requirement of autologous primary cells which have to
be prepared under cGMP conditions in accordance to EU
legislation.
As a tool to overcome this hurdle we have developed completely
closed bag systems for cGMP compliant dendritic cell
generations. Using sterile docking, all steps starting from cell
isolation up to the final formulation and cryo-conservation
of adenovirally modified DC can be realized without opening
the system. In addition to the development of the cultivation
system we investigated whether the stimulatory capacity of
the DC vaccine is enhanced by infection with M. bovis BCG
using flow cytometry, expression profiling and confocal and
electron microscopy and how the balance between anti-tumour
immunity and autoimmunity after DC vaccination can
be controlled in a transgenic murine model system.
So far, the bag system only allowed the cultivation of suspension
cells. Adherently growing cells like mesenchymal stem
cells require adherence-compatible modified surfaces. This
modification can be achieved using dielectric barrier discharge
(plasma). Plasma treatment in the presence of suitable
precursors such as silanes or amino-functionalized compounds
allows selective modification of surface properties.
We have identified suitable surface coatings, which support
adherence and survival of MSC whereas cells in unmodified
bags aggregated and eventually died.
Modified bag MSC in comparison to conventional flask
MSC MSC grown on modified bag surfaces were extensively
analysed in comparison to conventional flask MSC. Analysis
of cell specific surface markers, gene transfer efficiency,
adipogenic and osteogenic differentiation and global expression
profiles did not reveal significant differences between
bag- and flask-MSC. Bag modification using dielectric barrier
discharge therefore opens up new possibilities for the cultivation
of therapeutic cells in a closed system.
Furthermore, secondary surface modification (e.g. by
coupling of cell-specific antibodies) has been used to isolate
CD14 + monocytes from peripheral blood. Leukocytes and
structured modification of the surface allowed localized
adherence and genetic modification providing a tool for the
analysis of interactions between different cell types.
Garritsen,H.S., Macke,L., Meyring,W., Hannig,H., Pägelow,U., Wörmann,B., Geffers,R.,
Dittmar,K.E., and Lindenmaier,W. (2010). Efficient generation of clinical-grade genetically
modified dendritic cells for presentation of multiple tumor-associated proteins.
Transfusion. 50, 831-842.
Adherent growth on modified surface and karyotype stability
of MSC A) Electron micrograph of MSC growing adherently on
modified bag surface (Coop.: M. Rohde, HZI).
B) Even after passage 22 a normal diploid karyotype is
maintained. (Coop.: K. Miller, MHH).
Macke,L., Garritsen,H.S., Meyring,W., Hannig,H., Pagelow,U., Wörmann,B.,
Piechaczek,C., Geffers,R., Rohde,M., Lindenmaier,W., and Dittmar,K.E. (2010). Evaluating
maturation and genetic modification of human dendritic cells in a new polyolefin
cell culture bag system. Transfusion. 50, 843-855.
Dittmar,K.E.J., Simann,M., Zghoul,N., Schön,O., Meyring,W., Hannig,H., Macke,L.,
Dirks,W., Miller,K., Garritsen,H.S.P., and Lindenmaier,W. (2010). Quality of Cell
Products: Authenticity, Identity, Genomic Stability and Status of Differentiation. Transfusion
Medicine and Hemotherapy 37, 57-64.

SCIENTIFIC REPORTS | Infection and Immunity | Strategies for Prevention and Therapy 107
04.6 Molecular Diagnostics of Microbial Pathogens
PROJECT LEADER | Priv.-Doz. Dr. Manfred Höfle | Department of Vaccinology and Applied Microbiology |
mho@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Ingrid Brettar | Dr. Erika Harth-Chu | Karsten Henne | Leila Kalisch | Rolf Kramer
Molecular diagnostics of microbial pathogens focused on the
development and application of high resolution molecular
diagnostic tools for the study of foodborne and waterborne
bacterial pathogens. These tools aim at the identification
of single strains which are responsible for infections of
individual patients or cause outbreaks of infectious diseases.
They are based on the genome-wide analysis of a set of Variable
Number of Tandem Repeats (VNTR) which are used as
highly specific and sensitive taxonomic markers. A set of
these VNTR markers from across the bacterial genome is
selected and can be used to identify the targeted bacterial
species at the clonal level. This method, called Multi-Loci
Variable Number of Tandem Repeats Analysis (MLVA), is
a new tool to understand the infection routes of bacterial
pathogens from food and water. Additionally, this genomic
fingerprinting method provides insights into the evolution
and epidemiology of pathogenic bacteria in clinical and
natural environments.
Foodborne bacterial pathogens We have developed a
MLVA method for Vibrio parahaemolyticus based on comparative
genome analysis of the two sequenced genomes. V.
parahaemolyticus is a marine bacterium causing cholera-like
diarrhea if contaminated mussels are eaten raw or semicooked.
It causes outbreaks around the Pacific and in the
USA; recently, even occurrences in European countries have
been reported, probably due to global warming. We validated
the method with a set of Chilean isolates in collaboration
with the University of Santiago de Chile. Clinical and
environmental strains from the South and North of Chile had
a common evolutionary history and only Northern Chilean
strains showed an evolutionary relationship to Asian strains.
These findings indicate that, despite the pandemic occurrence
of V. parahaemolyticus,the local evolution of a pathogen
is of relevance for its infectivity.
Waterborne bacterial pathogens Legionella pneumophila
is a freshwater bacterium abundant in man-made systems
such as cooling towers, air conditions and drinking water
supply systems (DWSS). It can cause atypical pneumonia
in humans with high mortality stemming from the inhalation
of contaminated aerosols. Detection and identification
of L. pneumophila is hampered by its fastidious growth on
agar plates and its tendency to go into a so-called viable but
non-culturable (VBNC) state. We adapted the approved MLVA
method specific for L. pneumophila to finished drinking
water. Critical was the development and application of high
Epifluorescence microscopy of drinking water bacteria (blue
cells) and immunofluorescence of Legionella pneumophila
serogroup 1 cells (green) in biofilm from a cooling tower.
Photo: Leila Kalisch, HZI
resolution electrophoresis to obtain sequences of single
VNTRs. This approach allowed the detection and identification
of specific MLVA-genotypes in drinking water without
cultivation of the respective strains. We demonstrated the
applicability of this approach to common drinking water
with low levels of L. pneumophila (< 1 cell/liter). Using this
culture-independent method we were able to identify several
new clones in addition to isolated strains from the same
drinking water. This novel tool for in-situ diagnostics of environmental
bacterial pathogens provides a new avenue for the
surveillance of waterborne pathogens after outbreaks have
occurred and advances molecular epidemiology of bacterial
pathogens with high relevance to public health.
Harth-Chu, E. Especio R. T., Christen, R., Guzman, C.A. & M.G. Höfle (2009) Multiplelocus
variable-number of tandem-repeats analysis for clonal identification of Vibrio
parahaemolyticus isolates using capillary electrophoresis. Applied and Environmental
Microbiology 75, 4079-4088
Kahlisch, L. K. Henne, L. Groebe, J. Draheim, M.G. Höfle & I. Brettar (2010) Molecular
analysis of the bacterial drinking water community with respect to live/dead status.
Water Science & Technology-WST 61.1, 9-14
Kahlisch, L. K. Henne, J. Draheim, I. Brettar & M.G. Höfle (2010) High-resolution in situ
genotyping of Legionella pneumophila populations in drinking water by Multiple-Locus
Variable-Number of Tandem Repeat Analysis (MLVA) using environmental DNA. Applied
and Environmental Microbiology 76, 6186-6195

108 SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research
05 Pharmaceutical Research
TOPIC SPEAKER | Prof. Dr. Rolf Müller | Research Group Microbial Drugs | rom@helmholtz-hzi.de |
Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS) | Department of Microbial Natural Products
Infectious diseases are one of the most important causes of death worldwide and contribute considerably to the need for clinical
treatment. Strong interdisciplinary collaboration in anti-infective research is necessary to translate hits from initial stages of
the drug development process to drugs in the clinic (“bench to bedside”) and to cope with the increasing antibiotic resistance of
pathogens. Isolation, identifi cation and optimisation of small molecules interfering with processes involved in infectious diseases
and modelling of the molecular interaction is crucial for understanding the infection process. The knowledge of the transport
mechanism and the drug-target interaction mechanism is required to optimise the impact of the drug while reducing unwanted
side effects. Most of the commercial drugs currently in use are derived from natural products or small molecules mimicking their
pharmacophore. The topic “Pharmaceutical Research” investigates these compounds with chemical biology approaches in order to
provide new classes of sustainable anti-infectives to the drug development pipeline.
The research topic was newly created after the establishment of the “Helmholtz-Institute for Pharmaceutical Research Saarland”
(HIPS) in late summer 2009. It was founded to harvest the potential of a fostered interaction between the HZI and the Department
of Pharmacy at Saarland University (UdS). The incorporation of HIPS adds pharmaceutical sciences to the Health research programme
of the Helmholtz Association. As translational research was identifi ed as one of the most important tasks for future health
research, the foundation of HIPS is a signifi cant contribution to close the translational gap in natural product based anti-infective
research.
The topic is composed of seven groups working at both sites, in Braunschweig (HZI) and Saarbrücken (HIPS). The three departments
at HIPS, namely “Microbial Natural Products” (MINS), “Drug Development and Optimization” (DDOP) and “Drug Delivery”
(DDEL) now commit their respective pharmaceutical expertise to the fi eld of infectious diseases. While MINS has traditionally
strong ties with the group “Microbial Diversity and Natural Product Discovery” (MWIS) at HZI, DDOP and DDEL have built up collaborations
with HZI groups, which already lead to the attraction of third party funding and exchange of scientists. The HIPS departments
are presented separately within this report.
The group “Chemical Biology of Infectious Diseases” (CBIO) as well as the groups “Medicinal Chemistry” (MCH) and “Identifi cation
of Molecular Targets of Anti-infectives”(BISA) were integrated into the topic “Pharmaceutical Research” because they engage in
highly overlapping activities: CBIO uses various chemical biology approaches to characterise the activity and the mode of action of
natural products. MCH uses medicinal chemistry and synthetic approaches to optimise these natural products as lead structures
and make them available in signifi cant quantities. BISA synergistically works on the identifi cation of antifungal compounds and on
deciphering their mode of action.
The group “Chemical Biology of Infectious Diseases” (CBIO) at HZI focuses on the elucidation of molecular mechanisms of infectious
processes by low-molecular drugs. They develop new, miniaturize and run existing activity assays to identify lead structures
and elucidate the mode of action of hits from compound libraries. These libraries consist of approx. 90,000 compounds with
unique collections of natural products from myxobacteria, compounds provided by cooperation partners, commercially available
libraries as well as substances synthesized in-house. CBIO has developed a semi-automated microscopic assay that provides hints
on the mode of action from the changes in the phenotype, coupled with statistical analysis of huge data sets. By software-based
photo analysis of the cells, a biological profi le of the compounds is generated. In a hierarchical cluster analysis, new substances
group with known compounds and similar profi les. The procedure could be validated and already gave valuable hints on new
modes of action and potential new drugs, which will be further investigated.

SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research
109
In another project, a high-throughput microscopic adhesion assay was established to investigate the interaction of the pathogenic
bacterium Staphylococcus aureus with human epithelia cells. After screening of 4,000 substances and mixtures, new promising
compounds could be identifi ed which will now be tested in further studies: with human nasale primary cells on their effi ciency and
in cell-based assays on their toxicity.
The group “Identifi cation of Molecular Targets of Anti-Infectives” (BISA) at HZI deals with commensal and persistent microorganisms.
These microorganisms like Candida albicans and Mycobacterium bovis (BCG) are adopted to survive in presence of human
cells and are only causing infections if the immune defence of the human is weakened. To identify new sites of action for anti-infectives,
BISA simulates the conditions found in humans to screen potential sources for new drugs like libraries of chemical compounds,
extracts from myxobacteria and commercially available inhibitors. One project investigates, if the elimination of C. albicans
by phagocytes can be infl uenced by the treatment with kinase inhibitors. It could indeed be shown that all steps relevant for
the course of the infection can be controlled by chemical substances that in part are formed even by the organism itself, like Genistein
from soya. Another project shows that the histidine kinase CaNik1 of C. albicans is the site of action for fungicides. Unexpectedly,
CaNik1 causes a change in the metabolism but not in growth inhibition. The deletion of several HAMP domains in CaNik1
affects the sensitivity of the fungicides depending on the specifi c compound.
The group “Medicinal Chemistry” (MCH) at HZI deals with syntheses of anti-infectives as well as derivatives and analogues thereof
to optimise the biological profi le and to get insights into the structure-activity relationships. Recently, the absolute confi guration of
the natural product Chondramid C could be confi rmed. On the route to Corallopyronin the closely related natural product Myxopyronin
could be synthesised by a stereoselective and convergent strategy. The successful synthesis of Chivosazol confi rms the proposal
for the assignment of 10 stereo centres. The key step is an unusual Stille coupling for the formation of the macrocycle.
The research topic “Pharmaceutical Research” further develops the close interaction of the group “Microbial Diversity and Natural
Product Discovery” (MWIS) at HZI and the department “Microbial Natural Products” (MINS) at HIPS to further utilise the potential
of gliding bacteria for the production of natural products. Methods of classical microbiology, functional genomics, secondary metabolomics,
advanced screening, and structure elucidation are being employed and further developed. In addition, novel assays
for interference with bacterial cell-cell communication are developed. MWIS collaborates with CBIO, MCH and BISA to extend the
chemical and biological profi le of strains from their rich collection of myxobacteria.
By staining the nuclei of the epithelial cell line A549 with DAPI (blue) and the bacteria with fluorescent antibodies (green), the adhesion
can be visualized. After taking pictures with the automatic microscope (A + B), a software module allows the quantification of the nuclei
and the adherent bacteria. If a substance reduces the bacterial adhesion, the effect can be quantified. Picture B shows the result after
incubation of the cell line with the bacteria in the presence of compound HZI-MW195 at 5 μM concentration in comparison to the control
A without added substance. The concentration dependence of the effect is shown in figure C (see page 112).

110 SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research
05.1 Microbial Diversity and Natural Product Discovery
PROJECT LEADER | Prof. Dr. Rolf Müller | Research Group Microbial Drugs | rom@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Klaus Gerth | Dr. Herbert Irschik | Dr. Rolf Jansen | Wolfgang Kessler |
Dr. Kathrin Mohr | Heinrich Steinmetz
Since the closure of the department of natural product
research in 2006, the new group “Microbial Drugs” is focusing
on the discovery and characterization of novel bioactive
compounds from gliding bacteria. Since 2009, it concentrates
on the reactivation, conservation, and screening of our strain
collection.
Elansolid A1
Elansolid A1 and A2
Elansolid A2
Strain collection: For the first time we got a general idea on
the status of the complete strain collection of gliding bacteria
at HZI. We developed strategies how to improve the value of
the collection and at the same time reduce the number of
strains. So far we saved about 700 strains and prepared fresh
culture extracts which were analysed by modern techniques
of HPLC-UV-HRMS. We isolated DNA for genome sequencing
and the search for silent PKS gene clusters and then characterized
the isolates physiologically.
Natural product biology: About 50 strains of myxobacteria
have been isolated from fresh soil samples and screened.
Based on morphology and 16S rDNA analysis, one of these
strains could not be classified to any known group of myxobacteria.
First inhibition tests against different bacteria, yeasts
and fungi revealed a strong unknown antimicrobial and
fungicidal activity. New groups of myxobacteria hold a good
potential for the discovery of new secondary metabolites.
Fermentation: Two major tasks are covered by the fermentation
facility, provision of raw material for structure
elucidation and all other requests from the chemical pipeline
and external partners on one hand and the improvement of
fermentation processes on the other hand.
Basic studies on the improvement of the fermentation process
were done with Chitinophaga Fx GBF13, the producer
of elansolids. The influence of different carbon- and nitrogen
sources, of different pH values of the culture broth and the
effect of oxygen limitations on elansolid A1 production were
studied systematically.
Natural product chemistry: Gephyronic acid was characterized
as a selective inhibitor of eukaryotic protein synthesis
with a nanomolar cytostatic effect against a range
of mammalian cell lines. Using new MS data and chemical
derivatisation the structure was revised and the absolute
configuration of all asymmetric centres was determined.
Parallel fermentations with Fx GBF 13 at different pO 2
. Photo: HZI
1. Jansen, R., Irschik, H., Huch, V., Schummer, D., Steinmetz, H., Bock, M., Schmidt, T.,
Kirschning, A., & Müller, R. (2010) Carolacton -a macrolide ketocarbonic acid reducing
biofilm by the caries- and endocarditis-associated bacterium Streptococcus mutans.
European Journal of Organic Chemistry 7, 1284.
Development of an anhydrous isolation procedure for a novel
elansolid A isomer surprisingly revealed a common precursor
of the atropisomers elansolids A1 and A2. Genetic studies
now are expected to show which elansolid A isomer really
represents the final product of the biosynthesis.
2. Buntin, K., Irschik, H., Weissman, K. J., Luxenburger, E., Blöcker, H., & Müller, R.
(2010) Biosynthesis of Thuggacins in Myxobacteria: Comparative cluster analysis
reveals basis for natural product structural diversity. Chemistry & Biology 17, 342.
3. Irschik, H., Kopp, M., Weissman, K. J., Buntin, K., Piel, J., & Müller, R. (2010) Analysis
of the sorangicin gene cluster reinforces the utitlity of a combined hylogenetic/
retrobiosynthetic analysis for deciphering natural product assembly by transATPKS.
ChemBioChem 11, 1840.
4. Nawrath, T., Gerth, K., Müller, R., & Schulz, S. (2010) The biosynthesis of the aroma
volatile 2-methyltetrahydrothiophen-3-one in the bacterium Chitinophaga Fx 7914.
ChemBioChem 11, 1014.
5. Menche, D., & Irschik, H. (2010). Design, synthesis and biological evaluation of simplified
analogues of the RNA polymerase inhibitor etnangien. Bioorganic & Medicinal
Chemistry Letters 20, 939.
6. Bü low, L ., Nickeleit, I., Gi rbig, A .- K ., Brod ma n n, T., Rentsch, A ., Egger t, U., Sasse,
F., Steinmetz, H., Frank, R., Carlomagno, T., Malek, N.P., & Kalesse, M. (2010).Synthesis
and biological characterization of argyrin F. ChemMedChem 5, 832.

SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research 111
05.2 Medicinal Chemistry of Natural Products
PROJECT LEADER | Prof. Dr. Markus Kalesse | Department of Medicinal Chemistry |
mka05@helmholtz-hzi.de | markus.kalesse@oci.uni-hannover.de
PROJECT MEMBERS | Christina Brünjes | Dr. Jutta Niggemann | Dr. Evgeny Prusov | Tang Wufeng
In the frame of this project, synthetic routes will be
developed to provide new antibiotics. This will enable the
construction of derivatives and analogues for improving the
biological profile of the lead structure as well as allowing
conclusions about structure activity relationships to be
drawn.
In this context, through synthesis the configuration of the
chonramides could be unambiguously assigned. In addition,
through the synthesis of the four diastereomers in the
polyketide segment of this natural product the contribution
of these segments for its biological activity could be made.
It was shown that the polyketide segment of this natural
product is the structural element, adjusting the peptide portion
of the molecule so that the optimal distance and torsion
angle between the two ends of the peptide can be attained.
In the case of corallopyronin, a similar approach will be
used. The first task is to develop a stereoselective and
convergent synthesis to access the natural product. Initially,
the related natural product myxopyronin was synthesised.
Through this synthesis we were able to successfully address
the two main challenging aspects in the myxopyronin
structure and we are now in the process of transferring
these results to the synthesis of corallopyronin and its
derivatives. In comparison to myxopyronin, corallopyronin
has an alternative side-chain which has also been synthesised,
and now the two segments need to be coupled.
New projects which have begun are the syntheses of
pellasoren and angiolam. For both syntheses, the central
transformation should be an asymmetric protonation of an
aldehyde-enolate.
Influence of chondramide C (30) on the actin cytoskeleton
of A-498 kidney cancer after different incubation times. A,B:
control cells with a dividing cell in the centre, in metaphase (A),
and in telophase (B). Cells that were incubated with chondramide
(100 ng/ml) showed abnormal metaphase cells (C, after 2
hours), and a strengthened contractile ring in late telophase (E,
after 18 hours). Spot of F-actin became visible especially at focal
adhesion points (D, after 4 hours), stress fibers became stronger
and flakes of actin appeared (E and F, after 18 hours). (Kalesse
et al (2008) Angewandte Chemie International Edition 47,
6478-6482)
In the synthesis of chloronitril derivatives it was necessary
to make water soluble analogues. We envisioned that the
decalin part of the molecule could be replaced by a simple
double bond. Unexpected difficulties were experienced in
this synthesis and, therefore, to date, no derivatives have
been obtained. This project could be taken further with an
alternative synthesis concept.
The synthesis of chivasozole was successfully completed
this year and confirmed our structure proposal and the
chirality at the 10 stereocentres. The convergent synthesis
is characterised by an unusual Stille reaction which was
used as the macrolactonisation step.
Brodmann, T., Janssen, D., & Kalesse, M. (2010) Total synthesis of chivosazole F. Journal
of American Chemical Society 132, 13610-13611.
Schäckel, R., Hinkelmann, B., Sasse, F., & Kalesse, M. (2010) The synthesis of novel
disorazoles. Angewandte Chemie 122, 1663-1666; Angewandte Chemie International
Edition 49, 1619-1622.
Bülow, F.L., Nickeleit, I., Girbig, A.-K., Brodmann, T., Rentsch, A., Eggert, U., Sasse, f.,
Steinmetz, H., Frank, R., Carlomagno, T., Malek, N.P., & Kalesse, M. (2010) Synthesis
and biological characterization of argyrin. ChemMedChem 5, 832-836.

112 SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research
05.3 Chemical Biology of Infectious Disease
PROJECT LEADERS | Dr. Ronald Frank | Department of Chemical Biology | rfr@helmholtz-hzi.de
Dr. Florenz Sasse | Department of Chemical Biology | fsa@helmholtz-hzi.de
PROJECT MEMBERS | Jihad Al-Qudsi | Ulrike Beutling | Randi Diestel | Dr. Michelle Fountain | Dr. Raimo Franke |
Dr. Bernd Hofer | Denis Koska | Michael Mrosek | Dr. Irina Nickeleit | Dr. Mahtab Nourbakhsh | Dr. Marc Reboll |
Saad Shaaban | Galina Sergeev | Dennis Schwab | Dr. Dr. Werner Tegge | Dr. Peter Washausen | Marina Wöhl
The aim of the project is to elucidate the molecular mechanisms
of infection processes by interfering small molecules.
Special assay systems and screening techniques are developed
to select bioactive substances out of chemical libraries
whose mode of action will be further analysed. These
analyses should lead to new antibiotics, chemotherapeutics,
and immune modulators. The knowledge about their mode
of actions will open new ways of therapy. Within the framework
of the „Chemical Pipeline“, we built an infrastructure
that allows a rapid and effective search in miniaturised
assay systems. Our library of about 90,000 samples includes
a unique collection of natural compounds isolated from
myxobacteria (see 05.1), substances from collaboration
partners, purchased compound collections, and substances
that were synthesised by us. The infrastructure is also at
the disposal of external scientists and to other applications
via the German ChemBioNet (www.chembionet.de).
Phenotypes characterise the mode of action There is
no easy way to elucidate the mode of action of a biologically
active compound. Each biologically active compound
changes the metabolic and signalling pathways within the
Biologically active substances induce phenotypical changes in
treated cells which are characteristic for their mode of action.
This is shown for chivosazol A. The green actin filament structures
of the treated cells in B clearly differ from the control cells
in A. In C we see a hierarchical cluster analysis of substances
from myxobacteria. D gives a detail which shows that chivosazol
A clusters together with a second sample that was given “blind”,
a clear proof of principle.
complex biochemical network of a target cell. This can lead
to striking changes in the morphology of the cell and the
cell organelles or in the protein pattern. These phenotypical
changes can be visualised via antibodies or specific dyes.
Comparing phenotypes is one way to get hints about the
mode of action. Now we have strengthened this method
by automated microscopy and statistical analysis of large
amounts of data. By automated microscopy we can take a lot
of pictures of treated cells, evaluate them by software-based
image analysis, and form a profile of the biologically active
compound. By comparing the profiles of known and unknown
compounds we get hints about the mode of action of
the new substance. This is done by software-based statistical
methods, e.g., hierarchical cluster analysis. Substances,
which have the same mode of action, group together then.
In the meantime we have also got useful hints concerning
the mode of action of new substances. First follow-up
experiments confirmed the first hints.
Inhibitors of the adherence of Staphylococcus aureus to
human epithelial cells In the context of the BMBF funded
consortium “SkinStaph” a high-throughput amenable adhesion
assay was established, which provides the possibility
to search for new inhibitors of the interaction between the
pathogenic bacterium Staphylococcus aureus and human
epithelial cells (see Fig. page 109). Highly reliable data is
obtained by a newly developed method that employs an
automated microscope. Epithelial cells are incubated with
the bacteria in the presence of potentially interfering compounds,
followed by the removal of non-adherent bacteria
and the staining of the cells. The nuclei of the epithelial
cells are stained with a DNA specific dye (DAPI) and the
bacteria are labelled with specific fluorescent antibodies.
Several pictures for every compound are taken with the
auto mated microscope. With a software module the nuclei
and the bacteria are automatically quantified and set into
relation. This allows the identification of substances that
reduce the adhesion of the bacteria. After screening more
than 4,000 individual substances and mixtures, several
promising compounds were identified. These drug candidates
are currently investigated with primary human nasal
cells and their toxicity is determined with cell-based assays.
Diestel, R., Irschik, H., Jansen, R., Khalil, M.W., Reichenbach, H., & Sasse, F. (2009)
Chivosazole A and F, cytostatic macrolides from Myxobacteria, interfere with actin.
ChemBioChem 10, 2900-2903.

SCIENTIFIC REPORTS | Infection and Immunity | Pharmaceutical Research 113
05.4 Identification of Molecular Targets of Antiinfectives
PROJECT LEADER | Prof. Dr. Ursula Bilitewski | Research Group Biological Systems Analysis | ubi@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Nina Klippel | Dr. Bianca Lüderitz | Anna Buschart | Shuna Cui | Mohammed El-Mowafy |
Daniela Evers | Katja Gremmer | Rabeay Hassan | Hani Kaba | Carolin Lewark | Dörthe Sokolis
New therapies of infections require new antimicrobial
compounds, which exploit new molecular targets to avoid
already existing antibiotic resistances. We focused on commensal
and persistent microorganisms, and chose the yeast
Candida albicans and the bacteria Mycobacterium bovis
(BCG) as substitute of the tuberculosis pathogen Mycobacterium
tuberculosis. These microorganisms are adapted to the
survival in the presence of host cells, and become infectious
in patients with an impaired immune system.
Histidine kinases as molecular targets of anti-infectives
Histidine kinases are mainly found in bacteria, but also
in fungi. Mostly they are sensor proteins and induce the
adaptation of the microorganisms to their environment. We
had previously shown that the histidine kinase CaNik1 from
Candida albicans is the molecular target of the myxobacterial
secondary metabolites ambruticin VS3 and jerangolid
A, as we transferred sensitivity for these fungicides to the
otherwise resistant yeast S. cerevisiae by heterologous
functional expression of the CaNik1 gene. We identified a
pairwise sequence of 9 so-called HAMP-domains, each of
which comprises approximately 50 amino acids. Deletion of
selected pairs of these domains from CaNik1 reduced the
sensitivity for fungicides, with the magnitude of the effect
being dependent on the compound. At present investigations
are performed to elucidate further details of the interaction
of the protein with fungicides. Gene expression analysis
showed that in the S. cerevisiae-transformants fungicides,
which target CaNik1, mainly activate the signal transduction
pathway of the osmotic stress defense network, which
includes effects on cell cycle and thus on cell proliferation.
Exploitation of new targets for anti-infectives Candida
albicans usually colonizes human skin and mucosa (mouth,
gastro-intestinal tract, vagina). Systemic infections
include penetration of the underlying cell layers, followed
by invasion of deeper tissues and spread to inner organs
via the bloodstream. An essential pathogenicity factor is
the capability to switch morphology between yeast and
hyphal forms. Among primary immune defense reactions
is phagocytosis and stimulation of the immune system by
activation of macrophages and neutrophils. Macrophages
are also involved in the immune defense against infections
of the lung by mycobacteria. However, these bacteria can
survive phagocytosis as persistent, dormant organisms and
are hardly eliminated by present antibiotics.
To empirically exploit new targets for anti-infectives, we
simulate environmental conditions, which the microorganisms
face in the human host. We test whether small molecules
can enhance the clearing efficiency of phagocytes,
The interaction between the yeast Candida albicans and cells
of the epithelium (cell line A431) in the presence of the peptide
LL37. LL37 belongs to the anti-microbial peptides and does not
inhibit the growth of C. albicans, but it supports the adhesion
of the yeast to human cells. After an incubation time of 6 hours
(conditions of cell cultures), C. albicans forms hyphen which
can be found in the cells of the epithelium as well, where they
also continue to grow. Photo: HZI, Rohde
block the morphology change of C. albicans and the survival
of dormant mycobacteria. Sources of those compounds are
libraries of chemical compounds, extracts from myxobacteria
and commercially available inhibitors. We could show
that all steps of the infectious process can be influenced
by chemical compounds, some of which are endogenous
products of the organisms. Human cells produce peptides,
which manipulate the adhesion of C. albicans to surfaces, in
particular to components of the extracellular matrix. The
formation of hyphae is suppressed by compounds which are
formed by C. albicans as quorum sensing compounds. Finally,
we discovered that genistein, a well-known constituent of
soybeans, stimulates phagocytosis of genistein-treated
C. albicans. Further studies aim at the elucidation of underlying
molecular mechanisms.
Wesolowski,J., Hassan,R.Y.A., Reinhardt,K., Hodde,S. & Bilitewski,U. (2010) Antifungal
compounds redirect metabolic pathways in yeasts: Metabolites as indicators of modes of
action, Journal of Applied Microbiology 108, 462 - 471
Klippel,N., Cui,S., Gröbe,L. & Bilitewski,U. (2010) Deletion of the Candida albicans histidine
kinase gene CHK1 improves recognition by phagocytes through an increased
exposure of cell wall ß-glucans, Microbiology 156, 3432 – 3431
Buschart,A., Gremmer,K., v.d.Heuvel,J. Müller,P.P. & Bilitewski,U. (2011) Repetitive
HAMP domains at the N-terminus of the CaNik1 histidine kinase of Candida albicans
mediate specific interactions with fungicides, Molecular Microbiology, submitted

114 SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups
New Project Groups
The HZI has given young researchers with very good ideas the possibilities to do their research at the HZI facilities when their
scope has been within the forthcoming R&D developments of the Centre. Since 2009 further new projects groups have started
their work at the HZI. During the next 5 years they will try to establish new research ways and present excellent results. The fi r s t
results and further ideas of this 2 nd generation of new R&D groups are presented on the next pages.
The platform in the Gründerzentrum of the HZI, where new project groups have the possibilty of presenting their research results.
The photo was taken during a visit of Nobel Laureate Prof. Dr. Kurt Wütherich (La Jolla, USA / Zürich, Switzerland) (left side at the
first poster) in 2010. Photo: HZI

SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups 115
01 Regulation and Herpesviral Immune Modulation of
Toll-like Receptor Signalling
PROJECT LEADER | Dr. Melanie M. Brinkmann | Junior Research Group Viral Immune Modulation |
mbr10@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Bettina Gruhne | Elisa Reimer
Herpesviruses Herpesviruses establish lifelong persisting
infections. Whereas immunocompetent individuals are
able to control herpesviral infections immunocompromised
persons such as transplant recipients or AIDS patients can
encounter severe medical problems. An infection with the
betaherpesvirus cytomegalovirus (CMV) during pregnancy is
the most prevalent cause for virus-induced abnormalities of
the fetus. Further, the human gammaherpesviruses Epstein-
Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus
(KSHV) can cause tumours. In order to develop improved
prophylactic and strategic therapies it is crucial to improve
our understanding how herpesviruses are recognized by the
host’s immune system and how herpesviruses modulate the
immune response to avoid their detection and elimination.
Innate Immune System The innate immune system allows
an early and potent antiviral response and plays an important
role for the induction of the adaptive immune response.
One group of pathogen sensors of the innate immune system
are members of the family of Toll-like receptors (TLRs). TLR9
senses herpesviral DNA and induces a potent antiviral interferon
response. The cellular UNC93B protein is essential for
proper functioning of TLR9: it delivers TLR9 from the endoplasmic
reticulum, where it is synthesized and stored until
needed, to a specialized intracellular compartment called
the endosome. There TLR9 meets the DNA of the invading
pathogen and initiates a signalling cascade that results in
the activation of an antiviral response. Whereas the downstream
signalling cascade of TLRs is well characterized little
The membrane protein UNC93B interacts with Toll-like receptor
9 (TLR9) in the endoplasmic reticulum and delivers it to
the endolysosomal compartment where TLR9 meets DNA of
invading pathogens. Upon arrival in the endolysosome, TLR9
undergoes proteolytic cleavage which is the prerequisite for
signalling and the induction of an antiviral response.
Copyright Melanie M. Brinkmann
TLR9 localizes to the endolysosomal compartment after TLR
agonist stimulation. Primary bone marrow derived macrophages
were isolated from TLR9-GFP transgenic mice. Live
cells were stimulated with LPS and analysed with a spinning
disc confocal microscope in real-time. Endolysosomes were
visualised with lysotracker (in red). Copyright Melanie M. Brinkmann
is known about the regulation of intracellular trafficking in
infected antigen presenting cells.
The Project Our research focuses on the mechanisms
herpesviruses evolved to modulate their recognition by the
host’s immune system, thereby allowing them to establish
chronic infections. We have evidence that herpesviruses can
modulate the antiviral response initiated by TLRs. We aim to
identify the responsible herpesviral genes and characterize
their mechanism of action. We further study the regulation
of intracellular trafficking of TLR9 upon infection with
herpesviruses. Using fluorescently labelled virions and
fluorescently labelled TLR9 and UNC93B proteins, the group
will analyse intracellular trafficking in real-time in primary
antigen presenting cells by confocal lasermicroscopy. In a
biochemical approach, using herpesviral DNA as bait, we
aim to identify further DNA sensors that are crucial for the
detection of herpesviruses. Furthermore, we identified novel
interacting partners of the UNC93B protein and will characterize
their role for trafficking of the UNC93B-TLR9 complex
from the endoplasmic reticulum to the endolysosomal compartment
where TLR9 signalling is initiated.
Boyoun Park, Melanie M. Brinkmann, Eric Spooner, Clarissa C. Lee, You-Me Kim and
Hidde L. Ploegh, (2008) Nature Immunology, 9 (12), 1407-1414
You-Me Kim, Melanie M. Brinkmann, Marie-Eve Paquet and Hidde L. Ploegh (2008)
Nature, 452 (7184), 234-238
Melanie M. Brinkmann, Eric Spooner, Kasper Hoebe, Bruce Beutler, Hidde L. Ploegh
and You-Me Kim, (2007) Journal of Cell Biology, 177 (2), 265-275

116 SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups
02 Systems Immunology
PROJECT LEADER | Prof. Dr. Michael Meyer-Hermann | Department of Systems Immunology | mmh10@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Marc Thilo Figge | Dr. Valerii Sukhorukov | Dr. Arndt Telschow | Sebastian Binder |
Jaber Dehghany | Harald Kempf
Systems Immunology What is Systems Biology and why do
we need it? It is not possible to understand the functionality
of a system by just knowing its constituents, as derived from
„omics“-approaches. The major question is how all these
constituents interact with each other and how they give rise
to a meaningful and functional concept like an organism.
The key to this question is the dynamics of the system under
investigation. Systems Immunology is the science of developing
mathematical models of the dynamic behaviour and
interaction of complex systems related to health. The future
of research in Biology will rely on the iterative discussion of
theoretical prediction and experimental result, inspired by
the success of Physics.
of GC. We tested in silico with the aforementioned model
how disruption of TLR4-signalling would change the GC
and predicted that the mutation rate has to be reduced. This
prediction was confirmed in experiment. [3]
Directions The novel department Systems Immunology
at the HZI will work on models of dynamic systems in the
immune system and in the field of infection and inflammation.
In the coming year pre-established research fields, like
modelling of adaptive immune responses and of pancreatic
betacells in the context of diabetes, will be continued and it
is planned to develop new collaborations, in particular, with
emphasis on chronic inflammatory diseases.
Methodology As it is impossible to understand the mechanisms
of a biological system from the knowledge of its
constituents alone, we are restricted to start from phenomenological
descriptions of the whole system. These should
be as simple as possible and as complex as needed to allow
comparison with available experimental data. In an iterative
optimisation process the model assumptions are evolved up
to the point that the in silico model behaves as the in vivo
model. If theory and experiment disagree, research may take
advantage of Systems Immunology: Either the model has to
be extended or an assumption in the model has to be reconsidered.
In both cases the knowledge of how the biological
system works is improved.
The advantage of such a phenomenological approach is that
at any level of mathematical description reached so far, the
in silico model remains functional. Ultimately this approach
based on an increasingly complex picture of the biological
system will enable to (i) design new experiments that are
conclusive for specific questions, (ii) predict the outcome of
planned experiments in order to verify the consistency of the
current knowledge, and (iii) predict optimal treatment strategies
in deregulated systems.
Example generation of specific antibodies The immune
system contains a specific environment, the germinal centre
(GC), in which B lymphocytes (BC) are induced to mutate
their antibody encoding genes. Subsequently, mutated BC
undergo a selection process in order to either produce highaffinity
antibodies or to contribute to the immune memory.
The general conviction was that selection is controlled by
the availability of antigen. We have developed a mathematical
model for the GC dynamics and predicted that it is not
the antigen but affinity dependent TC help that is limiting
[1]. This prediction was verified in a series of experiments
[2] and, thus, a new concept of the GC reaction was induced.
Also toll-like-receptor-4 (TLR4) impacts on the development
In silico agent-based simulation of a germinal centre reaction.
A 20 micron thick slice of the germinal centre at day 5 after
onset of proliferation. Every object represents a cell: Dividing
and mutating BC (blue), non-dividing Ig-expressing BC (dark
green), BC with positive signals from FDC (light green), plasma
cells (white), T follicular helper cells (red), FDC (yellow)
where dendrites are not shown. The cells interact according
rules derived from present knowledge and move in response
to chemokines (not shown) and in agreement with recent intravital
two-photon imaging data. Previously published in [4].
[1] Meyer-Hermann, M., Figge, M.T., & Toellner, K.M. (2009) Trends in Immunology 30, 157-164.
[2] Victora, G.D., Schwickert, T.A., Fooksman, D.R., Meyer-Hermann, M., Dustin, M.L., &
Nussenzweig, M.C. (2010) Cell 143, 592-605.
[3] Garin1, A., Meyer-Hermann1, M., Contie, M., Figge, M.T., Buatois, V., Gunzer, M., Toellner,
K.-M., Elson, G., & Kosco-Vilbois, M.H. (2010) Immunity 33, 84-95. 1: shared first author.
[4] Figge, M.T., Garin, A., Gunzer, M., Kosco-Vilbois, M., Toellner, K.-M., & Meyer-Hermann,
M. (2008). Journal of Experimental Medicine 205, 3019-3029.

SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups 117
03 The National (“Helmholtz”) Cohort
PROJECT LEADER | Priv.-Doz. Dr. Frank Pessler | Department of Infection Genetics | fpe09@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Manas Akmatov | Matthias Preuße
Prospective cohort studies The rise of chronic diseases
such as cancer, atherosclerosis, diabetes, osteoarthritis and
Alzheimer’s disease represents an ever-growing challenge
to the health care systems of industrialized countries. The
causes of these disorders are multi-factorial and consist of
environmental, lifestyle and genetic risk factors. Epidemiologists
are presented with the challenge to develop strategies
for the early detection of risk factors and diseases,
ideally at a time when interventions are most effective.
Because of their prospective character, which allows assessing
risk factors and biomarkers in the preclinical stage of
disease development, cohort studies are the most powerful
tools of modern epidemiology for this purpose.
The National (“Helmholtz”) Cohort The Helmholtz As sociation
has been spearheading a nationwide cohort study to
address the health care challenges of the 21 st century. Ultimately,
this undertaking will involve recruiting 200,000
human volunteers throughout Germany, constructing a
central biobank near Munich, and the follow-up of the
subjects for 10-20 years. A start-up budget of 20 million
Euros from the Helmholtz Association is being used by the
five participating Helmholtz Centres to help plan and pilot
the study protocols for this largest health-research-related
endeavor in all of Germany’s history, to establish and equip
study centres, and to begin recruiting and phenotyping
participants in the middle of 2012.
2009 2012 2017
Planning
& piloting;
feasibility
studies
Recruitment,
construction
of biobank
Follow-up
visit #1
2022
Follow-up
visit #2 (?)
2027
Follow-up
Detection of incident cases of disease
20+
years?
Currently proposed time line for the planning, recruitment
and follow-up phases of the Cohort. Graphic: HZI
The HZI is and will be contributing to the Cohort as follows:
• By recruiting 10, 000 study participants during 2012-
2017 as part of the Recruitment Cluster North-West
Germany.
• By developing and optimizing protocols (SOPs) for collecting
and storing biomaterials, in particular nasal swabs,
saliva, peripheral blood leukocytes, and stool.
• By initiating, developing and supporting aspects of the
Cohort that relate to infectious diseases and disorders of
immunity. Our approach will follow two tiers: firstly, to
evaluate microorganisms as risk factors for the development
of common chronic diseases such as cardiovascular
and neurodegenerative diseases; secondly, to identify risk
factors for the development of acute and chronic infections
and disorders of immunity.
• By implementing novel epidemiologic tools for the study
of infectious diseases and disorders of immunity. For
instance, we are testing whether study participants can
reliably collect their own biospecimens for diagnostic
testing (e.g., nasal swabs or stool samples) during an
acute illness and then send it to the reference lab. In
the same projects, we are studying the use of modern
communication tools such as SMS and email as reminder
systems to maintain compliance and to obtain real-time
information about symptomatic infections.
• By evaluating and adapting novel biomarkers (e.g., micro-
RNAs) and biostatistical approaches.
Networking within the Cohort We represent the HZI in
the national Epidemiologic Planning Committee for the
Cohort, in the writing committee for the international
evaluation which is anticipated in 2011, and in the Thematic
Working Groups “Infection & Immunity”, “Musculoskeletal
Disorders”, “Biobank” and “Questionnaires”. In the current
feasibility phase of the Cohort we are coordinating a multicentre
feasibility study on novel measurement instruments
for infectious disease epidemiology.
Akmatov, M.K. & Pessler F. (2011) The potential of self-collected nasal swabs to detect
infection and colonization in population-based epidemiological studies. International
Journal of Infectious Diseases (accepted for publication).
Akmatov, M.K., Pessler, F. & Brzoska P. (2011) Attitudes among women in low- and
middle-income countries towards people living with HIV/AIDS – the situation after
three decades of AIDS. BMC Public Health (under review).
Ogdie, A., Li, J., Dai, L., Yu, X., Diaz-Torne, C., Akmatov, M., Schumacher, H.R., & Pessler,
F. (2010) Identification of broadly discriminatory synovial tissue biomarkers with
binary and multicategory receiver operating characteristic analysis. Biomarkers 15(2),
183-90.
Slansky, E., Li, J., Häupl, T., Morawietz, L., Krenn, V., & Pessler, F. (2010) Quantitative
determination of the diagnostic accuracy of the synovitis score and its components.
Histopathology 57(3), 436-443.

118 SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups
04 Immune Aging
PROJECT LEADER | Dr. Dr. Luka Cicin-Sain | Junior Research Group Aging and Chronic Infections |
lcs09@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Linda Ebermann | Iryna Dekhtiarenko | Anja Drabig | Thomas Marandu | Ilona Paprotta |
Franziska Schreiner
The loss of immunological function caused by the aging
process is termed immune senescence. There is a consensus
that the adaptive immune response to emerging infections is
compromised in the elderly. Naïve lymphocytes can recognize
a broad and divergent array of potential pathogens
because they are clonally diverse. Aging results in a loss
of naïve T-cells and of T-cell clonal diversity, increasing the
susceptibility to novel infections and decreasing the ability
to generate protective immunity upon a vaccination. Therefore,
the loss of clonal diversity in the T-lymphocyte pool is a
prominent change associated with immunosenescence. The
causes of immune senescence remain incompletely understood.
While thymic involution plays a prominent role in the
aging process, the role of the oxidative metabolism and environmental
stressors remain less defined. Understanding the
key environmental contributors to the aging process would
allow us to define targets for strategies delaying immune
senescence.
Cytomegalovirus Cytomegalovirus (CMV) is a ubiquitous
pathogen, infecting the majority of the human population
worldwide. CMV establishes a lifelong latent infection in
infected hosts, and can reactivate in immunosuppressed or
immunodeficient individuals. Interestingly, CMV elicits a
T-cell response that is unparalleled in its strength, which
dominates the repertoire of the memory T-cell compartment,
and where CMV-specific CD8 cells appear to accrue
in older age groups. Hence, the immune system may devote
a significant fraction of its adaptive capacity to the control
of one single pathogen and the mechanisms leading to these
uniquely strong immune responses may have consequences
in our understanding of the homeostatic processes that
govern the interaction between a host and its microbiological
environment.
The Project Epidemiological longitudinal studies of elderly
volunteers identified immune-risk phenotypes that predicted
poor survival, and these phenotypes coincided with
seropositivity to (CMV). If a ubiquitous agent, such as CMV,
contributed to immune aging, this would have a huge impact
on our understanding of the immune aging process in the
general population.
Our preliminary data show that mice carrying latent CMV
exhibit weaker CD8 responses to superinfections with
emerging pathogens, such as Influenza Virus, than control
(uninfected) animals. Therefore, chronic CMV infection may
contribute to the immune aging process. Our group aims to
define the effects of persistent virus infections on the host
immune system, by addressing two specific objectives: I. elucidate
the cellular and molecular mechanisms of the induction
of the adaptive immune system by CMV infection, and II.
assess the broader biological relevance of this phenomenon.
I. Mechanism: It is not clear if the poor CD8 response in
CMV infected mice is indeed caused by changes intrinsic
to the CD8 subset, or by changes in their immunological
environment. This question can be addressed by adoptive
transfer experiments. Moreover, it is not clear if this loss is
due to persistent antigenic stimulation or to inflammation
caused by the virus. This can be defined by recombinant
viruses with conditional and/or gene-specific expression of
immunodominant T-cell Ag.
II. Relevance: It is not clear if the poor immune response
relates exclusively to virus challenges of CMV infected hosts,
or if this is a broader phenomenon. We will test the T-cell
repertoire and immune function in other chronic infections,
and we will test if the chronic infections will result in poor
cellular immune responses to non-viral challenges, such as
challenges with (a) bacterial intracellular pathogens (e.g.
Listeria monocytogenes, Mycobacterium bovis), or (b) tumour
antigens.
1.Cicin-Sain, L., Bubic, I., Ruzsics, Z., Schnee, M., Mohr, C., Jonjic, S. & Koszinowski, U.H.
(2007) Targeted deletion of regions rich in immune-evasive genes from the cytomegalovirus
genome as a novel vaccine strategy. Journal of Virology 81(24),13825-34.
2.Cicin-Sain, L., Messaoudi, I., Park, B., Currier, N., Planer, S., Fischer, M, Tackitt, S.,
Nikolich-Zugich, D., Legasse, A., Axthelm, M.K., Picker, L.J., Mori, M. & Nikolich-Zugich J.
(2007). Dramatic Increase in Naïve T-Cell Turnover is Linked to Loss of Naïve T-cells from
Old Primates. PNAS 104(50),19960-5.
3.Cicin-Sain, L., Smyk-Pearson, S., Fisher, M.B., Currier, N., Koudelka, C., Nikolich-Zugich,
D., Legasse, A., Tackit, S., Axthelm, M.K., Mori, M. Lewinsohn, D., & Nikolich-Zugich, J.
(2010) Loss of naïve T-cells and repertoire constriction predict poor response to vaccination
in old primates. Journal of Immunology 184(12), 6739-45.

SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups 119
05 Development of Novel Diagnostic and Therapeutic
Tools for Tuberculosis Control and Prevention
PROJECT LEADER | Prof. Dr. Mahavir Singh | Department of Gene Regulation and Differentiation |
msi@helmholtz-hzi.de
PROJECT MEMBERS | Cyril Alozieuwa | Dr. Sabin Bhuju | Ayssar Elamin | Dr. Matthias Stehr | Hanaa Wanas
The aim of the project is to develop new diagnostics, therapeutics
and vaccines against tuberculosis and HIV. For the diagnosis
and the development of a vaccine, we aim to identify
new, highly specific antigens. The search for lead structures
is carried out by the screening of compound libraries against
virulence associated proteins and enzymes of the pathogen.
Tuberculosis Tuberculosis is a bacterial infectious disease.
Every year two million people die from tuberculosis and
another nine million are newly infected. Africa and Asia
are the most affected regions in the world. In Germany the
number of new cases of tuberculosis is declining. However,
about 15 years ago new, multi-resistant strains have emerged.
The so-called MDR (multidrug resistant) and XDR (Extreme
Drug Resistant) strains spread rapidly and can already be
found worldwide. Most of the known antibiotics are not effective
against MDR and XDR strains, thus new highly effective
substances must be developed in order to prevent the further
spread of drug resistant tuberculosis.
Identification of new leads for the development of new
antibiotics Supported by the EU-funded project “NOPERSIST”
we are identifying new lead compounds that act against two
virulence associated target enzymes of the pathogen. The
alanine dehydrogenase (AlaDH) synthesizes building blocks
for cell wall biosynthesis. We have developed a method,
which is capable to screen around 10,000 compounds from
the HZI compound library per week. So far we have identified
50 candidate inhibitors which are currently validated in
follow-up assays for further analysis.
The second target is the thiol peroxidase (TPX), an enzyme of
the mycobacterial peroxide defense system. Currently, we are
developing a high throughput assay for substance screening
against the TPX enzyme.
New antigens for rapid diagnosis tools and vaccine
candidates For tuberculosis control modern and rapid molecular
diagnostic tools are needed. Current methods take up
to four weeks. The high-throughput cloning and expression
allows the production of hundreds of pure mycobacterial proteins
per week. The cellular and humoral immune response
to these candidate proteins is analysed by the partners of
the consortium of the EU funded project”NOPERSIST”. From
the pool of candidate proteins the vaccine and diagnostic
candidates will be identified.
Transvac We are partners in an infrastructure project
“TRANSVAC” funded by the EU. Transvac is the European
platform for vaccine development for HIV, TB and malaria.
We are currently performing transcriptome mapping of samples
from BCG vaccinated human individuals and healthy
controls.
Schematic representation of the structure of the drug target
alanine dehydrogenase. The bound cofactor NAD+ and
pyruvate are depicted as ball-and-stick. (Ågren, D., Stehr, M.,
Berthold, C.L., Kapoor, S., Oehlmann, W., Singh, M. and Schneider,
G. (2008) Three-dimensional structures of apo- and
holo-L-alanine dehydrogenase from Mycobacterium tuberculosis
reveal conformational changes upon coenzyme binding.
Journal of Molecular Biology, 377, 1161-1173.)
Rosenkrands, I., Aagaard, C., Weldingh, K., Brock, I., Dziegiel, M.H., Singh, M., Hoff, S.,
Ravn, P., & Andersen, P. (2008) Identification of Rv0222 from RD4 as a novel serodiagnostic
target for tuberculosis. Tuberculosis (Edinb) 88, 335-343.
Ågren, D., Stehr, M., Berthold, C.L., Kapoor, S., Oehlmann, W., Singh, M., & Schneider, G.
(2008) Three-dimensional structures of apo- and holo-L-alanine dehydrogenase from
Mycobacterium tuberculosis reveal conformational changes upon coenzyme binding.
Journal of Molecular Biology 377, 1161-1173.
Elamin, A.A., Stehr, M., Oehlmann, W., & Singh, M. (2009) The mycolyltransferase 85A,
a putative drug target of Mycobacterium tuberculosis: Development of a novel assay and
quantification of glycolipid-status of the mycobacterial cell wall. Journal of Microbiological
Methods 24, 358-363.
Von Groll A., Martin A., Stehr M., Singh M., Portaels F., da Silva P.E., & Palomino J. C.
(2010) Fitness of Mycobacterium tuberculosis strains of the W-Beijing and Non-W-Beijing
genotype. PLoS One 5, e10191.

120 SCIENTIFIC REPORTS | Infection and Immunity | New Project Groups
06 Exploring and Exploiting Microbial Proteomes
PROJECT LEADER | Prof. Dr. Katharina Riedel | Research Group Microbial Proteomics |
katharina.riedel@helmholtz-hzi.de | ak.riedel@tu-bs.de
PROJECT MEMBERS | Dr. Isabel Hartmann | Dr. Martin Kucklick | Thomas Langer | Christian Lassek
Proteomics – a powerful tool to investigate microbial
pathogens Proteins are the effector molecules that mediate
basic cellular functions and are often the direct targets
of antimicrobial drugs. The analysis of the entirety of all
expressed proteins of microbes (“microbial proteomics”) has
been proven as an ideal tool to investigate various aspects in
medical microbiology, e.g. the molecular basis of bacterial
infections or the adaptation of microbes to antibiotic stresses.
We employ two-dimensional polyacrylamide gel electrophoresis,
gel-free liquid chromatography, and mass spectrometrybased
technologies for microbial proteome analyses.
Evaluation of novel anti-infectiva The emergence of drugresistant
microbes has led to an urgent need of novel anti-infective
agents. Cell-to-cell communication systems (quorum
sensing, QS), which operate in many pathogens as central
regulators for the expression of virulence factors and biofilm
formation, represent attractive targets for the design of novel
therapeutics. The major advantage of this strategy is that it
circumvents the problem of resistance, which is intimately
connected to conventional antibiotics. We have developed
specific sensor strains to screen compound libraries for their
potential to interfere with QS of two important pathogens,
Pseudomonas aeruginosa and Burkholderia cenocepacia. To
elucidate the compounds efficacy and specificity comparative
proteome analyses are employed. Moreover, in close
collaboration with Prof. J. Robinson (University of Zürich) we
have recently unraveled the mode-of-action of peptidomimetics
that specifically inhibit P. aeruginosa.
In situ proteomics to study the molecular basis of microbial
infections Eighty percent of microbial infections in
humans are mediated by biofilms which are highly resistant
against antimicrobial therapies. Deciphering the crucial
command lines of biofilm formation under infection-like conditions
will therefore contribute to a better understanding of
bacterial pathogens. We have established in situ proteome
analyses of P. aeruginosa during colonization of non-mammalian
and mammalian pathogenicity models to identify
virulence factors specifically expressed during the infection
process and to observe changes in the host proteome
induced by the bacteria. A long term goal of the collaborative
project with Prof. M. Givskov (University of Copenhagen) is
to apply our in situ proteomics analyses for target validation
of linked multi-drugs, which exhibit two or more antimicrobial
features.
Workflow of a typical proteomics experiment starting with
protein extraction, followed by separation via gel-based or gelfree
approaches, identification & quantification by mass spectrometry,
and data validation by complementary approaches
(e.g. phenotypical assays).
Metaproteome analyses of uropathogenic catheter biofilms
Urinary tract infections are among the most common
nosocomial infections. Because of the ability of uropathogens
to develop resistances during antibiotic treatment, catheter
biofilms are difficult to eradicate and often become chronic.
P. aeruginosa as well as Escherichia coli have been reported
to cause about 40% of complicated catheter-associated infections;
however, recent investigations indicate that catheter
biofilms are highly complex and consist of various different
species. Our group is part of the UroGenOmics consortium,
which aims on the elucidation of regulatory and metabolic
strategies during colonization and infection. We are currently
employing metaproteome analyses to investigate
the interplay between composition and function of mixed
catheter biofilms, to investigate temporal changes and spatial
distributions during the infections process, and to unravel
strain-specific strategies to adapt to urinary tract conditions.
The gained knowledge will help to develop new analytical
tools, to identify novel drug targets, and to improve catheter
surface design.
Srinivas, N., Jetter, P., Ueberbacher, B.J., Werneburg, M., Zerbe, K., Steinmann, J., Van der
Meijden, B., Bernardini, F., Lederer, A., Dias, R.L., Misson, P.E., Henze, H., Zumbrunn, J.,
Gombert, F.O., Obrecht, D., Hunziker, P., Schauer, S., Ziegler, U., Käch, A., Eberl, L., Riedel,
K., Demarco, S.J., Robinson, J.A. (2010). Peptidomimetic Antibiotics Target Outer-Membrane
Biogenesis in Pseudomonas aeruginosa. Science 327: 1010-1013.
Schneider, T., Riedel, K. (2010). Environmental proteomics: Analysis of structure and
function of microbial communities. Proteomics 10: 785-798.
Riedel, K., Köthe, M., Kramer, B., Saeb, W., Gotschlich, A., Ammendola, A., Eberl, L. (2006).
Computer-aided design of agents that inhibit the cep quorum sensing system of Burkholderia
cenocepacia. Antimicrob. Agents Chemotherapy 50: 318-323.

SCIENTIFIC REPORTS | PoFII Independent Research
121
POFII INDEPENDENT RESEARCH
Some projects at the HZI are indepedently performed of the PoF-programme. In this Research Report we present three projects.
Ken Timmis, who is going to retire in 2011, presents fi nal results from the research of his group on “Functional genomics and
niche specifi ty”.
Inari Kursula is coordinating a research group within the newly established “Centre for Structural Systems Biology” (CSSB) at the
German Electron Synchrotron (DESY), Hamburg. This is a new external HZI research group after a joint venture agreement was
signed in 2010. She is presenting results on “Structural biology of the cytoskeleton”.
Wolf-Dieter Schubert, until 2009 head of a research group at the HZI, presents research results which he obtained jointly with the
group of Prof. Dirk Heinz fi nalizing his former activities at the HZI: “Structural analysis of the innate immune system”.
An aerial photograph of the German Electron Synchrotron, Hamburg, where Prof. Inari Kursula, HZI, is working. Photo: DESY

122 SCIENTIFIC REPORTS | PoFII Independent Research
01 Functional Genomics and Niche Specificity
PROJECT LEADER | Prof. Dr. Kenneth N. Timmis | Research Group Environmental Microbiology |
kti@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Sagrario Arias Rivas | Dr. Mónica Bassas Galia | Simrita Cheema |
Dr. Thorben Dammeyer | Dr. Isabel Hartmann | Dr. Stefanie Libnow | Dr. Gabriella Molinari | Dr. Daniela Näther |
Kateryna Selezska | Dr. Johannes Sikorski | Dr. Joao Vieira de Castro | Montri Yasawong.
The microbial biota of the human gastro-intestinal (GI) tract
has been designated an additional human organ and a major
compartment of the human “biome”, to reflect the multitude
of diverse contributions and services it provides and the extent
to which it influences the human phenotype. Despite its
key roles in health and disease, it remains a diverse, variable,
difficult to study and poorly understood entity. Current large
scale metagenome projects aim to shed light on the community
composition and functional genomics of these microbial
ecosystems, but tend to focus on the abundant members,
which are more accessible and considered to play the more
important roles.
Inflammatory bowel disease (IBD), encompassing the syndromes
Crohn’s disease, ulcerative colitis and other undetermined
forms of colitis, are complex, multifactorial inflammatory
diseases. They are known to be determined by multiple
genetic loci and thought to triggered and/or exacerbated by
the microbial gut flora. As part of a collaborative study with
the University Clinic Kiel (Schreiber, Ott) and the University
of the Balearic Isles (Nogales, Lanfranconi) seeking microbial
correlates of intestinal bowel disease, we obtained mucosal
biopsies from the inflamed parts of the sigmoid region of
the colon of patients suffering from diverse forms of IBD.
We established 16S ribosomal RNA gene clone libraries,
sequenced the clones, and determined their phylogenetic
assignments. Though bacteria dominate the colonic microbial
community, archaeal methanogens are minor members.
Therefore, in addition to creating libraries of bacterial 16S
rRNA gene sequences, we also created libraries of archaeal
16S rRNA genes obtained by low-stringency amplification using
archaeal domain primers. Surprisingly, this yielded from
a number of biopsies, in addition to expected sequences
of methanogens, sequences of members of the Halobacteriaceae.
This was unexpected not only because the GI tract
is not considered to be a salty environment and had not
previously been reported to harbour halophilic archaea, but
also because of the high diversity and novelty of the cloned
sequences: 269 Halobacteriaceae-related sequences obtained
from the biopsies grouped into 15 distinct phylotypes (which
covered almost the entire phylogenetic range of this family),
9 of which were new.
Colonoscopy requires prior intestinal lavage with a solution
containing polyethylene glycol and salts, so one possibility
was that the haloarchaeal sequences originated from the
lavage solution. We obtained two unopened lavage preparations
from the hospital in which the biopsies were collected
and, for comparison, one from the same manufacturer from a
pharmacy in Palma, Mallorca, and one from a different manufacturer
from the same pharmacy, and repeated the DNA
isolation, amplification and sequencing of library clones. We
also analysed 8 fecal specimens from IBD patients who had
not undergone intestinal lavage. Halobacteriaceae-like sequences
were obtained from 2 of the 8 fecal samples (76 sequences)
and from all of the lavage samples (125 sequences),
the latter of which grouped into 17 phylotypes, none of which
corresponded to phylotypes detected in patient samples.
Another potential origin of halophiles in the human intestine
is the table salt assimilated in food. We repeated the procedure
used above with 3 table salt samples and obtained 34
haloarchaeal sequences representing 11 phylotypes. None of
these was the same as any phylotype from the lavage preparations,
and only one was the same as that from a patient.
The experimental approach used does not distinguish
between dead cells/free DNA and viable metabolically active
cells. Therefore we set up enrichment cultures for halophiles
from a patient biopsy. This yielded haloarchaeal sequence
types, and fluorescence in situ hybridization analysis subsequently
revealed viable archaeal cells in these cultures.
Thus: while Haloarchaea are continuously introduced into
the GI tract via food, the lack of relatedness of table salt and
GI tract phylotypes found here failed to identify either table
salt or lavage solutions as a source of GI tract phylotypes.
These results point to the possibility that haloarchaea are
functional members of the human colonic biota, which would
seem to be counter-intuitive, given that this environment is
not considered to be particularly salty. The colon environment
is however rather heterogeneous: while the intestinal
contents of healthy individuals have an average salinity similar
to that of plasma (135–145 mM sodium), small pockets of
concentrated luminal ions (reported to be in the order of 15
mM above that of the surrounding fluids) may form within
the lateral intercellular spaces (LIS) and crypts as part of
the normal processes of fluid transport in the intestinal
epithelium. Regions like the LIS may thus provide suitable
micro-niches favourable for colonization by haloarchaea. If
the haloarchaea discovered here in colonic biopsies are
indeed active, then their metabolism and products may have
significant consequences for mucosal physiology in general,
and the diseased bowel, in particular, and vice versa – as has
been previously suggested for methanogens in conditions
of colorectal cancer and diverticulosis. In this regard, it is
worth noting that altered bowel physiologies are a prominent
feature of IBD. Impaired absorption of sodium and organic
solutes, like short-chain fatty acids such as butyrate, result
in increased solute concentrations and osmolarity in the
lumen. These results suggest that investigation of a potential
physiological basis of the haloarchaeal-intestinal mucosal association
may lead to new insights into GI health and disease.

SCIENTIFIC REPORTS | PoFII Independent Research 123
02 Structural Biology of the Cytoskeleton
PROJECT LEADER | Prof. Dr. Inari Kursula | Centre for Structural Systems Biology (CSSB) at DESY, Hamburg |
iku09@helmholtz-hzi.de
PROJECT MEMBERS | Dr. Alexander Ignatev | Dr. Petri Kursula | Dr. Thorsten Mengesdorf | Saligram Prabhakar Bhargav |
Moon Chatterjee | Gopinath Muruganandam | Nele Vervaet
We are interested in how cytoskeletal proteins recognize
and bind to each other and how actin polymerization in
eukaryotic cells is regulated. In particular, we want to understand
how pathogenic parasites use their actin cytoskeleton
for motility and host cell invasion and to find ways to
interfere with these processes. Another focal point in our
work is histone deacetylases, especially those with cytosolic
substrates. We use X-ray crystallography and small-angle
X-ray scattering for protein structure determination and
complementary biophysical and biochemical methods for
the functional characterization of proteins and complexes.
We are also interested in the development of new synchrotron-based
applications for visualizing large cytoskeletal
complexes at high resolution.
Actin-based motility of the malaria parasite Malaria is
one of the most devastating worldwide health threats. More
than a million people die of malaria and up to 300 million
people are infected every year, most of them young children
and pregnant women. Malaria is also a significant economical
burden, trapping the poorest areas in a downward spiral
of poverty. Resistance to existing anti-malarial drugs is
a growing problem in nearly all malaria-endemic areas.
Therefore, there is an urgent need for new drug and vaccine
candidates.
Malaria is caused by Plasmodium spp., a group of unicellular,
eukaryotic, intracellular parasites, belonging to the
phylum Apicomplexa. These parasites use actin for both
motility and host cell invasion. Their cytoskeleton differs
significantly from that of higher eukaryotes and is,
therefore, an attractive target for anti-malarial research.
The parasite actin filaments are extremely short and their
rapid turnover is regulated by a limited set of actin-binding
proteins, which are poorly conserved with their mammalian
homologues.
Plasmodium actin-binding proteins with divergent
structures and functions Profilins are ubiquitous, small,
actin-binding proteins that work together with formins to
facilitate actin polymerization by recruiting polymerizable
actin monomers close to the growing end of the filament.
We have determined the crystal structure of P. falciparum
profilin (PfPfn) and shown that it possesses the key functionalities
of profilins and is essential for the survival of
the parasite. Structurally, PfPfn differs significantly from
all other profilins. The large structural rearrangements
close to the actin-binding face point towards a possibly
novel binding mode to actin. Our work aims to find out the
regions required for the interaction between Plasmodium
profilin and actin and to determine the three-dimensional
structure of the complex.
Once we have the crystal structure available, it is possible
to start looking for compounds, which specifically disrupt
the Plasmodium profilin-actin complex. We are also working
on the two Plasmodium forming isoforms to find out what
their roles in the regulation of actin polymerization are and
if they work together with profilin in these parasites.
Plasmodium has two actin depolymerization factors (ADFs),
which differ from each other in structure and function. The
major isoform in apicomplexan parasites, ADF1, is essential
for survival, does not bind F-actin and seems to work rather
as a nucleotide exchange factor than a filament destabilizing
protein. ADF2 is present only in Plasmodium spp. and is not
essential but is structurally similar to the canonical members
of the ADF family.
a) We have determined the crystal structure of Plasmodium
falciparum profilin in complex with an octa-proline peptide.
The structure contains a large b-hairpin insertion, which is not
present in any other profilins, as seen in the superposition of
Plasmodium and mouse profilins (small picture below, where
green is Plasmodium and gray mouse profilin).
b) Homology modeling suggests that the b-hairpin protrusion
may be involved in actin binding.
c) A close-up view of the putative interactions of the b-hairpin
protrusion in Plasmodium profilin (green) with actin (gray).
The superimposed mouse profilin is shown in pink.
Graphics: Inari and Petri Kursula
Kursula, P., Kursula, I., Massimi, M., Song, Y.H., Downer, J., Stanley, W.A., Witke, W. &
Wilmanns, M. (2008) High-resolution structural analysis of mammalian profilin 2a
complex formation with two physiological ligands: formin homology 1 domain of mDia1
and the proline rich domain of VASP. Journal of Molecular Biology 375, 270-290.
Kursula, I., Kursula, P., Ganter, M., Panjikar, S., Matuschewski, K. & Schüler, H. (2008)
Structural basis for parasite-specific functions of the divergent profilin of Plasmodium
falciparum. Structure 16, 1638-1648.
Huttu, J., Singh, B., Bhargav, S.P., Sattler, J., Schüler, H. & Kursula, I. (2010) Crystallization
and preliminary structural characterization of the two actin depolymerization
factors of the malaria parasite. Acta Crystallographica Section F 66, 583-587.

124 SCIENTIFIC REPORTS | PoFII Independent Research
03 Structural Analysis of the Innate Immune System
PROJECT LEADER | Prof. Dr. Wolf-Dieter Schubert | Former Research Group Molecular Host-Pathogen
Interactions | Department of Biotechnology, University of the Western Cape, Cape Town, South Africa |
wschubert@uwc.ac.za
PROJECT MEMBERS | Nils Kuklik | Clive Mketsu | Mujaahida Mohammed | Edukondalu Mullapudi | Lilia Polle |
Donné Simpson | Jason Stark
In the Research Group “Molecular Host-Pathogen Interactions”
(MHPI) we previously focussed on the molecular
details of human defence mechanisms against invading
pathogens (innate immunity) as well as the corresponding
molecular strategies of pathogens in infecting humans.
Novel Listerial Virulence Factors InlJ and Auto The bacterium
Listeria monocytogenes infects humans following the
consumption of contaminated food. To breach the intestinal
barrier and for all steps of the infection process Listeria has
evolved a dedicated set of highly specific protein factors that
it produces and secretes in a highly coordinated manner. To
understand how their physical structure relates to their physiological
function, we have undertaken to investigate their
three-dimensional structure at the atomic level of resolution.
“Auto” The cell wall of Gram-positive bacteria is a complex
envelope that helps cells to maintain their shape and to
counteract the turgor pressure of their cytosol. It consists of
long, linear chains of alternating sugar residues (N-acetylglucosamine
and N-acetylmuramic acid) crosslinked by short
peptides. Cell growth, cell division and many other processes
crucially depend on the constant remodelling of the cell wall.
Enzymes involved in this process are generally referred to as
autolysins.
Studies revealed that only one of the autolysins is involved
in the pathogenesis of Listeria monocytogenes. This autolysin
named “Auto” is essential for listerial entry into numerous
eukaryotic cell lines.
Left: Crystal structure of autolysin Auto of L. monocytogenes
is depicted in cartoon style and with transparent surface.
The active site of the pro-enzyme is blocked by an N-terminal
α-helix (red). Right: Crystal structure of InlJ (right) with enlarged
view of the cysteine ladder (left): reduced cysteines align
in the hydrophobic core (Sulphur - yellow). Graphics: HZI
Auto is an N-acetylglucosaminidase, meaning that it cleaves
the sugar backbone of the cell wall exposing the reducing
end of N-acetylglucosamine. Solving the crystal structure
of Auto we observed that it is structurally related to lytic
transglycosylases and some lysozymes despite catalysing an
unrelated chemical reaction. By mutating residues potentially
involved in catalysis, we identified the glutamic acid residues
Glu122 and Glu156 as being essential for the enzymatic
reaction. Fascinatingly, we found that Auto, as originally produced,
is autoinhibited by an N-terminal α-helix. Its specific
N-terminal cleavage by an as yet unidentified protease following
an extracellular signal may, therefore, allow its rapid
and coordinated activation. In addition, Auto has a highly
acidic pH optimum making it essentially inactive at a pH of 7.
Overall we interpret our data to indicate that Auto participates
in liberating the pathogen from the partially acidified
phagolysosome. By increasing the porosity of the cell wall it
would help to coordinate the release of other virulence factors
such as listeriolysin. Their activity in turn would result
in the breaching of the phagolysosomal membrane, raising
the pH to physiological levels and de-activating Auto.
InlJ The internalin family of proteins constitutes a group
of virulence factors of Listeria monocytogenes. The founding
member, Internalin or InlA, is central to the forced uptake
of the bacterium into epithelial cells of the human small intestine.
Only recently this family was found to possess an additional,
previously unrecognized member, InlJ. This protein
has been shown to be crucial to bacterial adhesion to host
cells. InlJ is distinct from other family members in that its 16
LRR units mostly comprise 21 rather than the standard 22
residues and by one LRR-defining hydrophobic residue being
replaced with a conserved cysteine. Solving the crystal structure
of this protein we could show that the cysteines line up
to form a unique cysteine ladder in consecutive repeats. InlJ
thus presents a very rare case of an extracellular protein
(oxidizing environment) bearing a large number of reduced
cysteine residues. This implies that the cysteines must serve
to significantly stabilize the structure, offsetting the danger
of its unfolding followed by cysteine oxidation.
Bröcker, M.J., Schomburg, S., Heinz, D.W., Jahn, D., Schubert, W.-D., & Moser, J (2010)
Crystal structure of the nitrogenase-like dark operative protochlorophyllide oxidoreductase
catalytic complex (ChlN/ChlB)2. Journal of Biological Chemistry 285, 27336-27345.
Heinemann, I.U., Schulz, C., Schubert, W.-D., Heinz, D.W., Wang, Y.G., Kobayashi, Y.,
Awa, Y., Wachi, M., Jahn, D., & Jahn, M. (2010) Structure of the heme biosynthetic
Pseudomonas aeruginosa porphobilinogen synthase in complex with the antibiotic
alaremycin. Antimicrobial Agents and Chemotherapy 54, 267-272.
Bublitz, M., Polle, L., Holland, C., Heinz, D.W., Nimtz, M., & Schubert, W.-D. (2009)
Structural basis for autoinhibition and activation of Auto, a virulence-associated peptidoglycan
hydrolase of L. monocytogenes. Molecular Microbiology 71, 1509–1522.

SCIENTIFIC REPORTS | Technological Platforms
125
TECHNOLOGICAL PLATFORMS
A number of platform technologies essential for research and development carried out at the Helmholtz Centre for Infection
Research are made available to the scientifi c projects as centralised facilities. In the context of national and international research
programmes, these platforms provide services not only to internal projects, but also to scientifi c collaborators from other Helmholtz
research centres, universities, other public research institutes, and industry. On the following pages the most important
platforms are described in detail.
Dr. Matthias Negri and Jennifer Hermann, HIPS, discussing a drug-target interaction. Photo: HIPS/HZI

126
SCIENTIFIC REPORTS | Technological Platforms
01 Central Animal Facility
HEAD | Dr. Hermann Riedesel | Central Animal Facility | hri07@helmholtz-hzi.de
The Central Animal Facility provides state-of-the-art
laboratory animal care and service for the scientific needs
of the HZI in compliance with the animal welfare guidelines.
This facility consists of 24 animal and 13 adjacent
procedure rooms with a total capacity of 15,000 cages which
equals 37,500 mice. Exclusively laboratory mice are housed
as single species. All animals are kept in individually
ventilated cage systems in seven holding units of five
different hygienic and three biosafety levels. Five units are
constructed as a complete specific-pathogen-free (SPF)-
barrier, two units are registered as animal biosafety level 2
(ABSL2) and one as ABSL3 facility for infection experiments.
The average daily animal census is currently about
15,000 mice consisting of 450 different strains and
genetically engineered lines. The health status, which is
monitored on a quarterly basis, complies with the FELASA
recommendations.
The following services are offered:
• Basic animal care
• Breeding services and colony management
• Assistance with experimental procedures
• Health monitoring
• Animal procurement
• Organization of national and international mouse
shipments
• Training programme for animal care technicians
• Animal welfare services for about 60 animal experimental
protocols1
• Consultation and training in laboratory animal science
• Quarantine and rederivation of imported 50 lines
• In vitro fertilization (IVF) for rescue, speed expansion and
rederivation of mouse lines
• Embryo-cryopreservation of mouse lines and banking of
germplasm
• Generation of knock out lines by blastozyst injektion
In 2010 the new mouse facility T2 has been put into operation
starting with the core breeding unit in May. By the end
of July all breeding colonies were completely housed in this
new unit. In October the S2 unit in T2 has been started. The
S3 unit will be operative by the beginning of 2011. After
moving out most of the animals, the T1 building will be
refurbished and reorganized for future use as experimental
holding facility. One floor of this building can be used for
preliminary S2 experiments with animals from outside
sources. This unit harbours an imaging unit equipped with
an IVIS system.
In 2010 the Transgenic Unit has successfully generated 30
new knock out lines. 50 mouse lines from four new groups
have been successfully imported into our breeding units by
means of IVF and/or Embryo transfer. The 2 step controlled
rate freezing method with propandiole as cryoprotectant
was successfully implemented and is now also working successfully
in combination with IVF.
Animal house at TWINCORE At the TWINCORE Hanover
the refurbished mouse facility was commissioned in summer
and put into operation in September 2010. This satellite
mouse facility consists of four animals rooms with floor
area of 96,5 m 2 and a cage capacity of 2,300 IVC cages and
has ABSL 1, 2 and 3 capabilities. The core breeding of all
TWINCORE lines is done at the HZI, so that the TWIN-
CORE facility will be mainly used for expansion breeding
and experimental holding. By this means mice kept at the
TWINCORE will exhibit the same hygienic status as at the
HZI. Four animal technicians are working at the TWIN-
CORE facility offering basic animal care, breeding and
colony management and experimental assistance. All the
other above mentioned services are performed by the HZI
animal facility.
The mouse house. Photo: HZI, Krämer

SCIENTIFIC REPORTS | Technological Platforms 127
02 Recombinant Protein Expression
HEAD | Dr. Joop van den Heuvel | Research Group Recombinant Protein Expression | jvh@helmholtz-hzi.de
SCIENTIFIC COLLABORATOR | Dr. Konrad Büssow | Dr. Volker Jäger | Prof. Dr. Ursula Rinas
The recombinant protein expression platform (RPEX) is
essential for the production of ultra pure protein for high
resolution structural analysis using X-ray crystallography
and NMR spectroscopy. Four major expression systems have
been established in the RPEX facility: E. coli, P. pastoris,
insect cell and mammalian cell culture. This allows the
production of “simple proteins” as well as proteins with
complicated modifications.
For production of the challenging mammalian protein
targets an animal cell culture facility has been established.
The size of the cell culture reactors ranges from 1.6 to 6
litres in order to meet the increasing requirements for
routine production of proteins on a 10-50 mg scale. For the
purification of poorly expressed recombinant multi-protein
complexes a 30-litres-bioreactor has been set up that allows
the pilot-scale production of protein from several hundred
litres of medium.
The system includes all the necessary equipment for continuous
media exchange via tangential flow microfiltration
or internal membrane perfusion and subsequent protein
purification steps. As a supplement to recombinant protein
production, large amounts of HeLa cell biomass can be
produced as a source for purification of naturally occurring
human protein complexes.
Research A research group within the PSPF is focussing
on the development of new and fast strategies for creating
stable expression cell lines. This group has already established
a new and unique methodology for rapidly generating
highly overproducing stable mammalian cell lines (RMCE)
(Wilke et al., 2010). The expression level varies between
1-5 mg/L and can be easily, and reproducibly, scaled up to
10-100 L. In cooperation with the project group of K. Büssow
the newly developed CHO-Lec-RMCE cell lines will be
improved in robustness and validated for their efficiency
of generating a substantial set of new expression cell lines.
The performance of these production cell lines in cell culture
bioreactors will be characterized. Development of new
cell lines for the co-expression of multi-protein complexes
on the basis of the CHO-Lec-RMCE cell lines will be one of
the major challenges in future.
PSPF-Training course in basic techniques in insect cell
cultivation and baculoviral expression of recombinant protein.
Photo: HZI, Bierstedt
Support The Protein Sample Production Facility (PSPF)
is a part of the HZI protein expression platform and was
established in 2007 in cooperation with the Max Delbrück
Centre for Molecular Medicine (MDC). The main goal of this
Helmholtz cooperation is the support of structural biologists
in Germany via circumvention of the major bottleneck of
protein production. The PSPF service is offered either on a
subsidised fee-for-service billing scheme or as a scientific
cooperation (www.pspf.de). Currently, 50% of the capacity of
the unit is used for internal HZI projects.
During 2010, a total of 20 PSPF projects have been processed.
Eight of these projects have been successfully finished, four
have been terminated and eight are still in progress. In 2010,
three scientists were trained and supported to establish the
expression of their target gene in the baculovirus expression
system in our lab. Several in-house HZI cooperation projects
for the analysis of multi-protein complexes have been undertaken
and include: TLRs (Schubert (University of the Western
Cape)/Heinz), HGF-cMet receptor interaction (Krausze/
Heinz/Gherardi), HCV Protease NS3/4 complexed with novel
inhibitory peptides (Collins/Schmelz) and the ABCB6 membrane
protein (Menzel/Heinz). The production of these proteins
is based either on the baculovirus expression system or
the Acembl/Multibac expression system, animal stable glycosylation
deficient CHO cell line cultivation or expression in
Pichia pastoris. Many projects are on-going as the production
of sufficient amounts of soluble and stable protein is still
limiting and requires extensive redesign and recloning of the
expression systems. For several of the projects the process of
crystallisation is still continuing and requires a continuous
supply of fresh and pure material.

128
SCIENTIFIC REPORTS | Technological Platforms
03 Gene Expression Analysis
HEAD | Dr. Robert Geffers | Research Group Genome Analytics | rog@helmholtz-hzi.de
This platform is a central service unit for our internal
investigators as well as for HZI collaboration partners. We
are offering microarray technology for transcriptional and
genomic analysis utilizing the following applications:
Gene expression For gene expression analysis we provide
microarray technology ordered from the market leaders
Affymetrix and Agilent Technologies, respectively. Moreover,
the core unit is furnished with a gene arrayer which
we use to manufacture microarrays in customer desired
format. Most of the microarrays used are so-called “whole
genome” microarrays. More than 40,000 probes (probe
sets) arranged on each microarray are sufficient to cover
the transcriptomes of species like human and mouse. The
straight forward Agilent microarray technology allows for
processing of up to eight samples simultaneously on only
one slide. High flexibility in probe design helps to generate
new microarray formats very cost effective in any complexity
for any organism. The core unit takes responsibility
for the probe design and supervises the steps during the
manufacturing processes. Gene expression analysis can
be performed in different ways depending on the “Target”
that should be measured. Usually 3´ expression arrays
are used. Here the probe design depends on sequences
nearby the 3´end of each transcript. Such microarray can
be used for quantification of the expressed RNA, but failed
to detect splice variants per gene. Exon arrays (Affymetrix)
are composed of probes tiled across the exon sequences of
each gene in the genome. Consequently, these arrays can
be used to quantify and qualify splice variants at a genome
wide level. Also for this approach the core unit takes over
responsibility, if desired, for experimental setup of the
projects and gives advice during the steps of primary data
acquisition.
Tiling arrays Tiling arrays are composed of partially overlapping
probes across defined genomic sequences. These
selected genomic regions either represent regulatory DNA
regions (promoter, enhancer) or DNA regions underlying
epigenetic modifications (CpG isles). Comparative microarray
analysis determines target sequence enrichments based
on chromatin immunoprecipitation (ChIP or MeDIP) of DNA
binding proteins resulting in the identification of either genomic
DNA transcription factor binding sites or epigenetic
modifications within CpG islands. Some tiling arrays are
designed to cover the complete genome (aCGH: array based
comparative genome hybridization analysis). Comparative
genome hybridization, i.e. tumour vs. normal tissue, can determine
genetic instability established during tumourigenesis
of individual tumour entities. The core unit offers besides
catalog aCGH arrays also the opportunity to design focused
versions with higher resolution. Thus, for any complexity of
the project a proper design can be adapted. Moreover, the
core unit will support also the primary data mining process.
In course of internal and external collaborations new analyses
were placed in several areas: tumourigenesis and tumour
typing, pathogen-host interaction of streptococcus, pseudomoniae
and mycobacteria and on immunological issues.
Agilent arrayCGH: Human, Mouse and Rat microarrays can be
offered fort he Agilent SurePrint Systems. Investigators have
the choice between general catalogue arrays or customized
aCGH formats. Left: 244k aCGH; right: 105k aCGH. Graphic: HZI
microRNA expression MicroRNAs are known to play
a pivotal role in gene regulation. Incorporated into the
“RISC”-complex (RNA inducing Silencing Complex) miRNA
molecules mediate the breakdown of mRNA or the reduction
of ribosomal RNA translation. Our core unit offers
continuously updated murine and human microRNA expression
chips based on the newest releases from the Sanger
miRBase. Complete service is provided including microRNA
extraction and array analysis. Finally, a systematically
approach linking changes of microRNA expression with
predicted target gene modification, aims to uncover their
regulative effects applied to the transcriptome.
Genome-wide expression: Human, Mouse and Rat on either
Affymetrix GeneChip or Agilent SuerPrint Systems.Graphic: HZI

SCIENTIFIC REPORTS | Technological Platforms 129
04 Peptide Synthesis
HEAD | Dr. Dr. Werner Tegge | Department of Chemical Biology | wte@helmholtz-hzi.de
SCIENTIFIC COLLABORATOR | Dr. Ronald Frank
Since its inauguration as a service unit in 1990, the platform
generates synthetic peptides, both in soluble form
and immobilised in the form of arrays, for many different
HZI projects and external collaborations. State-of-the-art
equipment is employed for the synthesis, characterisation
and purification. By pursuing own research projects, the
methodological repertoires are continuously updated and
extended.
Developments in this context include:
• New methodologies for the generation of peptide arrays
(e.g. the SPOT method);
• Methods for the preparation of phosphorylated and thiophosphorlylated
peptides
• The utilisation of new biocompatible solid supports
• New selectively cleavable peptide linkers
• Methods for the synthesis of branched peptides
• Methods for the generation of libraries of linear and
cyclic peptides
• Assays for the utilisation of soluble and immobilised
peptides in biological systems
Soluble peptides To date, over 3,000 soluble peptides with
a length of two to over fifty amino acids have been generated
in the platform. Soluble peptides are characterised
using HPLC and mass spectrometry. If necessary, further
characterisation is carried out by amino acid analysis,
protein sequencing, special mass spectrometry techniques
and NMR in the HZI Division of Structural Biology.
Depending on the intended usage and desired quality of
the products, purifications are carried out, usually by preparative
HPLC. For special applications, the platform also
offers peptide modifications like fluorescence labelling,
phosphorylation, biotinylation, lipid conjugation, PEGylation,
branched peptides and cyclisations.
SPOT-arrays In the platform, immobilised peptides in the
form of arrays are generated to facilitate the systematic
and empirical search for ligands. For the successful design
of such arrays a thorough understanding of the biological
problem is essential, which is attained via close co-operation
and collaboration with the users. The SPOT-arrays
are generated semi- and fully automatically on cellulose
membranes or other polymeric supports. Each year, approximately
15,000 peptides and peptide mixtures are
generated in an array format and utilised for the investigation
of e.g. protein-protein interaction, including epitope
mapping and enzyme-substrate recognition.
Method developments for parallel combinatorial chemical
synthesis and screening are based on the SPOT synthesis
performed on cellulose membranes. Photo: HZI, Bierstedt
Tegge, W., Bonafe, C.F.S., Teichmann, A., & Erck, C. (2010) Synthesis of peptides from
α- and β-tubulin containing glutamic acid side chain linked oligo-Glu with defined length.
International Journal of Peptides, Article ID 189396.
Nickl, C.K., Raidas, S.K., Zhao, Sausbier, M., Ruth, P., Tegge, W. Brayden, J.E. & Dostmann,
W.R. (2010) (D)-Amino acid analogues of DT-2 as highly selective and superior inhibitors of
cGMP-dependent protein kinase I-alpha. Biochimica et Biophysica Acta (BBA) – Proteins and
Proteomics 1804, 524-532.
Weiß, S.M., Ladwein, M., Schmidt, D., Ehinger, J., Lommel, S., Städing, K., Beutling, U.,
Disanza, A., Frank, R., Jänsch, L., Scita, G., Gunzer, F., Rottner, K. & Stradal, T.E.B. (2009)
IRSp53 links Tir to EspF U
/N-WASP-mediated actin assembly in EHEC pedestal formation.
Cell Host Microbe 15, 244-258.

130
SCIENTIFIC REPORTS | Technological Platforms
05 Histo-Pathology Platform
HEAD | Prof. Dr. Klaus Schughart | Department of Infection Genetics | kls@helmholtz-hzi.de
SCIENTIFIC COLLABORATOR | Dr. Verena Haist | University of Veterinary Medicine, Hannover
The histo-pathology platform represents a central service
unit at the HZI. It supports several projects and research
groups which require support for histological analyses and
pathology expertise.
Many research projects at the HZI are performing infection
challenge experiments in mice and studying mechanisms
of the host defense in genetically diverse mouse strains
and mutant lines. The histo-pathology unit offers a central
customized service and provides the entire necessary
infrastructure in a single unit. Scientists from the HZI can
either use a complete service – from embedding, sectioning,
staining, archiving of tissues, and review by a pathologist –
or take advantage of its infrastructure and perform some of
the tasks themselves.
At present, paraffin sections and histochemical staining are
offered on a routine basis. Cryostats are available and a set
of immune-histochemical analyses is offered which allows
the detection of immune cells in the mouse during infection
and inflammation. A new vibratom was recently acquired
which allows to cut non-treated samples and therefore
can be used for further experiments, for example in organ
cultures.
Immunohistochemical staining for influenza nuclear protein
(brown) showing infection of cells in the lungs of mice.
Photo: HZI
The unit is operated by Anna Rinkel and it maintains close
collaboration with experts from the Institute of Pathology
at the University of Veterinary Medicine, Hannover, (Dr.
Verena Haist) and the Institute of Veterinary Pathology at
the Free University of Berlin (Prof. Dr. Achim Gruber), who
provide pathological expertise and support for the planning
of experiments and interpretation of results.
Ziehl-Neelsen staining of liver section showing the presence of
acid-resistant bacteria (red). Photo: HZI

SCIENTIFIC REPORTS | Technological Platforms 131
06 Analytical Instruments
HEAD | Dr. Victor Wray | Research Group Biophysical Analysis | vwr@helmholtz-hzi.de
SCIENTIFIC COLLABORATORS | Dr. Heinrich Lünsdorf | Dr. Manfred Nimtz | Priv.-Doz. Dr. Manfred Rohde
The platform provides a facility for determining the three
dimensional structure of all types of natural products and
has instrumentation for mass spectrometry (MS), nuclear
magnetic resonance (NMR) spectroscopy, X-ray crystallography
(X-ray), protein sequencing (PS), electron microscopy
(EM) and confocal laser microscopy.
MS and NMR spectroscopy For the majority of lowmolecular
weight natural products the total structure is
elucidated in a routine manner using a combination of MS
and NMR spectroscopy. The direct analysis of large, intact
biomolecules such as proteins, oligonucleotides and complex
carbohydrates, is routinely carried out using MALDIand
ESI-MS. MS has the important advantage of providing
information for very small amounts of compound. Automated
MS micro-techniques are used for the identification and
characterization of proteins from 2D gels and from “gel-less”
techniques for “Proteomics”, through the determination of
the molecular weight of their proteolytic fragments using
MALDI/TOF-MS/MS and HPLC-ESI-MS/MS. The secondary
and tertiary structure of peptides and proteins can be
elucidated when appropriately labeled material ( 15 N and 13 C)
is available through the application of multidimensional
NMR spectroscopy.
Platform: Analytical Instruments
NMR
X-ray
Small Natural Products
Macromolecules:
Proteins
Molecular Complexes:
Protein-ligand
Protein-Protein
MS
PS
X-ray crystallography The main emphasis in X-ray crystallography
is the structural analysis of proteins. A pipetterobot
and an X-ray unit with an area detector and rotating
anode are available for crystallization and data collection.
The measurement of high resolution data and phase determination
using anomalous dispersion is available through
the use of external synchrotron facilities.
Electron microscopy This technique includes a field
emission scanning electron microscope (FESEM) with an
installed EDX analysis system for elemental analysis and
an attached cryo-stage for cryo-FESEM work. The main
mission is to provide a competitive modern spectrum of
methodology to develop and perform techniques for high
resolution morphological analysis and immune localization
of proteins. High resolution FESEM techniques are especially
used to visualize the adherence to and invasion of host
cells by a wide range of pathogens. Preparation protocols
have been customized to fulfil the wide requests of different
researchers on the campus. New methodologies have
been developed to immune localize pathogenicity factors on
the bacterial cell surface or the interface between bacterial
and host cell membranes using FESEM. Furthermore, goldparticles
coated with isolated proteinaceous pathogenicity
factors have been proven to be a useful tool for studying
the crosstalk between pathogen and host cell. In addition,
conventional and an energy filtering transmission electron
microscopes (TEM, EF-TEM) are used to support findings
by FESEM studies especially for immune localization of
pathogenicity factors inside bacteria or host cells and for
following the intracellular trafficking of pathogens. TEM is
used for studying the quaternary structure of proteins by
negative-staining techniques and is applied for high resolution
elemental localisation studies by electron spectroscopic
imaging and electron energy loss spectroscopy.
EM
Super macromolecules:
Organelles
Bacteria
Platform techniques available for investigation of all types of
natural products Photo: HZI
Wang,Y., Edrada-Ebel,R., Tsevegsuren,N., Sendker,J., Braun,M., Wray,V., Lin,W., & Proksch,P.
(2009) Dihydrostilbene derivatives from the Mongolian medicinal plant Scorzonera radiata.
Journal of Natural Products 72, 671-675.
Xu,J., Kjer,J., Sendker,J., Wray,V., Guan,H., Edrada,R., Lin,W., Wu,J., & Proksch,P. (2009)
Chromones from the endophytic fungus Pestalotiopsis sp. isolated from the chinese mangrove
plant Rhizophora mucronata. Journal of Natural Products 72, 662-665.
Zelena,K., Zorn,H., Nimtz,M., & Berger,R.G. (2009) Heterologous expression of the msp2
gene from Marasmius scorodonius. Archives of Microbiology 191, 397-402.

132
SCIENTIFIC REPORTS | The Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS)
The Helmholtz-Institute
for Pharmceutical Research
Saarland (HIPS)
MANAGING DIRECTOR | Prof. Dr. Rolf Müller | Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS)
within Helmholtz Centre for Infection Research (HZI) |
Campus Saarland University, Building C2 3
, 66123 Saarbrücken | Germany | rom@helmholtz-hzi.de
STAFF | Dr. Markus Ehses | David Hofmann | Birgitta Lelarge | Natja Mellendorf | Claudia Thiele
The Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS) was founded in August 2009 jointly by the Helmholtz Centre
for Infection Research (HZI) and Saarland University (UdS). It is established as branch of HZI on the campus of UdS with the aim
to accelerate the identifi cation of new drug candidates for antiinfective research, their optimization by methods of molecular
biology and medicinal chemistry and the improvement of the transport to the site of action. It is especially the emerging of new
and often multi-resistant pathogens that requires an effi cient approach to the development of antiinfectiva. The founding of HIPS
is the result of an initiative by the Helmholtz Association to complement translational research within the research fi eld “health”.
The combination of the internationally renowned high quality of infection research at HZI and of pharmaceutical sciences at UdS
creates a signifi cant added value. Close cooperations with regional partner institutes allow covering the wide range of drug development
starting from early drug candidate identifi cation to clinical studies all within one research centre. The implementation of
the complete development pipeline will considerably speed up the use of natural products in clinical studies.
The range of scientific activities at HIPS comprises genetic
and genome-analytic methods for optimizing natural product
producers and lead compounds as well as methodologies
to improve the transportation of pharmaceutical agents to
their target. The combination of the expertise in infectious
diseases and in pharmaceutical research at HZI and HIPS
takes a unique position in Germany and Europe, especially
regarding the development of antiinfectives. The three
departments of HIPS, “Microbial Natural Products” (MINS,
Prof. R. Müller), “Drug Design and Development” (DDOP,
Prof. R.W. Hartmann), and “Drug Delivery” (DDEL, Prof.
C.-M. Lehr), originate each from a pharmaceutical research
group at UdS, with approximately 140 employees in total, of
which at the end of 2010 30 were financed by HIPS. Finally,
100 collaborators will move to the new building, which
should be constructed within three to four years on the
campus of UdS.
Another mission of HIPS is the promotion of young scientists.
Currently, a structured doctoral programme is developed,
which besides courses in soft and professional skills
will feature an optional one-year-stay abroad. Furthermore,
three young researcher groups will be established. The
Helmholtz Young Investigators Group “Metabolic Engineering
of Actinomycetes” (AMEG), headed by Andriy Luzhetskyy,
moved to HIPS in January 2011. By the follow-up
appointments to the three vacant university chairs and
further new junior professor positions at UdS, the number
of groups with pharmaceutical focus at Saarbrücken will
raise from currently five full professors and two young researcher
groups to a total of 15 groups in the mid-term. The
The founding heads of departments of HIPS (from left to right):
Claus-Michael Lehr (DDEL), Rolf Müller (MINS) and
Rolf W. Hartmann (DDOP) Photo: HZI
research approaches, methods and apparatus in this pool of
pharmaceutical competence will work together synergistically
under the roof of the intended “Centre for Pharmaceutical
Sciences Saarland”.
The close linking of HIPS to HZI is promoted with high
priority and is being realized by initiation of research collaborations
and implementation of compatible administrative
workflows and infrastructure for controlling, administration
of personnel, IT and public relations. For these
measures an administrative manager, a scientific assistant
as well as an administrative assistant support the director
of the institute.

SCIENTIFIC REPORTS | The Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS)
133
The Department “Microbial Natural Products” (MINS)
Head of Department | Prof. Dr. Rolf Müller | Helmholtz-
Institute for Pharmaceutical Research Saarland (HIPS) within
Helmholtz Centre for Infection Research (HZI) | Department
of Microbial Natural Products (MINS) rom@helmholtz-hzi.de
Scientific Collaborators | Dr. Daniel Krug | Dr. Silke Wenzel
The department “Microbial Natural Products” investigates
the chemistry, biological activity, production and regulation
of natural products mainly from myxobacteria. Myxobacteria
are soil-living organisms and represent a promising
source of natural products, which are now as much valued
as the long-established actinomycetes. Myxobacterial
natural products exhibit diverse biological activities and
modes-of-action and are thus useful for various therapeutical
applications – examples include their use as antibiotics,
anti-cancer drugs, immunosuppressants or anti-parasitics.
Myxobacteria are known to secrete a high number of natural
products, so-called secondary metabolites, at least in
part to presumably inhibit growth of their competitors. In
order to uncover the potential of myxobacteria as producers
of secondary metabolites, scientists at MINS employ
a broad spectrum of techniques including microbiological,
molecular-biological, genetic, biochemical, biotechnological
and analytical methods.
Sonja Burkhard checks a culture in the shaker. Photo: HZI, Bellhäuser
(A) Swarming of the myxobacterium Byssovorax cruenta
starting from a piece of cellulose on agar plate and (B) microscope
image of a fruiting body from Chondromyces crocatus.
Photos: R. Garcia
Newly isolated myxobacterial strains from locations worldwide,
together with the existing myxobacterial strain collection
at HZI (harbouring over 8000 isolates), form the basis
for our interdisciplinary efforts to discover novel natural
products. To date, a number of entirely new myxobacterial
species, genera and families as well as many potentially
novel natural products have been isolated successfully.
Isolation of a novel myxobacterium usually requires careful
optimization of microbiological methods and cultivation
conditions, since these bacteria exhibit a rather complex
“lifestyle”: Myxobacteria are able to glide on surfaces in a
swarm-like pattern and they collectively feed on other microorganisms
by actively degrading their prey. When star vation
conditions are encountered, many cells can aggregate to form
a multicellular structure, termed a “fruiting body”, containing
a high number of heat- and dryness resistant myxospores,
which may survive unfavourable environmental conditions
even for a prolonged period of time.
Once a new myxobacterial strain has been successfully
adapted to growth under laboratory conditions, MINS uses
state-of-the-art mass spectrometric techniques to screen
the strain’s metabolite profile with respect to the presence
of known myxobacterial secondary metabolites. At the same
time the department MINS performs a range of biological
assays, using indicator organisms as well as cell-based
screening approaches to discover novel compounds exhibiting
a potentially interesting activity. Following optimization
of the production of candidate compounds, cultivation of
the producing strain is upscaled. The target compounds are
purified using liquid chromatography methods. Thereafter,
structure elucidation using a panel of analytical techniques
including multidimensional NMR spectroscopy is carried out.
In addition, further cell-based experiments are performed
in order to investigate in detail the mode-of-action of a novel
natural product. This work is performed in close collaboration
with the work group MWIS at HZI.

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Another core expertise of MINS is the investigation of
the biosynthetic pathways underlying the formation of
myxobacterial natural products, including the biochemical
characterization of the involved enzymatic machinery. The
scientists at MINS aim to improve the fundamental understanding
of the mechanisms directing secondary metabolite
biosynthesis in myxobacteria on the molecular level.
Thereafter, we apply this knowledge in order to increase
product yields via the manipulation of regulatory processes
and for the targeted manipulation of biosynthetic pathways,
resulting in the production of structurally modified or novel
compounds. The researchers underpin their efforts with
sequencing of the exceptionally large genomes of a number
of myxobacterial model strains. Having reported the
sequence of the to date largest bacterial genome from
Sorangium cellulosum a couple of years ago, every additional
genome highlights again the enormous genetic
capacity of myxobacteria for the production of secondary
metabolites: the number of predicted biosynthetic pathways
regularly outnumbers the natural products previously
known from a strain. Thus, the bioinformatic analysis of
myxobacterial genomes forms the basis for the discovery of
novel secondary metabolites by using a combined genomemining
and “secondary metabolome mining” approach,
implementing highly sensitive mass spectrometric methods.
Moreover, the sequence data allow the researchers to optimize
the production of myxobacterial metabolites and even
enable the expression of complete biosynthetic pathways
in heterologous hosts, thereby increasing product yield.
Furthermore, it is possible to introduce alterations to a
compound’s structure by genetic engineering. Last but not
least, the availability of whole-genome information is an
important prerequisite to deepen our understanding of the
host microbiology of myxobacteria.
The treatment of infectious diseases is of growing significance
in health care as an increasing number of pathogens
become multi-resistant. Thus, the development of new
antibiotic drugs has become one major topic for pharmaceutical
research, in particular because efficient medications
fighting against infectious diseases are still missing. Conventional
antibiotics target essential processes in bacteria
to kill these pathogens. An alternative strategy aims at
blocking the cell-cell communication by inter ference of the
drug with the communication system. This communication
system is based on the exchange of signal molecules and
regulates the production of the virulence factor production
and biofilm formation, which could thus both be reduced or
even prevented.
Recent advances in microorganism research have resulted
in the identification of an increasing number of natural
compounds showing antibiotic activity. However, most of
these structures are not suitable for becoming drugs due
to unfavourable pharmacokinetic properties, such as poor
solubility or difficult large-scale synthesis. The department
DDOP is focusing on the development of novel synthetic
antibiotics, which can either be derivatives of the natural
hit compounds with improved drug likeness, or synthetic
compounds based on a rational drug design. Different medicinal
chemistry strategies are applied, ranging from simplification
of the molecular complexity of the compounds to
computer-aided methods in drug design.
At DDOP two main projects are currently developed, targeting
either the bacterial growth or their cell-cell communication
system.
The Department “Drug Design and Optimisation” (DDOP)
Head of Department | Prof. Dr. Rolf W. Hartmann | Helmholtz-Institute
for Pharmaceutical Research Saarland (HIPS)
within Helmholtz Centre for Infection Research (HZI) |
Department of Drug Design and Development (DDOP) |
rwh09@helmholtz-hzi.de
Scientific Collaborators | Dr. Johannes De Jong | Dr. Jörg
Haupenthal | Dr. Matthias Negri | Dr. Anke Steinbach |
Dr. Christiane Zimmer
Jeannine Jung and Cenbin Lu from the department DDOP
discuss the synthesis of a new inhibitor. Photo: HZI, Bellhäuser

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Development of potent bacterial RNA polymerase
inhibitors The project focuses on the development of
inhi bitors that block the transcription of bacterial genes.
These molecules are intended to target the bacterial RNA
polymerase (RNAP). The novel RNAP inhibitors should
also be effective against bacteria that are resistant to other
antiinfectives.
RNAP is the enzyme responsible for the formation of RNA
in all living cells. The bacterial form is a complex multisubunit
enzyme with a molecular weight of about 400 kDa.
It consists of four different subunits, which form the core
enzyme (α2ββ‘ω). The latter is activated for the transcription
by interaction with the so-called sigma factor. As RNAP
is essential for bacterial growth and selective to bacteria, it
is a highly attractive target for the development of a broadspectrum
antibiotic against severe diseases with humans.
One important example is Mycobacterium tuberculosis (Mtb),
the pathogen of tuberculosis, which causes two million
deaths per year worldwide. However, RNAP inhibitors that
are in clinical use have evoked resistant strains, which
increase the demand for new drugs enormously. The aim
of the project is to develop a new inhibitor of the bacterial
RNAP, to characterise its mode-of-action and to optimise
the hits. By this approach, the inhibitor becomes a highly
potent drug that invades pathogenic bacteria to finally
unfold its antibiotic function in humans.
In the development of novel RNAP inhibitors DDOP follows
two strategies:
The first strategy consists of direct inhibition of the core enzyme.
Different binding sites have been described for RNAP,
which are prone to inhibitors that can act either by directly
blocking the active site or by influencing the flexibility of
RNAP via allosteric mechanisms. Since the 3D structure of
RNAP is known, computer-aided drug design strategies are
used to screen virtual compound databases and to rationally
select compounds for subsequent experimental screening.
Virtual hits showing effectiveness in vitro will then pass
over to the hit-to-lead phase. First validated screening hits
have already been identified and novel synthetic projects
have been launched to chemically optimize the hits.
The second strategy aims at preventing the holoenzyme
formation, i.e. the assembly of the core enzyme with the
sigma factor. Part of the interaction interface of the two
subunits has been characterized by means of mutagenesis
studies, and it has been shown that this interface might be
a suitable target for enzyme inhibition. This information
Structure of RNA polymerase exhibiting five subunits. Ligandand
structure- based approaches are followed in the design of
new RNAP inhibitors.
is used by DDOP to identify a suitable regions for binding
and hence to develop an appropriate inhibitor. First RNAP
inhibitors, in part short peptides, have already been identified.
The aim of the project is to optimize the inhibition
effect and various pharmacokinetic properties of these
compounds. Finally, this should lead to a highly potent drug
for use in therapeutic applications in humans.
RNAP from both E. coli and Mycobacterium tuberculosis will
be cloned and used for the biological testing of the synthesized
compounds. The characterization of the putative
binding site(s) of novel potent inhibitors will be performed
by site-directed mutagenesis.
Development of compounds interfering with PQS quorum
sensing communication in Pseudomonas aeruginosa In
this project, the scientists at DDOP develop compounds
that interfere with the PQS quorum sensing communication
system in Pseudomonas aeruginosa. Lung infections by
this opportunistic pathogen are considered a major cause
of death for patients with cystic fibrosis. Resistance to
conventional antibiotics is often associated with the formation
of biofilms, which prevent the penetration of antibiotics
through this physical barrier and thus reduce the immune

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response. Both the formation of biofilms and the production
of virulence factors are controlled by a complex quorum
sensing communication system. By this communication
system, bacteria detect their population density and control
a kind of population behaviour, amongst others. Besides the
two quorum sensing systems, LasR and RHLR that exist in
many bacterial species, in P. aeruginosa, a third, unique
communication system is regulated by PQS (Pseudomonas
Quinoline Signal, 2-heptyl-3-hydroxy-4-quinolone) as
signal molecule. The aim of the project is the development
of compounds that interfere with the PQS quorum sensing
communication system in order to prevent the formation of
biofilms and the production of different virulence factors –
this, however, without affecting the growth of bacteria. Two
approaches are followed in this project as well: On the one
hand the effect of PQS on the receptor PqsR is antagonized
and on the second hand, the biosynthesis of HHQ, the direct
precursor of PQS, should be blocked by inhibition of the
enzyme PqsD.
Development of antagonists of the transcription regulator
PqsR Initial drug discovery efforts are focused on the
synthesis of close analogues of PQS intended to bind to
and block PqsR. The transcription regulator PqsR drives
the expression of virulence determinants after activation
by PQS. The PQS analogues will be evaluated in a recently
established β-galactosidase reporter gene assay. After
activation of the PqsR transcriptional regulator by PQS, the
transcriptional activation of the PqsA promoter should be
detected. Furthermore, the SPR methodology for the kinetic
characterization of hits will be developed with the soluble
truncated version of PqsR, ( C87 PqsR) and applied for medium
throughput screening of fragment libraries. Hits from the
SPR screening are further thermodynamically characterized
using ITC (isothermal titration calorimetry).
Inhibition of the second enzyme of the PQS biosynthesis
PqsD For the biochemical evaluation of potential PqsD
inhibitors, a medium-throughput PqsD LC-MS/MS based
inhibition assay has been established in our lab. Novel PqsD
inhibitors, structural analogues of the native substrate
anthraniloyl-CoA, have been synthesized. Currently, an
SPR-based medium-throughput screening of the in-house
library and of commercially available fragment-libraries is
performed followed by the kinetic characterization of hits.
The same libraries are also virtually screened on PqsD with
the aim to identify the putative binding mode of the fragment
hits. Initial active fragments are identified and used as
novel scaffolds in ongoing synthesis projects. The identification
of fragments and their binding mode will be further explored
by high resolution NMR-spectroscopy. Further, kinetic
studies, both computational and biochemical (biological assays,
SPR, ITC), should promote the development of PqsD-inhibitors.
Especially the evaluation of the kinetic mechanisms
by molecular dynamic simulations will provide information
about the flexibility of the protein and may contribute to the
elucidation of possible induced-fit mechanisms.
The Department “Drug Delivery” (DDEL)
Head of Department | Prof. Dr. Claus-Michael Lehr |
Helmholtz-Institute for Pharmaceutical Research Saarland
(HIPS) within Helmholtz Centre for Infection Research
(HZI) | Department of Drug Delivery (DDEL) | cle09@
helmholtz-hzi.de
Scientific Collaborators | Dr. Eva-Maria Collnot |
Dr. Nicole Daum | Dr. Steffi Hansen | Dr. Brigitta Loretz
Advances in molecular biotechnology and medicinal chemistry
have led to the discovery of new drug candidates with
potential affinity to target receptors. However, developing
such molecules into drug products necessitates first screening
of their per meability, transport and bioavailability
through biological barriers and second establishing new
technologies to ensure their safe and effective delivery to
the site of action. Therefore, the main focus of research in
the department Drug Delivery is on the one hand exploring
the biological barriers, in particular the lungs, the gastrointestinal
tract, and the skin. On the other hand, DDEL
develops the appropriate carriers capable of crossing these
epithelial barriers and delivering the active molecule to the
target.
Nico Mell from the department DDEL attaches flasks with
samples at the lyophilisator for freeze drying. Photo: HZI, Bellhäuser

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Development of a new in vitro model of the air-bloodbarrier
The so-called “air-blood-barrier” represented by the
alveolar epithelium of the lung is crucial for the pulmonary
delivery of drugs and plays an important role in the context
of aerosol transmitted infectious diseases (e.g. swine flu,
tuberculosis). To mimic the in vivo situation several in vitro
cell models are available (e.g., tumour cells, primary cells
from animal origin), most of them lacking biological relevance
due to impaired barrier properties. Primary human
cells reflect the in vivo situation best, but are not available
in sufficient amounts and the isolation procedure is time
and cost intensive.
Hence, a new cell line is being developed to establish a nontumour
derived in vitro model with intact barrier properties.
This approach will help to elucidate infection pathways
across the respiratory tract and to facilitate the discovery of
drug compounds capable of treating these diseases.
Needle-free transdermal vaccination Traditional vaccination
requires intramuscular injection. However, this kind
of vaccination does not ensure the optimal delivery of the
vaccine to the skin antigen presenting cells that elicit the
immune response. Current strategies of needle-free vaccination
via the dermal route reduce the protective stratum
corneum barrier for a significant time. This makes them
suboptimal for certain applications, such as mass vaccination
campaigns in countries with critical hygienic conditions.
Recently, an alternative pathway for targeting the skin
immune system via the hair follicles by using nanoparticles
has been reported. As it is well known that pollen-antigens
rapidly cross the sebum filled follicle and cause an allergic
reaction with susceptible persons, the scientists at DDEL
hypothesize that mimicking the “delivery strategy” of pollen
with artificial particulate carriers is a promising strategy
for non-invasive vaccination across the intact skin barrier.
Drug loaded nano- and microparticles for the treatment
of inflammatory bowel diseases Inflammatory bowel
diseases such as ulcerative colitis are autoimmune diseases
of the intestine. Patients suffer from recurring acute pain,
gastric and intestinal spasms, and heavy, often bloody diarrhea.
In addition, the risk of developing intestinal cancer is
substantially increased.
It could be shown in animal studies that nano- to microsized
particles accumulate specifically in inflamed areas of the
intestine. Therefore, nano- and microparticle based medicines
may help to improve the therapy of inflammatory bowel
Caco-2 Zelle with propylstarch-nanopartikels adhered at the
cell membrane Photo: HIPS / HZI
diseases in the near future. Currently, such new formulations
are being developed for a number of anti-inflammatory drugs
at DDEL. To better understand the mechanism of particle accumulation
in the inflamed tissue a novel cell culture model
of the inflamed intestinal mucosa has been established.
Design of new gene delivery carriers In the field of nonviral
genetic therapies the current focus shifted from the
cure of genetic disorders more towards vaccination or allergy
treatment. Additional new interest arose with the discovery
of RNA interference. It concerns a mechanism for sequencespecific
inhibition of target genes, which can be used as
therapy for the treatment of a variety of diseases. There
are physicochemical and biological barriers, which hinder
the delivery of free oligo- or poly-nucleotides. Their use is,
therefore, reliant on suitable systems to achieve efficient,
safe and clinically appropriate delivery to overcome biological
barriers. The scientists at DDEL develop new formulations
from current approaches in polymer chemistry and
nanotechnology. In these formulations, the drug is either
bound coordinatively (polyrotaxanes as cationic partner
complexing anionic nucleic acid polymers) or encapsulated
in nanoparticles (core-shell carrier particles on the basis
of biodegradable polymers like chitosan-coated PLGA particles).
The aim is the development of carrier systems that
can be adapted to various application routes by adjustable
physical and pharmacokinetic properties. Such transport
systems are optimized for the intake via the lung or, in a
future project, via the intestinal tract.

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TWINCORE, Centre for
Experimental and Clinical
Infection Research GmbH
EXECUTIVE DIRECTOR | Prof. Dr. Ulrich Kalinke
HEAD OF EXPERIMENTAL INFECTION RESEARCH | Prof. Dr. Ulrich Kalinke
HEAD OF EXPERIMENTAL VIROLOGY | Prof. Dr. Thomas Pietschmann
HEAD OF INFECTION IMMUNOLOGY | Prof. Dr. Tim Sparwasser
HEAD OF PATHOPHYSIOLOGY OF BACTERIAL BIOFILMS | Prof. Dr. Susanne Häußler
HEAD OF CELL AND GENE THERAPY | Prof. Dr. Michael Ott
TWINCORE, the translation centre of HZI and MHH TWINCORE, Centre for Experimental and Clinical Infection Research GmbH,
is a joint venture between the Helmholtz Centre for Infection Research (HZI), Braunschweig, and the Hannover Medical School
(MHH). The goal pursued in the founding of TWINCORE is to promote and further develop the outstanding expertise of HZI and
MHH in the fi eld of infection research in a joint centre with a specifi c focus on translational research. The purpose of translational
research is a dual one, on one hand facilitating the path of the latest fi ndings from basic research to the patients and, on the
other hand, enabling unanswered questions from clinical practice to make their way back to researchers. A key part of the work
at TWINCORE is also the scientifi c investigation of regulatory concerns regarding approval and implementation of clinical trials.
Complex issues frequently arise in advance of clinical trials, for example regarding the relevance of preclinical experiments for the
safety and effi ciency of new medicines. TWINCORE helps to enable the development of new treatment options for the prophylaxis
and therapy of infectious diseases ensuring that a solid scientifi c basis for the minimisation of risk exists prior to the testing of
new approaches on humans. Research at TWINCORE focuses on the analysis of pathogen-host interactions. New fi ndings in this
area lead to new approaches regarding mechanisms of pathogen inhibition and to the development of new vaccination strategies.
TWINCORE holds laboratories which are adapted to conduct experiments up to bio safety level S3**. During the 2 nd TWINCORE
Symposium entitled “Antimicrobials and Vaccines”, which was held on August 12, 2010, the freshly renovated animal house was
inaugurated (see Figure 1). The animal house can accommodate more than 2,000 mouse cages and is also adequate for performing
experiments up to bio safety level S3**. In addition to the annual TWINCORE Symposium a varied programme of lectures has
been established, in which translational and basic researchers report on their recent research results.
1. Analysis of pathogen-host interactions During long
periods of co-evolution, pathogens and hosts developed
complex strategies to enable survival of both the host population
and the pathogen population. At the cellular level
intrinsic immune mechanisms play a role. At TWINCORE
investigations are conducted to establish the influence such
factors have on host- and tissue-specificity of pathogens. For
several years it has been acknowledged that, in addition to
the recognition of “foreign”, the communication of “danger
signals” via pattern recognition receptors (PRR) plays a
central role in the induction of protective immunity. The
analysis of how innate immunity is triggered by stimulation
of PRR and the consequences on pathogen-specific immunity
are the subject of intensive investigations. In this process
both acute and chronic courses of infections and their
associated inflammatory reactions are examined. Pathogens
have developed various strategies to evade host immunity.
Pathogen-encoded factors that modulate immune responses
are sought. Furthermore, the influence of regulatory cells
on the course of infection is also analysed.
2. New mechanisms of pathogen inhibition Following the
triumphant march of antibiotics in the treatment of bacterial
infections, over the past decades key breakthroughs have
been made in the development of antiviral substances. New
approaches to the inhibition of pathogen reproduction are
sought at TWINCORE. In collaboration with the HZI and the
University of Hannover, biological compound libraries are
being examined for antiviral and antibacterial substan ces.
This involves the utilisation of new cell culture methods, for
example permitting a targeted search for inhibitors of HCV
replication. A further core focus is the search for inhibitors

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Fig. 1. As part of the 2 nd TWINCORE Symposium “Antimicrobials and Vaccines” the newly renovated animal house was inaugurated on
August 12, 2010 at TWINCORE (left picture). The TWINCORE main entrance with the symposium transparent (right image). The “red
tape” was jointly cut through at the opening of the new animal house by Prof. Bitter-Suermann (MHH), Prof. Wehland (HZI),
Mr. Gevers (MWK) and Prof. Kalinke (TWINCORE) (from left to right). Photos: HZI/Twincore
of bacterial biofilm formation which occur in chronic infections.
Similarly, new gene-therapy approaches are also being
sought for the treatment of infectious diseases. Moreover, it
is also being investigated whether pathogen-encoded immune
modulators constitute potential target structures for
new therapeutic approaches.
3. New vaccination strategies To the general public, vaccination
is one of the most successful medical advances. Never -
theless, there are still numerous infectious diseases for
which no vaccine is available. Consequently, at TWINCORE
new vaccination strategies are being developed. Virus-like
particles are investigated for utilisation as vaccine vectors,
as well as the specific in vivo charging of specialised crosspresenting
dendritic cells with antigens. One interesting
option is the reinforcement of immune responses by influencing
regulatory T cells. There are currently comparatively
few approved adjuvants for the enhancement of immune
responses following vaccination. Therefore, new adjuvants
are being investigated in collaboration with partners at HZI.
Similarly, investigations are also underway to determine
whether cytokines are suitable natural adjuvants for certain
vaccination protocols. A further key focus is the analysis
of mechanisms that support the induction of long lasting
IgG responses.
4. New preclinical models New therapeutic or prophylactic
approaches developed in basic research need to be subjected
to extensive preclinical testing prior to first being tested in
humans. At TWINCORE new models are being developed
that should enable improved prediction with regard to reactions
in humans. An important point in this respect is the
humanisation of mice. To this end, on the one hand, mice
are treated with human cells to enable components of the
human immune system to develop in the animals. On the
other hand, mouse models are also being developed in which
human precursor cells contribute to the formation of liver
tissue. A further aspect is the genetic humanisation of mice,
for example by bacterial artificial chromosome (BAC)-mediated
transgenesis. Using this method, human receptors and
allelic forms of them found in the human population may be
expressed in order to investigate their function in an animal
model. Another subject is the investigation of effects that
are mediated by constant portions of human antibodies. This
subject is particularly relevant in the context of the development
of new therapeutic monoclonal antibodies.
A view of the TWINCORE building, Centre for Experimental and
Clinical Infection Research GmbH. Alongside modern laboratories
and superb equipment, TWINCORE also offers optimal rooms
in which to hold scientific meetings and seminars. Photo: HZI/Twincore

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Research groups at TWINCORE
TWINCORE was officially inaugurated in 2008 and all key
leading positions were occupied in 2009. Currently, Prof. Dr.
Ulrich Kalinke is working as Executive Director of TWIN-
CORE and Director of the Institute for Experimental Infection
Research, Prof. Dr. Thomas Pietschmann as Head of the
Department for Experimental Virology, Prof. Dr. Tim Sparwasser
as Director of the Institute for Infection Immunology,
and Prof. Dr. Susanne Häußler as Head of the Department
for Pathophysiology of Bacterial Biofilms. Furthermore, the
Translational Research Group for Cell and Gene Therapy
headed by Prof. Dr. Michael Ott of the Clinic for Gastroenterology,
Hepatology and Endocrinology of the MHH, directed
by Prof. Manns, has been delegated to TWINCORE. A total
of 98 employees are working currently at TWINCORE.
The TWINCORE research group leaders (from left to right):
Profs. Susanne Häußler, Michael Ott, Thomas Pietschmann,
Executive Director Ulrich Kalinke, and Tim Sparwasser.
Photo: HZI, Gramann
Research group Prof. Dr. Ulrich Kalinke |
ulrich.kalinke@twincore.de Following viral infection,
within hours type I interferon responses are usually
induced which secure the initial survival of the host. It is
only about a week later that adaptive immunity is activated
to an extent that it is able to eradicate pathogens. In earlier
projects we have shown that following an infection with the
vesicular stomatitis virus (VSV) a small number of highly
specialised cells, also addressed as plasmacytoid dendritic
cells (pDC), are activated via PRR triggering to produce
large quantities of protective type I interferon. Interestingly,
practically all viruses examined more closely developed
countermeasures that inhibit the induction of such type I
interferon responses. A key focus of our work is how different
viruses induce type I interferon responses and which
type I interferon-inhibiting factors they encode. Local conditions
of type I interferon responses decisively influence
the course of disease. In that line we are investigating how
type I interferon inhibits the spread of pathogens within
the central nervous system. In more recent investigations,
we discovered that type I interferon can also have direct
effects on immune cell functions. We are currently studying
how type I interferon induced upon vaccination with
virus-like particles influences the formation of long lasting
IgG antibody responses. Antibodies are comparatively
large molecules, the variable parts of which bind antigens
specifically, whereas the constant parts may be bound via
so-called Fc receptors expressed by certain immune cells
and other body cells and confer antibody function. We
investigate how Fc receptors interact with different subclasses
of IgG antibodies and which immune functions are
influenced this way. This issue is particularly relevant for
therapeutically-employed monoclonal antibodies because
such reagents play an increasingly important role as innovative
therapeutic agents in the treatment of tumours,
Theresa Frenz working at the clean bench Photo: Twincore/HZI
Claudia Soldner analysing the results of an experiment
Photo: Twincore/HZI

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The workgroup of Prof. Dr. Ulrich Kalinke: (from left to right)
Ulrich Kalinke, Robert Bittorf (last row), Linda Semmler,
Theresa Frenz, Marius Döring (3rd row), Claudia Soldner,
Anette Kessler, Christian Mers, Jennifer Paijo, Claudia Detje,
Lukas Graalmann, Patrick Bartholomäus (2nd row),
Sabrina Heindorf, Stephanie Vogel, Julia Heinrich (1st row).
Photo: Twincore/HZI
are involved in the development of a new immuno therapy
approach for the treatment of chronic HCV infection and
HCV-associated hepatocellular carcinoma. A new focus is
forming our investigations of the HCV species and tissue
tropism. HCV replicates in the liver and only infects
humans and chimpanzees. The mechanisms which are
responsible for this restricted species and tissue tropism
have been scarcely studied up to now and thus are only
barely understood. With our research on this topic we aim
to understand how extra hepatic virus reservoirs can affect
virus pathogenesis and the response to therapy, how HCV
makes use of essential cellular cofactors, and which host
factors control the HCV replication.
autoimmune diseases and infections. In order to make even
better use of the results of the above mentioned approaches
for the application and implementation of clinical trials,
regulatory research is performed in collaboration with the
Paul-Ehrlich-Institut, Germany.
Research group Prof. Dr. Thomas Pietschmann |
thomas.pietschmann@twincore.de Infection with HCV
which belongs to the family of flaviviruses is one of the
major causes of chronic liver disease. According to WHO
estimates, up to 170 million people have had HCV contact
worldwide. Of these, around 100 to 130 million people
are considered to be chronically infected. We develop
new cell culture techniques for the investigation of HCV
replication. The goal of this research is to investigate the
molecular mechanisms of HCV replication in liver cells. In
particular, early stages of HCV infection are studied which
are critically involved in virus entry into liver cells. We
are also analysing processes that lead to the packing of the
viral genetic material in progeny viruses and their release
from the host cell. In this manner we aim to draw up the
fundamental basis of the infection strategy of this humanpathogenic
virus in order to subsequently generate new approaches
and perspectives for the development of therapies.
In TWINNING projects with partners at the MHH and the
HZI we employ the HCV cell culture system to identify new
agents that inhibit HCV replication. Furthermore, within
the Helmholtz Alliance on Immunotherapy of Cancer we
The workgroup of Prof. Dr. Pietschmann: (from left to right)
Kathrin Hüging, Nikoleta Bodosoglou, Juliane Dörrbecker,
Dorothea Bankwitz, Christina Grethe, Juliane Gentzsch, Sibylle
Haid, Nicolas Menzel, Patrick Chhatwal, Eike Steinmann, Thomas
Pietschmann, Gabrielle Vieyres, Stephanie Pfänder, Anne
Frentzen (missing: Joachim Hain, Martina Friesland, Sandra
Ciesek) Photo: Twincore/HZI
Research group Prof. Dr. Tim Sparwasser |
tim.sparwasser@twincore.de Our focus lies upon the investigation
of the significance of PRR, for example from the
family of Toll-like receptors (TLRs) and the C-type lectins,
for the activation of the most important positive regulators
of the immune system and initiators of adaptive immunity,
i.e. the dendritic cell system (DCs). A further focus is upon
regulatory T cells (Tregs) which may be regarded as the
principal counterparts to DCs: Tregs use mechanisms as
yet not completely understood to inhibit an overshooting
immune response and limit the proliferation of T effector
cells. Optimal vaccination strategies against pathogens
may comprise the activation of specific DC subpopulations
whilst avoiding Treg expansion or induction. Vaccination
studies aimed towards Tregs and DCs in the murine model

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Matthias Lochner is currently establishing a junior research
team on the analysis of the role of DCs in the induction and
control of inflammatory (Th17) and regulatory T-cell populations
in infection and inflammation reaction of the intestine.
Jointly analysing scatter plots (from left to right): Nicole Fisch,
Venkateswaran Ganesh, Wiebke Ginter, Catharina Schrauf,
Christian Mayer und Ina Lu Photo: Twincore/HZI
system have several limitations, one of which is that in vivo
analysis of Tregs and DC subpopulations is extremely difficult.
For example, subpopulations of DCs exist in extremely
low numbers in various lymphatic organs that have highly
specialised tasks in several cases including the induction
of tolerance. As these “regulatory immune cells” usually
react highly sensitively to ex vivo isolation and have thus far
proved only inadequately manipulable in vivo, the knowledge
of function and significance of Tregs and DC subpopulations
for adaptive immunity remains incomplete. A further complicating
factor is the expression of different pattern recognition
molecules or different expression profiles of PRRs to DC sub -
populations between humans and mice. A key objective is,
therefore, the development of molecular tools that allow the
genetic manipulation of DCs and Tregs. We wish to use these
models to investigate the function of Tregs and subpopulations
of DCs in infection, allergy and tolerance. In “humanised”
models the role of PRRs such as human TLR9 and DC-SIGN
is analysed and vaccination strategies aimed at these molecules
tested in vivo. Within the Sparwasser Institute Dr.
Research group Prof. Dr. Susanne Häußler |
susanne.haeussler@twincore.de The successes of modern
medicine are increasingly affected by opportunistic bacterial
infections. In chronic bacterial infections the pathogens may
often come together within so-called biofilms and are then
better protected against attack by cells of the immune system
or antibiotics. The life within the population of the bacteria
also provides additional adaptation mechanisms to stress
situations that go beyond the usual reactions of individual
cells. Pseudomonas aeruginosa is the most dominant bacterial
pathogen causing chronic lung infection in cystic fibrosis
(CF) patients. Although most patients are colonised with only
one P. aeruginosa clone, various morphotypes can be isolated.
This diversity seems to play an important role in the persistence
of the germ and thus in the formation of a chronic infection.
Our research focuses on the elucidation of the molecular
mechanism underlying this diversity. In CF patients with a
chronic P. aeruginosa infection of the lungs so-called small
colony variants (SCVs) are frequently found which form
biofilms particularly efficiently. To identify the mutations
that lead to the formation of such SCVs, we sequenced the
genomes of P. aeruginosa clinical isolates using the so-called
next generation sequencing technology. The comparative
analysis of chromosomal DNA sequence of P. aeruginosa
isolates with similar phenotypic characteristics; we look for
typical base substitutions and check whether they are causally
involved in the progression of the phenotype.
In the future we aim to study clinically relevant mutations
that occur in P. aeruginosa under in vitro biofilm growth
conditions and in vivo in the course of a chronic infection.
The knowledge of the genotypes that are selected at different
stages of infection can be helpful for the development of
new, promising therapy strategies. In addition, to two
The workgroup of Prof. Dr. Sparwasser: (from left to right) Christina Hesse, Christian Klemann, Amrita Nandan, Matthias Lochner,
Stefanie Pohl, Christian Mayer, Zuobai Wang, Franz Puttur, Catharina Schrauf, Luciana Berod, Christopher van Helt, Julia Huntenburg,
Wiebke Ginter, Nicole Fisch, Martina Thiele, Stephanie Dippel, Christine Jänke, Janika Quindt, Esther Ermeling, Abdul Mannan Baru,
Venkateswaran Ganesh, Siona Hauer, Tim Sparwasser. Photo: Twincore/HZI

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well-characterized homoserine lactone signal molecules P.
aeruginosa produces a third interbacterial signal molecule,
the Pseudomonas quinolone signal (PQS). PQS is involved
in cell density dependent virulence factor regulation – as
well as the homoserine lactone – and is essentially involved
in the establishment of P. aeruginosa biofilms. However, the
molecular mechanism of the implementation of the PQS signal
in a bacterial behaviour at the signal cell level is largely
unknown. Here, a pqsE encoded enzyme, the last gene in
the PQS biosynthetic operon, seems to play a central role.
The elucidation of the function of PqsE is a key research
focus of our group.
The workgroup of Prof. Dr. Ott: (from left to right) Johan Waern,
Michael Rothe, Martin Pacher, Michael Ott, Urda Rüdrich,
Gesa Riedel, Quinggong Yuan, Ina Rittelmeyer (missing:
Michael Bock). Photo: Twincore/HZI
Prof. Häußler and her team: (from left to right) Vera Nöding,
Kathi Klimmek, Susanne Häußler, Mathias Müsken, Andrea
Blanka Photo: Twincore/HZI
Research group Prof. Dr. Michael Ott |
ott.michael@mh-hannover.de We develop cell and gene
therapy procedures for the treatment of hereditary liver
disease. Another focus lies on the development of mouse
models with chimeric human/murine liver tissue and human
immune system for researching vaccination strategies
against HIV and HCV. The repopulation of the liver with
human liver cells and the transplantation of human blood
stem cells into immune deficient mice continue to represent
a major scientific challenge. In order to investigate vaccination
strategies in the alb-uPA transgene immune deficient
(RAGyc) mouse it is necessary that human cells of a donor
are utilised in the repopulation of the mice. Where primary
tissue is used as starting material the isolation of both cell
populations using foetal liver tissue exclusively is possible.
In our transplantation experiments it emerged that the transplantation
of foetal hepatoblasts is significantly less efficient
than that of adult primary hepatocytes. Alternatively, the
research group enabled the transplantation of foetal human
liver tissue beneath the capsule of the recipient liver to be
tested for the first time. The first combined transplantations
of human blood stem cells and foetal liver tissue to a newlydeveloped
mouse strain are set to be performed shortly. Due
to the lack of availability of primary human cell material for
the “humanisation” of mice, as well as for cellular therapies
in humans, the research group is conducting intensive research
into alternative cell sources. In association with MHH
groups and international partners, research is underway into
hepatic differentiation protocols for embryonic stem cells
and iPS cells. In another project the risk of insertional mutagenesis
in lentiviral gene transfer is being analysed. Serial
transplantation of ex vivo genetically transduced hepatocytes
enables the incidence of liver tumours in dependence on the
number of lentiviral insertions to be investigated. Furthermore,
the research group is supervising a clinical study on
cell transplantation in patients with urea cycle disorder.
This project marks the world’s first controlled study on cell
therapy of hereditary metabolic diseases of the liver.
TWINCORE key publications In 2010 employees working at
TWINCORE published a total of 54 articles. In 2011 already
24 articles have either been published or are “in press” (see
“Complete list of articles published in 2010 as well as articles
published an “in press” in 2011 by employees working
at TWINCORE”). Particularly important research results
have been compiled in the following areas:
1. Regulatory T cells in the immune modulation therapy
So far, proven therapeutic vaccination against tumours
have shown only minimal or no success. Now we have succeeded
in improving the therapeutic vaccination success
in the treatment of malignant melanoma in a mouse model
by selective depletion of regulatory T cells (Klages et al.,
2010). Another project made an important contribution to a
better understanding of the regulatory mechanisms of the
immune system in the intestine (Sawa et al., 2011).

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SCIENTIFIC REPORTS | TWINCORE, Centre for Experimental and Clinical Infection Research GmbH
2. Mechanisms of hepatitis C virus replication and
immune control In a hepatitis C virus (HCV) infection
species-specific interactions with surface receptors such
as CD81 play a central role. According to this, only chimpanzees
and humans are naturally infected with HCV.
After adaptation of HCV to a mouse CD81, it was possible
to identify three mutations in the envelope glycoprotein of
HCV which enhance the infection of cells with murine CD81
100-fold and thereby allow the HC cell entry into mouse
cells without using human cofactors (Bitzegeio et al., 2010).
These studies raise the hope that it will one day be possible
to establish a permissive immuno-competent small animal
model for HCV infection.
3. Pathogen recognition and immune stimulation by dendritic
cells Virus-induced type I interferon (IFN) responses
may enhance cytotoxic T cell responses dramatically. This
finding is surprising because IFN-responses reach peak
serum levels within hours of infection, while cytotoxic T
cell responses are maximally induced after approximately
one week. The work by Frenz et al. has shown that IFNresponses
enhance T cell proliferation by the stimulation
of antigen-presenting cells as well as through the direct
stimulation of T cells (Frenz et al., 2010).
4. Immune subversion strategies In the context of chronic
bacterial infections biofilms may develop which cannot be
treated with normal antibiotics. We have succeeded in developing
a new in vitro test system that allows the targeted
search of novel compounds that inhibit biofilm formation
(Pommerenke et al., 2010).
5. Humanised mouse models Treatment options for various
liver diseases are limited by the fact that syngeneic
hepatocytes are not usually available. In the mouse system
we have succeeded in developing a new process that allows
the differentiation of induced pluripotent stem (iPS) in
hepatocyte-like cells (Sancho-Bru et al., 2010).
• Bitzegeio J, Bankwitz D, Hueging K, Haid S, Brohm C, Zeisel MB,
Herrmann E, Iken M, Ott M & Baumert TF, Pietschmann T (2010)
Adaptation of hepatitis C virus to mouse CD81 permits infection of
mouse cells in the absence of human entry factors. PLoS Pathogens
6: e1000978.
• Frenz T, Waibler Z, Hofmann J, Hamdorf M, Lantermann M, Reizis
B, Tovey MG, Aichele P, Sutter G & Kalinke U (2010) Concomitant
IFNAR-triggering of T cells and of DC is required to promote maximal
MVA-induced T-Cell expansion. European Journal of Immunology
40(10), 2769-2777.
• Klages K, Mayer CT, Lahl K, Loddenkemper C, Teng MW, Ngiow SF,
Smyth MJ, Hamann A, Huehn J & Sparwasser T (2010) Selective
depletion of Foxp3+ regulatory T cells improves effective therapeutic
vaccination against established melanoma. Cancer Research 70(20),
7788-7799.
• Pommerenke C, Müsken M, Becker T, Dotsch A, Klawonn F & Häussler
S (2010) Global genotype-phenotype correlations in Pseudomonas
aeruginosa. PLoS Pathogens 6(8). pii: e1001074.
• Sancho-Bru P, Roelandt P, Narain N, Pauwelyn K, Notelaers T,
Shimizu T, Ott M & Verfaillie C (2010) Directed differentiation of
murine-induced pluripotent stem cells to functional hepatocyte-like
cells. Journal of Hepatology 54(1), 98-107.
• Sawa S, Cherrier M, Lochner M, Satoh-Takayama N, Fehling HJ,
Langa F, Di Santo JP & Eberl G (2010) Lineage relationship analysis
of ROR t+ innate lymphoid cells. Science 330(6004), 665-669.
Contact
TWINCORE GmbH
Zentrum für Experimentelle und Klinische
Infektionsforschung
Feodor-Lynen-Str. 7 | 30625 Hannover
Telefon 0511 2200 27 102 | Fax 0511 2200 27 186
twincore@twincore.de | www.twincore.de

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Complete list of articles published in 2010 as well as
articles published and “in press” in 2011 by employees
working at TWINCORE
Research group Prof. Kalinke
• 1. Williams SK, Fairless R, Weise J, Kalinke U, Schulz-Schaeffer W &
Diem R (2011) Neuroprotective effects of the cellular prion protein in
autoimmune optic neuritis. American Journal of Pathology (in press).
• 2. Gratz MS, Suezer Y, Kremer M, Volz A, Majzoub M, Hanschmann
KM, Kalinke U, Schwantes A & Sutter G (2011) N1L is a virulence
factor of ectromelia virus and essential for in vivo spread upon
respiratory infection. Journal of Virology (in press).
• 3. Grabski E, Waibler Z, Schule S, Kloke BP, Sender LY, Panitz S,
Cichutek K, Schweizer M & Kalinke U (2011) Comparative analysis
of transduced primary human dendritic cells generated by the use
of three different lentiviral vector systems. Molecular Biotechnology
47(3), 262-269.
• 4. Kochs G, Bauer S, Vogt C, Frenz T, Tschopp J, Kalinke U &
Waibler Z (2010) Thogoto virus infection induces sustained type I
interferon responses that depend on RIG-I-like helicase signaling of
conventional dendritic cells. Journal of Virology 84(23), 12344-12350.
• 5. Frenz T, Waibler Z, Hofmann J, Hamdorf M, Lantermann M, Reizis
B, Tovey MG, Aichele P, Sutter G & Kalinke U (2010) Concomitant
IFNAR-triggering of T cells and of DC is required to promote
maximal MVA-induced T-Cell expansion. European Journal of
Immunology 40(10), 2769-2777.
• 6. Goossens P, Gijbels MJ, Zernecke A, Eijgelaar W, Vergouwe MN,
van der Made I, Vanderlocht J, Beckers L, Buurman WA, Daemen
MJ, Kalinke U, Weber C, Lutgens E & de Winther MP (2010) Myeloid
type I interferon signaling promotes atherosclerosis by stimulating
macrophage recruitment to lesions. Cell Metabolism 12(2), 142-153.
• 7. Lang PA, Recher M, Honke N, Scheu S, Borkens S, Gailus N,
Krings C, Meryk A, Kulawik A, Cervantes-Barragan L, Van Rooijen
N, Kalinke U, Ludewig B, Hengartner H, Harris N, Häussinger D,
Ohashi PS, Zinkernagel RM & Lang KS (2010) Tissue macrophages
suppress viral replication and prevent severe immunopathology in
an interferon-I-dependent manner in mice. Hepatology 52(1), 25-32.
• 8. Prinz M & Kalinke U (2010) New lessons about old molecules: how
type I interferons shape Th1/Th17-mediated autoimmunity in the
CNS. Trends in Molecular Medicine 16(8), 379-386. Review.
• 9. Murikinati S, Juttler E, Keinert T, Ridder DA, Muhammad S,
Waibler Z, Ledent C, Zimmer A, Kalinke U & Schwaninger M (2010)
Activation of cannabinoid 2 receptors protects against cerebral
ischemia by inhibiting neutrophil recruitment. FASEB Journal 24(3),
788-798.
• 10. Sender LY, Gibbert K, Suezer Y, Radeke HH, Kalinke U & Waibler
Z (2010) CD40 ligand-triggered human dendritic cells mount
interleukin-23 responses that are further enhanced by danger
signals. Molecular Immunology 47(6), 1255-1261.
• 11. Al Moussawi K, Ghigo E, Kalinke U, Alexopoulou L, Mege JL
& Desnues B (2010) Type I interferon induction is detrimental
during infection with the Whipple’s disease bacterium, Tropheryma
whipplei. PLoS Pathogens 6(1): e1000722.
• 12. Ilchmann A, Burgdorf S, Scheurer S, Waibler Z, Nagai R, Wellner
A, Yamamoto Y, Yamamoto H, Henle T, Kurts C, Kalinke U, Vieths S
& Toda M (2010) Glycation of a food allergen by the Maillard reaction
enhances its T-cell immunogenicity: role of macrophage scavenger
receptor class A type I and II. Journal of Allergy and Clinical
Immunology 125(1), 175-183 e171-111.
• 13. Kern M, Popov A, Scholz K, Schumak B, Djandji D, Limmer
A, Eggle D, Sacher T, Zawatzky R, Holtappels R, Reddehase MJ,
Hartmann G, Debey-Pascher S, Diehl L, Kalinke U, Koszinowski U,
Schultze J & Knolle PA (2010) Virally infected mouse liver endothelial
cells trigger CD8+ T-cell immunity. Gastroenterology 138(1), 336-346.
• 14. Malinarich FH, Grabski E, Worbs T, Chennupati V, Haas JD,
Schmitz S, Candia E, Quera R, Malissen B, Förster R, Hermoso M
& Prinz I (2010) Constant TCR triggering suggests that the TCR
expressed on intestinal intraepithelial γδ T cells is functional in vivo.
European Journal of Immunology 40(12), 3378-88.
• 15. Schneider CK, Salmikangas P, Jilma B, Flamion B, Todorova LR,
Paphitou A, Haunerova I, Maimets T, Trouvin JH, Flory E, Tsiftsoglou
A, Sarkadi B, Gudmundsson K, O’Donovan M, Migliaccio G, Ancans
J, Maciulaitis R, Robert JL, Samuel A, Ovelgonne JH, Hystad M, Fal
AM, Lima BS, Moraru AS, Turcani P, Zorec R, Ruiz S, Akerblom
L, Narayanan G, Kent A, Bignami F, Dickson JG, Niederwieser D,
Figuerola-Santos MA, Reischl IG, Beuneu C, Georgiev R, Vassiliou
M, Pychova A, Clausen M, Methuen T, Lucas S, Schussler-Lenz M,
Kokkas V, Buzas Z, MacAleenan N, Galli MC, Line A, Gulbinovic J,
Berchem G, Fraczek M, Menezes-Ferreira M, Vilceanu N, Hrubisko
M, Marinko P, Timon M, Cheng W, Crosbie GA, Meade N, di Paola
ML, VandenDriessche T, Ljungman P, D’Apote L, Oliver-Diaz O, Buttel
I & Celis P (2010) Challenges with advanced therapy medicinal
products and how to meet them. Nature Reviews Drug Discovery 9(3),
195-201. Review.
• 16. Buttel IC, Voller K & Schneider CK (2010) Immunogenicity and
its impact on benefit/risk considerations in the authorisation of
biopharmaceuticals. Current Drug Safety 5(4), 287-292. Review.
• 17. Goetz KB, Pfleiderer M & Schneider CK (2010) First-inhuman
clinical trials with vaccines-what regulators want. Nature
Biotechnology 28(9), 910-916. Review.
• 18. Schussler-Lenz M & Schneider CK (2010) [Clinical trials with
advanced therapy medicinal products]. Bundesgesundheitsblatt
Gesundheitsforschung Gesundheitsschutz 53(1), 68-74. Review.
Research group Prof. Pietschmann
• 1. Frentzen A, Hüging K, Bitzegeio J, Friesland M, Haid S, Gentzsch
J, Hoffmann M, Lindemann D, Zimmer G, Zielecki F, Weber F,
Steinmann E & Pietschmann T (2011) Completion of hepatitis C virus
replication cycle in heterokaryons excludes dominant restrictions in
human non-liver and mouse liver cell lines. PLoS Pathog (in press).
• 2. Gentzsch J, Hinkelmann B, Kaderali L, Irschik H, Jansen R, Sasse
F, Frank R & Pietschmann T (2011) Hepatitis C virus complete
life cycle screen for identification of small molecules with pro- or
antiviral activity. Antiviral Research 89(2),136-48. Epub 2010 Dec 15.
• 3. Montserret R, Saint N, Vanbelle C, Salvay AG, Simorre JP, Ebel
C, Sapay N, Renisio JG, Bockmann A, Steinmann E, Pietschmann
T, Dubuisson J, Chipot C & Penin F (2010) NMR structure and ion
channel activity of the p7 protein from hepatitis C virus. Journal of
Biological Chemistry 285(41), 31446-31461.
• 4. Steinmann E & Pietschmann T (2010) Hepatitis C virus P7 - a
viroporin crucial for virus assembly and an emerging target for
antiviral therapy. Viruses 2010 2(9), 2078-2095 (Review).
• 5. Bitzegeio J, Bankwitz D, Hueging K, Haid S, Brohm C, Zeisel MB,
Herrmann E, Iken M, Ott M & Baumert TF, Pietschmann T (2010)
Adaptation of hepatitis C virus to mouse CD81 permits infection of
mouse cells in the absence of human entry factors. PLoS Pathogens
6: e1000978.
• 6. Bürgel B, Friesland M, Koch A, Manns MP, Wedemeyer H,
Weissenborn K, Schulz-Schaeffer WJ, Pietschmann T, Steinmann
E & Ciesek S (2010) Hepatitis C virus enters human peripheral
neuroblastoma cells - evidence for extra-hepatic cells sustaining
hepatitis C virus penetration. Journal of Viral Hepatology Jun 23
[Epub ahead of print].

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• 7. Ciesek S, Friesland M, Steinmann J, Becker B, Wedemeyer H,
Manns MP, Pietschmann T & Steinmann E (2010) How stable is the
hepatitis C virus (HCV)? Environmental stability of HCV and its
susceptibility to chemical biocides. Journal of Infectious Diseases
201(12), 1859-1866.
• 8. Lemon SM, McKeating JA, Pietschmann T, Frick DN, Glenn JS,
Tellinghuisen TL, Symons J & Furman PA (2010) Development of
novel therapies for hepatitis C. Antiviral Research 86(1), 79-92
(Review).
• 9. Bankwitz D, Steinmann E, Bitzegeio J, Ciesek S, Friesland M,
Herrmann E, Zeisel MB, Baumert TF, Keck ZY, Foung SK, Pecheur
EI & Pietschmann T (2010) Hepatitis C virus hypervariable region 1
modulates receptor interactions, conceals the CD81 binding site, and
protects conserved neutralizing epitopes. Journal of Virology 84(11),
5751-5763.
• 10. Stegmann KA, Bjorkstrom NK, Veber H, Ciesek S, Riese P,
Wiegand J, Hadem J, Suneetha PV, Jaroszewicz J, Wang C, Schlaphoff
V, Fytili P, Cornberg M, Manns MP, Geffers R, Pietschmann T,
Guzman CA, Ljunggren HG & Wedemeyer H (2010) Interferon-alphainduced
TRAIL on natural killer cells is associated with control of
hepatitis C virus infection. Gastroenterology 138(5), 1885-1897.
• 11. Ciesek S, Steinmann E, Iken M, Ott M, Helfritz FA,
Wappler I, Manns MP, Wedemeyer H & Pietschmann T (2010)
Glucocorticosteroids increase cell entry by hepatitis C virus.
Gastroenterology 138(5), 1875-1884.
• 12. von Hahn T, Steinmann E, Ciesek S & Pietschmann T (2010)
Know your enemy: translating insights about the molecular biology
of hepatitis C virus into novel therapeutic approaches. Expert Review
on Gastroenterology & Hepatology 4(1), 63-79.
• 13. Steinmann J, Becker B, Bischoff B, Paulmann D, Friesland M,
Pietschmann T, Steinmann E (2010) Virucidal activity of 2 alcoholbased
formulations proposed as hand rubs by the World Health
Organization. American Journal of Infection Control 38(1), 66-68.
• 14. Haid S, Windisch MP, Bartenschlager R & Pietschmann T (2010)
Mouse-specific residues of claudin-1 limit hepatitis C virus genotype
2a infection in a human hepatocyte cell line. Journal of Virology
84(2), 964-975.
Research group Prof. Sparwasser
• 1. Hackl D, Loschko J, Sparwasser T, Reindl W & Krug A (2011)
Activation of dendritic cells via TLR7 reduces Foxp3 expression
and suppressive function in induced Tregs. European Journal of
Immunology (in press).
• 2. Paust HJ, Ostmann A, Erhardt A, Turner JE, Mittrücker HW,
Sparwasser T, Panzer U & Tiegs G (2011) Regulatory T cells control
the Th1 immune response in murine crescentic glomerulonephritis.
Kidney International (in press).
• 3. Sawa S, Lochner M, Satoh-Takayama N, Gaboriau-Routhiau V,
Dulauroy S, Bérard M, Kleinschek M, Cerf-Bensussan N, Cua D, Di
Santo JP & Eberl G (2011) RORgt+ innate lymphoid cells regulate
intestinal homeostasis by integrating negative signals from the
symbiotic microbiota. Nature Immunology Feb 20 [Epub ahead of
print].
• 4. Blankenhaus B, Klemm U, Eschbach ML, Sparwasser T,
Huehnn J, Kühl AA, Loddenkemper C, Jacobs T & Breloer M (2011)
Strongyloides ratti infection induces expansion of Foxp3+ regulatory
T cells that interfere with immune response and parasite clearance
in BALB/c mice. Journal of Immunology Feb 18 [Epub ahead of print].
• 5. Hadis U, Wahl B, Schulz O, Schippers A, Wagner N, Müller W,
Sparwasser T, Förster R & Pabst O (2011) Cooperation of lymph nodes
and intestinal lamina propria in FoxP3+ regulatory T cell mediated
intestinal tolerance. Immunity 34(2), 237-246.
• 6. Stagg J, Divisekera U, Duret H, Sparwasser T, Teng MW, Darcy PK
& Smyth MJ (2011) CD73-deficient mice have increased anti-tumor
immunity and are resistant to experimental metastasis. Cancer
Research Feb 3 [Epub ahead of print].
• 7. Ohnmacht C*, Marques R*, Presley L*, Sawa S*, Lochner M* &
Eberl G (2011) Intestinal microbiota, evolution of the immune system
and the bad reputation of pro-inflammatory immunity. Cellular
Microbiology Jan 28 [Epub ahead of print] Review.
• 8. Mayer CT, Floess S, Baru AM, Lahl K, Huehn J & Sparwasser T
(2011) CD8+Foxp3+ T cells share developmental and phenotypic
features with classical CD4+Foxp3+ regulatory T cells but lack
potent suppressive activity. European Journal of Immunology 41(3),
716-725.
• 9. Lahl K & Sparwasser T (2011) In vivo depletion of Foxp3+ Tregs
using the DEREG mouse model. Methods in Molecular Biology 707,
157-172 Review.
• 10. Berod L, Heinemann C, Heink S, Escher A, Stadelmann C, Drube
S, Wetzker R, Norgauer J & Kamradt T (2011) PI3K deficiency delays
the onset of experimental autoimmune encephalomyelitis and
ameliorates its clinical outcome. European Journal of Immunology
41(3), 833-844.
• 11. Pellegrini M, Calzascia T, Toe JG, Preston SP, Lin AE, Elford
AR, ShahinianA, Lang PA, Lang KS, Morre M, Assouline B, Lahl K,
Sparwasser T, Tedder TF, Paik J, DePinho RA, Basta S, Ohashi PS
& Mak TW (2011) IL-7 engages multiple mechanisms to overcome
chronic viral infection and limit organ pathology. Cell 144(4), 601-
613.
• 12. Dietze KK, Zelinskyy G, Gibbert K, Schimmer S, Francois S,
Myers L, Sparwasser T, Hasenkrug KJ & Dittmer U (2011) Transient
depletion of regulatory T cells in transgenic mice reactivates virusspecific
CD8+ T cells and reduces chronic retroviral set points.
Proceedings of the National Academy of Sciences U S A 108(6), 2420-
2425.
• 13. Vaeth M, Gogishvili T, Bopp T, Klein M, Berberich-Siebelt F,
Gattenloehner S, Avots A, Sparwasser T, Grebe N, Schmitt E, Hunig
T, Serfling E & Bodor J (2011) Regulatory T cells facilitate the
nuclear accumulation of inducible cAMP early repressor (ICER) and
suppress nuclear factor of activated T cell c1 (NFATc1). Proceedings
of the National Academy of Sciences U S A 108(6), 2480-2485.
• 14. Lochner M, Bérard M, Sawa S, Hauer S, Gaboriau-Routhiau V,
Fernandez F, Bousso P, Cerf-Bensussan N & Eberl G (2011) Restricted
microbiota and absence of cognate TCR antigen leads to an
unbalanced generation of Th17 cells. Journal of Immunology 186(3),
1531-1537 (Epub 2010 Dec 22)
• 15. Lochner M, Ohnmacht C, Presley L, Bruhns P, Si-Tahar M, Sawa
S & Eberl G (2011) Microbiota-induced tertiary lymphoid tissues
aggravate inflammatory disease in the absence of RORgt and LTi
cells. Journal of Experimental Medicine 208(1), 125-134 (Epub 20 Dec
2010)
• 16. Haque A, Best SE, Amante FH, Mustafah S, Desbarrieres L, de
Labastida F, Sparwasser T, Hill GR & Engwerda CR (2010) CD4+
natural regulatory T cells prevent experimental cerebral malaria via
CTLA-4 when expanded in vivo. PLoS Pathogens 6(12), e1001221.
• 17. Engel, D, Koscielny A, Wehner S, Maurer J, Schiwon M, Franken
L, Schumak B, Limmer A, Sparwasser T, Hirner A, Knolle P, Kalff J
& Kurts C (2010) Th1 memory cells disseminate postoperative ileus
over the entire intestinal tract. Nature Medicine 16(12), 1407-1413.
• 18. Hubert S, Rissiek B, Klages K, Hühn J, Sparwasser T, Haag F,
Koch-Nolte F, Boyer O, Seman M & Adriouch S (2010) Extracellular
NAD+ shapes the Foxp3 + regulatory T cell compartment through
the ART2/P2X7 pathway. Journal of Experimental Medicine 207(12),
2561-2568.

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• 19. Teng MW, Ngiow SF, von Scheidt B, McLaughlin N, Sparwasser T
& Smyth MJ (2010) Conditional regulatory T-cell depletion releases
adaptive immunity preventing carcinogenesis and suppressing
established tumor growth. Cancer Research 70(20), 7800-7809.
• 20. Klages K, Mayer CT, Lahl K, Loddenkemper C, Teng MW, Ngiow
SF, Smyth MJ, Hamann A, Huehn J & Sparwasser T (2010) Selective
depletion of Foxp3+ regulatory T cells improves effective therapeutic
vaccination against established melanoma. Cancer Research 70(20),
7788-7799.
• 21. Sawa S, Cherrier M, Lochner M, Satoh-Takayama N, Fehling HJ,
Langa F, Di Santo JP & Eberl G (2010) Lineage relationship analysis
of ROR t+ innate lymphoid cells. Science 330(6004), 665-669.
• 22. Navarro S, Cossalter G, Chiavaroli C, Kanda A, Fleury S, Lazzari
A, Cazareth J, Sparwasser T, Dombrowicz D, Glaichenhaus N &
Julia V (2010) The oral administration of bacterial extracts prevents
asthma via the recruitment of regulatory T cells to the airways.
Mucosal Immunology 4(1), 53-65 (Epub 01 Sept. 2010).
• 23. Arnold I, Lee J, Amieva M, Roers A, Flavell R, Sparwasser T
& Müller A (2010) Tolerance rather than immunity protects from
Helicobacter pylori-induced gastric preneoplasia. Gastroenterology
140(1), 199-209 (Epub 22 June 2010).
• 24. Baru AM. Hartl A, Lahl K, Krishnaswamy JK, Fehrenbach H,
Yildirim A, Garn H, Renz H, Behrens G & Sparwasser T (2010)
Selective depletion of Foxp3+ regulatory T cells during sensitization
phase aggravates experimental allergic airway inflammation.
European Journal of Immunology 40(8), 2259-2266.
• 25. Pechloff K, Holch J, Ferch U, Schweneker M, Brunner K, Kremer
M, Sparwasser T, Quintanilla-Martinez L, Zimber-Strobl U, Streubel
B, Gewies A, Peschel C & Ruland J (2010) The fusion kinase ITK-
SYK mimics a T-cell receptor signal and drives oncogenesis in
conditional mouse models of peripheral T cell lymphoma. Journal of
Experimental Medicine 207(5), 1031-1044.
• 26. Suttner K, Depner M, Wetzke M, Klopp N, von Mutius E, Illig
T, Sparwasser T & Kabesch M (2010) Genetic variants harbored in
the forkhead box protein 3 locus increase hay fever risk. Journal of
Allergy and Clinical Immunology 125(6), 1395-1399.
• 27. Boissonnas A, Scholer-Dahirel A, Simon-Blancal V, Pace L, Valet
F, Kissenpfennig A, Sparwasser T, Malissen B, Fetler L & Amigorenga
S (2010) FoxP3+ T cells induce perforin-dependent dendritic cell
death in tumor-draining lymph nodes. Immunity 32(2), 266-278.
• 28. Schildknecht A, Brauer S, Brenner C, Lahl K, Schild H,
Sparwasser T*, Probst HC* & van den Broek M* (2010) FoxP3+
regulatory T cells essentially contribute to peripheral CD8+ T cell
tolerance induced by steady state dendritic cells. Proceedings of
the National Academy of Sciences U S A 107(1), 199-203 *(equally
contributed) (Epub 2009).
• 29. Anz D, Koelzer VH, Moder S, Thaler R, Schwerd T, Lahl
K, Sparwasser T, Besch R, Poeck H, Hornung V, Hartmann G,
Rothenfusser S, Bourquin C & Endres S (2010) Immunostimulatory
RNA blocks suppression by regulatory T cells. Journal of Immunology
184(2), 939-946 (Epub 2009).
Research group Prof. Häußler
• 1. Schmidt, J, Müsken M, Becker T, Magnowska Z, Bertinetti D,
Möller S, Zimmermann B, Herberg FW, Jänsch L & Häussler S (2011).
The Pseudomonas aeruginosa chemotaxis methyltransferase CheR1
impacts on bacterial surface sampling. PLoS One (in press).
• 2. Häussler S (2010) Multicellular signalling and growth of
Pseudomonas aeruginosa. International Journal of Medical
Microbiology 300(8), 544-548.
• 3. Pommerenke C, Müsken M, Becker T, Dotsch A, Klawonn F
& Häussler S (2010) Global genotype-phenotype correlations in
Pseudomonas aeruginosa. PLoS Pathogens 6(8). pii: e1001074.
• 4. Müsken M, Di Fiore S, Römling U & Häussler S (2010) A 96-wellplate-based
optical method for the quantitative and qualitative
evaluation of Pseudomonas aeruginosa biofilm formation and its
application to susceptibility testing. Nature Protocols 5(8), 1460-1469.
• 5. Häussler S & Parsek MR (2010) Biofilms 2009: new perspectives
at the heart of surface-associated microbial communities. Journal of
Bacteriology 192(12), 2941-2949.
• 6. Dötsch A, Klawonn F, Jarek M, Scharfe M, Blocker H & Häussler S
(2010) Evolutionary conservation of essential and highly expressed
genes in Pseudomonas aeruginosa. BMC Genomics 11, 234.
• 7. Müsken M, Di Fiore S, Dotsch A, Fischer R & Häussler S
(2010) Genetic determinants of Pseudomonas aeruginosa biofilm
establishment. Microbiology 156(Pt 2), 431-441.
Research group Prof. Ott
• 1. Sharma AD, Narain N, Händel EM, Iken M, Singhal N, Cathomen
T, Manns M, Schöler HR, Ott M & Cantz T (2011) MicroRNA-221
regulates FAS-induced fulminant liver failure. Hepatology [Epub
ahead of print].
• 2. Sharma AD, Razvan I, Bock M, Cantz T, Manns MP & Ott M (2011)
Liver (chapter 33); G. Steinhoff (ed.) Regenerative Medicine: From
protocol to patient, pp. 773-803.
• 3. Razvan I, Rüdrich U, Rothe M, Kirsch S, Maasoumy B, Narain
N, Verfaillied CM, Sancho-Bru P, Iken M, Popescu I, Schambach
A, Manns MP & Bock M (2011) Induction of a mature hepatocyte
phenotype in adult liver derived progenitor cells by ectopic
expression of transcription factors. Stem Cell Research Feb 24 [Epub
ahead of print].
• 4. Becker PD, Legrand N, van Geelen CM, Noerder M, Huntington
ND, Lim A, Yasuda E, Diehl SA, Scheeren FA, Ott M, Weijer K,
Wedemeyer H, Di Santo JP, Beaumont T, Guzman CA & Spits H (2010)
Generation of human antigen-specific monoclonal IgM antibodies
using vaccinated “human immune system” mice. PLoS One 5(10).
• 5. Sancho-Bru P, Roelandt P, Narain N, Pauwelyn K, Notelaers T,
Shimizu T, Ott M & Verfaillie C (2010) Directed differentiation of
murine-induced pluripotent stem cells to functional hepatocyte-like
cells. Journal of Hepatology 54(1), 98-107.
• 6. Asgari S, Pournasr B, Salekdeh GH, Ghodsizadeh A, Ott M &
Baharvand H (2010) Induced pluripotent stem cells: a new era for
hepatology. Journal of Hepatology 53(4), 738-751.
• 7. Bitzegeio J, Bankwitz D, Hueging K, Haid S, Brohm C, Zeisel MB,
Herrmann E, Iken M, Ott M, Baumert TF & Pietschmann T (2010)
Adaptation of hepatitis C virus to mouse CD81 permits infection of
mouse cells in the absence of human entry factors. PLoS Pathogens
6, e1000978.
• 8. Ciesek S, Steinmann E, Iken M, Ott M, Helfritz FA,
Wappler I, Manns MP, Wedemeyer H & Pietschmann T (2010)
Glucocorticosteroids increase cell entry by hepatitis C virus.
Gastroenterology 138(5), 1875-1884.
• 9. Hettwer M, Reis-Fernandes MA, Iken M, Ott M, Steinberg P &
Nau H (2010) Metabolic activation capacity by primary hepatocytes
expands the applicability of the embryonic stem cell test as
alternative to experimental animal testing. Reproductive Toxicology
30(1), 113-120.
• 10. Stange MA, Tutarel O, Pischke S, Schneider A, Strassburg CP,
Becker T, Barg-Hock H, Basturk M, Wursthorn K, Cornberg M, Ott
M, Greten TF, Manns MP & Wedemeyer H (2010) Fulminant hepatic
failure due to chemotherapy-induced hepatitis B reactivation: role of
rituximab. Zeitschrift für Gastroenterologie 48(2), 258-263.

148 SCIENTIFIC REPORTS | Publications
Publications 2010-2011
Dept. Molecular Infection Biology | Prof. Dr. Petra Dersch
• Driouch,H., Roth,A., Dersch,P., & Wittmann,C. (2010) Filamentous
fungi in good shape: Microparticles for tailor-made fungal morphology
and enhanced enzyme production. Bioengineered Bugs 2.
• Driouch,H., Roth,A., Dersch,P., & Wittmann,C. (2010) Optimized bio -
process for production of fructofuranosidase by recombinant Aspergillus
niger. Applied Microbiology and Biotechnology 87, 2011-2024.
• Fleissner,A. & Dersch,P. (2010) Expression and export: recombinant
protein production systems for Aspergillus. Applied Microbiology and
Biotechnology 87, 1255-1270.
• Göpel,Y., Luttmann,D., Heroven,A.K., Reichenbach,B., Dersch,P., &
Görke,B. (2010) Common and divergent features in transcriptional
control of the homologous small RNAs GlmY and GlmZ in Enterobacteriaceae.
Nucleic Acids Research 39, 1294-1309.
• Heroven,A.K. & Dersch,P. (2010) Global virulence gene regulation
networks in enteropathogenic Yersiniae. In: Research Advances in
Molecular Microbiology (Mohan,R.M., ed.), Global Research Network,
Kerala, Indien, pp. 1-17.
• Homann,A., Qamar,R.-U., Serim,S., Dersch,P., & Seibel,J. (2010)
Bioorthogonal metabolic glycoengineering of human larynx carcinoma
(HEp-2) cells targeting sialic acid. Beilstein Journal of Organic
Chemistry 6.
• Quade,N., Dieckmann,M., Haffke,M., Heroven,A.K., Dersch,P., &
Heinz,D.W.*. (2011) Structure of the effector-binding domain of the
LysR-type transcription factor RovM from Yersinia pseudotuberculosis.
Acta Crystallographica Section D: Biological Crystallography 67,
81-90.
• Dersch,P. & Jahn,D. (2011) Mikrobielle Genetik. In Mikrobiologie
(Munk,K., ed.), Thieme., in press
RG Chronic Pseudomonas Infections | Prof. Dr. Susanne Häußler
• Dötsch,A., Klawonn,F., Jarek,M., Scharfe,M., Blöcker,H., & Häußler,S.
(2010) Evolutionary conservation of essential and highly expressed
genes in Pseudomonas aeruginosa. BMC GENOMICS 11, 234.
• Häussler,S. (2010) Multicellular signalling and growth of Pseudomonas
aeruginosa. International Journal of Medical Microbiology
300, 544-548.
• Häußler,S. & Parsek,M.R. (2010) Biofilms 2009: new perspectives at
the heart of surface-associated microbial communities. Journal of
Bacteriology 192, 2941-2949.
• Häußler,S. (2010) Multicellular signalling and growth of Pseudomonas
aeruginosa. International Journal of Medical Microbiology
300, 544-548.
• Moreno,A.M., Matz,C., Kjelleberg,S., & Manefield,M. (2010) Identification
of ciliate grazers of autotrophic bacteria in ammonia-oxidizing
activated sludge by RNA stable isotope probing. Applied and Environmental
Microbiology 76, 2203-2211.
• Müsken,M., Di,F.S., Dötsch,A., Fischer,R., & Häußler,S. (2010) Genetic
determinants of Pseudomonas aeruginosa biofilm establishment.
Microbiology 156, 431-441.
• Müsken,M., Di,F.S., Römling,U., & Häußler,S. (2010) A 96-well-platebased
optical method for the quantitative and qualitative evaluation
of Pseudomonas aeruginosa biofilm formation and its application to
susceptibility testing. Nature Protocols 5, 1460-1469.
• Pommerenke,C., Müsken,M., Becker,T., Dötsch,A., Klawonn,F., &
Häußler,S. (2010) Global genotype-phenotype correlations in Pseudomonas
aeruginosa. PLoS Pathogens 6, 89-90.
• Haddad,A., Jensen,V., Becker,T., & Häußler,S. (2011) The Pho regulon
influences biofilm formation and type three secretion in Pseudomonas
aeruginosa. Environmental Microbiology Reports 1, 488-494.
• Schmidt,J., Müsken,M., Becker,T., Magnowska,Z., Bertinetti,D.,
Moller,S., Zimmermann,B., Herberg,F.W., Jansch,L., & Häußler,S.
(2011) The Pseudomonas aeruginosa chemotaxis methyltransferase
CheR1 impacts on bacterial surface sampling. PLoS ONE 6, e18184.
RG Microbial Communication | Prof. Dr. Irene Wagner-Döbler
• Bodor,A.M., Jänsch,L., Wissing,J., & Wagner-Döbler,I. (2011) The luxS
mutation causes loosely-bound biofilms in Shewanella oneidensis.
BMC Research Notes 4, 180.
• Jaramillo-Colorado,B., Olivero-Verbel,J., Stashenko,E.E., Wagner-
Döbler,I., & Kunze,B. (2011) Anti-quorum sensing activity of essential
oils from Colombian plants. Natural Product Research, DOI:10.1080/1
4786419.2011.557376
• Kunze,B., Reck,M., Dotsch,A., Lemme,A., Schummer,D., Irschik,H.,
Steinmetz,H., & Wagner-Döbler,I. (2010) Damage of Streptococcus
mutans biofilms by carolacton, a secondary metabolite from the
myxobacterium Sorangium cellulosum. BMC Microbiology 10, 199.
• Lemme,A., Grobe,L., Reck,M., Tomasch,J., & Wagner-Döbler,I. (2011)
Subpopulation-specific transcriptome analysis of competence-stimulating-peptide
induced Streptococcus mutans. Journal of Bacteriology
193, 1863-1877.
• Lemme,A., Sztajer,H., & Wagner-Döbler,I. (2010) Characterization of
mleR, a positive regulator of malolactic fermentation and part of the
acid tolerance response in Streptococcus mutans. BMC Microbiology
10, 58.
• Nawrath,T., Dickschat,J.S., Kunze,B., & Schulz,S. (2010) The biosynthesis
of branched dialkylpyrazines in myxobacteria. Chemistry and
Biodiversity 7, 2129-2144.
• Reck,M., Rutz,K., Kunze,B., Tomasch,J., Surapaneni,S.K., Schulz,S.,
& Wagner-Döbler,I. (2011) The biofilm inhibitor carolacton disturbs
membrane integrity and cell division of Streptococcus mutans
through the serine/threonine protein kinase PknB. Journal of
Bac teriology 193, 5692-5706.
• Riedel,T., Tomasch,J., Buchholz,I., Jacobs,J., Kollenberg,M., Gerdts,G.,
Wichels,A., Brinkhoff,T., Cypionka,H., & Wagner-Döbler,I. (2010) Constitutive
expression of the proteorhodopsin gene by a flavobacterium
strain representative of the proteorhodopsin-producing microbial
community in the North Sea. Applied in Environmental Microbiology
76, 3187-3197.
• Schulz,S., Dickschat,J.S., Kunze,B., Wagner-Döbler,I., Diestel,R.,
& Sasse,F. (2010) Biological activity of volatiles from marine and
terrestrial bacteria. Marine Drugs 8, 2976-2987.

SCIENTIFIC REPORTS | Publications
149
• Thiel,V., Brinkhoff,T., Dickschat,J.S., Wickel,S., Grunenberg,J.,
Wagner-Döbler,I., Simon,M., & Schulz,S. (2010) Identification and
biosynthesis of tropone derivatives and sulfur volatiles produced by
bacteria of the marine Roseobacter clade. Organic and Biomolecular
Chemistry 8, 234-246.
• Tomasch,J., Gohl,R., Bunk,B., Suarez-Diez,M., & Wagner-Döbler,I.
(2011) Transcriptional response of the photoheterotrophic marine
bacterium Dinoroseobacter shibae to changing light regimes. ISME
Journal, DOI:10.1038/ismej.2011.68
• Vilchez,R., Lemme,A., Ballhausen,B., Thiel,V., Schulz,S., Jansen,R.,
Sztajer,H., & Wagner-Döbler,I. (2010) Streptococcus mutans inhibits
Candida albicans hyphal formation by the fatty acid signaling molecule
trans-2-decenoic acid (SDSF). ChemBioChem 11, 1552-1562.
• Wagner-Döbler,I., Ballhausen,B., Berger,M., Brinkhoff,T., Buchholz,I.,
Bunk,B., Cypionka,H., Daniel,R., Drepper,T., Gerdts,G., Hahnke,S.,
Han,C., Jahn,D., Kalhoefer,D., Kiss,H., Klenk,H.P., Kyrpides,N.,
Liebl,W., Liesegang,H., Meincke,L., Pati,A., Petersen,J., Piekarski,T.,
Pommerenke,C., Pradella,S., Pukall,R., Rabus,R., Stackebrandt,E.,
Thole,S., Thompson,L., Tielen,P., Tomasch,J., von,J.M., Wanphrut,N.,
Wichels,A., Zech,H., & Simon,M. (2010) The complete genome sequence
of the algal symbiont Dinoroseobacter shibae: a hitchhiker’s
guide to life in the sea. ISME Journal 4, 61-77.
• Xue,X., Sztajer,H., Buddruhs,N., Petersen,J., Rohde,M., Talay,S.R.,
& Wagner-Döbler,I. (2011) Lack of the delta subunit of RNA polymerase
increases virulence related traits of Streptococcus mutans.
PLoS ONE 6, e20075.
• Xue,X., Tomasch,J., Sztajer,H., & Wagner-Döbler,I. (2010) The delta
subunit of RNA polymerase, RpoE, is a global modulator of Streptococcus
mutans environmental adaptation. Journal of Bacteriology
192, 5081-5092.
• Brinkhoff,T., Fischer,D., Vollmers,J., Vogel,S., Beardsley,C., Thole,S.,
Mussmann,M., Kunze,B., Wagner-Döbler,I., Daniel,R., & Simon,M.
(2011) Biogeography and phylogenetic diversity of a cluster of exclusively
marine myxobacteria. ISME Journal, in press
• Xue,X., Li,J., Wang,W., Sztajer,H., & Wagner-Döbler,I. (2011) The
global impact of the delta subunit RpoE of the RNA polymerase on
the proteome of Streptococcus mutans. Microbiology, in press
Dept. of Gene Regulation and Differentiation |
Prof. Dr. Hansjörg Hauser
• Badar,M., Hemmen,K., Nimtz,M., Stieve,M., Stiesch,M., Lenarz,T.,
Hauser,H., Möllmann,U., Vogt,S., Schnabelrauch,M., & Müller,P.P.*.
(2010) Evaluation of madurahydroxylactone as a slow release antibacterial
implant coating. Open Biomedical Engineering Journal 4,
263-270.
• Becker,J., Pavlakovic,H., Ludewig,F., Wilting,F., Weich,H.A.*.,
Albuquerque,R., Ambati,J., & Wilting,J. (2010) Neuroblastoma progression
correlates with downregulation of the lymphangiogenesis
inhibitor sVEGFR-2. Clinical Cancer Research 16, 1431-1441.
• Bergmann,A., Ahmad,S., Cudmore,M., Gruber,A.D., Wittschen,P.,
Lindenmaier,W., Christofori,G., Gross,V., Gonzalves,A.C., Grone,H.J.,
Ahmed,A., & Weich,H.A.*. (2010) Reduction of circulating soluble
Flt-1 alleviates preeclampsia-like symptoms in a mouse model.
Journal of Cellular and Molecular Medicine 14, 1857-1867.
• Charbord,P., Livne,E., Gross,G., Haupl,T., Neves,N.M., Marie,P.,
Bianco,P., & Jorgensen,C. (2010) Human bone marrow mesenchymal
stem cells: a systematic reappraisal via the genostem experience.
Stem Cell Reviews and Reports 7, 32-42.
• Cohen-Haguenauer,O., Creff,N., Cruz,P., Tunc,C., Aiuti,A., Baum,C.,
Bosch,F., Blomberg,P., Cichutek,K., Collins,M., Danos,O., Dehaut,F.,
Federspiel,M., Galun,E., Garritsen,H., Hauser,H., Hildebrandt,M.,
Klatzmann,D., Merten,O.W., Montini,E., O’Brien,T., Panet,A.,
Rasooly,L., Scherman,D., Schmidt,M., Schweitzer,M., Tiberghien,P.,
Vandendriessche,T., Ziehr,H., Yla-Herttuala,S., von,K.C., Gahrton,G.,
& Carrondo,M. (2010) Relevance of an academic GMP Pan-European
vector infra-structure (PEVI). Current Gene Therapy 10, 414-422.
• Coroadinha,A.S., Gama-Norton,L., Amaral,A.I., Hauser,H., Alves,P.M.,
& Cruz,P.E. (2010) Production of retroviral vectors: review. Current
Gene Therapy 10, 456-473.
• Courties,G., Seiffart,V., Presumey,J., Escriou,V., Scherman,D.,
Zwerina,J., Ruiz,G., Zietara,N., Jablonska,J., Weiss,S., Hoffmann,A.,
Jorgensen,C., Apparailly,F., & Gross,G. (2010) In vivo RNAi-mediated
silencing of TAK1 decreases inflammatory Th1 and Th17 cells
through targeting of myeloid cells. Blood 116, 3505-3516.
• Dietrich,N., Rohde,M., Geffers,R., Kroger,A., Hauser,H., Weiss,S., &
Gekara,N.O. (2010) Mast cells elicit proinflammatory but not type I
interferon responses upon activation of TLRs by bacteria. Proceeding
of the National Academy of Sciences of the United States of America
107, 8748-8753.
• Dittmar,K.E.*., Simann,M., Zghoul,N., Schön,O., Meyring,W.,
Hannig,H., Macke,L., Dirks,W.G., Miller,K., Garritsen,H.S.P., &
Lindenmaier,W. (2010) Quality of cell products: Authenticity, identity,
genomic stability and status of differentiation. Transfusion
Medicine and Hemotherapy 37, 57-64.
• Ehlert,N., Hoffmann,A., Luessenhop,T., Gross,G., Mueller,P.P.*.,
Stieve,M., Lenarz,T., & Behrens,P. (2010) Amino-modified silica surfaces
efficiently immobilize bone morphogenetic protein 2 (BMP2)
for medical purposes. Acta Biomaterialia 7, 1772-1779.
• Ehlert,N., Müller,P.P.*., Stieve,M., & Behrens,P. (2010) Immobilization
of alkaline phosphatase on modified silica coatings. Microporous and
Mesoporous Materials 131, 51-57.
• Gama-Norton,L., Herrmann,S., Schucht,R., Coroadinha,A.S., Low,R.,
Alves,P.M., Bartholomae,C.C., Schmidt,M., Baum,C., Schambach,A.,
Hauser,H., & Wirth,D. (2010) Retroviral vector performance in defined
chromosomal Loci of modular packaging cell lines. Human
Gene Therapy 21, 979-991.
• Garritsen,H.S., Macke,L., Meyring,W., Hannig,H., Pagelow,U.,
Wormann,B., Geffers,R., Dittmar,K.E.*., & Lindenmaier,W. (2010) Efficient
generation of clinical-grade genetically modified dendritic cells
for presentation of multiple tumor-associated proteins. Trans fusion
50, 831-842.
• Kirschning,C.J., Dreher,S., Maass,B., Fichte,S., Schade,J., Koster,M.,
Noack,A., Lindenmaier,W., Wagner,H., & Boldicke,T. (2010) Generation
of anti-TLR2 intrabody mediating inhibition of macrophage
surface TLR2 expression and TLR2-driven cell activation. BMC Biotechnology
10, 31.
• Loew,R., Meyer,Y., Kuehlcke,K., Gama-Norton,L., Wirth,D., Hauser,H.,
Stein,S., Grez,M., Thornhill,S., Thrasher,A., Baum,C., & Schambach,A.
(2010) A new PG13-based packaging cell line for stable production of
clinical-grade self-inactivating gamma-retroviral vectors using targeted
integration. Gene Therapy 17, 272-280.
• Macke,L., Garritsen,H.S., Meyring,W., Hannig,H., Pagelow,U.,
Wormann,B., Piechaczek,C., Geffers,R., Rohde,M., Lindenmaier,W.,
& Dittmar,K.E.*. (2010) Evaluating maturation and genetic modification
of human dendritic cells in a new polyolefin cell culture bag
system. Transfusion 50, 843-855.
• May,T., Butueva,M., Bantner,S., Markusic,D., Seppen,J.,
MacLeod,R.A., Weich,H., Hauser,H., & Wirth,D. (2010) Synthetic gene
regulation circuits for control of cell expansion. Tissue Engineering –
Part A 16, 441-452.
• Nehlsen,K., Herrmann,S., Zauers,J., Hauser,H., & Wirth,D. (2010)
Toxin-antitoxin based transgene expression in mammalian cells.
Nucleic Acids Research 38, e32.
• Pulverer,J.E., Rand,U., Lienenklaus,S., Kugel,D., Zietara,N., Kochs,G.,
Naumann,R., Weiss,S., Staeheli,P., Hauser,H., & Koster,M. (2010)
Temporal and spatial resolution of type I and III interferon responses
in vivo. Journal of Virology 84, 8626-8638.
• Roming,M., Lünsdorf,H., Dittmar,K.E.*., & Feldmann,C. (2010)
ZrO(HPO(4))(1-x)(FMN)(x): quick and easy synthesis of a nanoscale
luminescent biomarker. Angewandte Chemie International Edition
49, 632-637.

150 SCIENTIFIC REPORTS | Publications
• Sandhu,U., Cebula,M., Behme,S., Riemer,P., Wodarczyk,C.,
Metzger,D., Reimann,J., Schirmbeck,R., Hauser,H., & Wirth,D. (2010)
Strict control of transgene expression in a mouse model for sensitive
biological applications based on RMCE compatible ES cells. Nucleic
Acids Research 39, e1.
• Schniedermann,J., Rennecke,M., Buttler,K., Richter,G., Stadtler,A.M.,
Norgall,S., Badar,M., Barleon,B., May,T., Wilting,J., & Weich,H.A.*.
(2010) Mouse lung contains endothelial progenitors with high capacity
to form blood and lymphatic vessels. BMC Cell Biology 11, 50.
• Schucht,R., Wirth,D., & May,T. (2010) Precise regulation of transgene
expression level and control of cell physiology. Cell Biology and
Toxicology 26, 29-42.
• Shahab-Osterloh,S., Witte,F., Hoffmann,A., Winkel,A., Laggies,S.,
Neumann,B., Seiffart,V., Lindenmaier,W., Gruber,A.D., Ringe,J.,
Haupl,T., Thorey,F., Willbold,E., Corbeau,P., & Gross,G. (2010) Mesenchymal
stem cell-dependent formation of heterotopic tendon-bone
insertions (osteotendinous junctions). Stem Cells 28, 1590-1601.
• Stammen,S., Schuller,F., Dietrich,S., Gamer,M., Biedendieck,R.,
& Jahn,D. (2010) Application of Escherichia coli phage K1E DNAdependent
RNA polymerase for in vitro RNA synthesis and in vivo
protein production in Bacillus megaterium. Applied Microbiology and
Biotechnology 88, 529-539.
• Stirnweiss,A., Ksienzyk,A., Klages,K., Rand,U., Grashoff,M.,
Hauser,H., & Kröger,A. (2010) IFN regulatory factor-1 bypasses IFNmediated
antiviral effects through viperin gene induction. Journal of
Immunology 184, 5179-5185.
• Sun,L., Hemgard,G.V., Susanto,S.A., & Wirth,M. (2010) Caveolin-1
influences human influenza A virus (H1N1) multiplication in cell
culture. Virology Journal 7, 108.
• Thorey,F., Menzel,H., Lorenz,C., Gross,G., Hoffmann,A., &
Windhagen,H. (2010) Enhancement of endoprosthesis anchoring
using BMP-2. Technology and Health Care 18, 217-229.
• Wirth,D., Gama-Norton,L., Schucht,R., Nehlsen,K., & Hauser,H. (2010)
Site-directed engineering of defined chromosomal sites for recombinant
protein and virus expression. BioPharm International 22, 32-39.
• Buttler,K., Schniedermmann,J., Weich,H.A.*., & Wilting,J. (2011)
Isolierung bipotenter endothelialer Vorläuferzellen aus der Lunge
adulter Mäuse [Isolation of bipotent endothelial precursor cells from
lungs of adult mice]. Lymphologie in Forschung und Praxis 14, 58-64.
• Carrondo,M., Panet,M., Wirth,D., Coroadinha,A.S., Cruz,P., Falk,H.,
Schucht,R., Dupont,F., Geny-Fiamma,C., Merten,O.W., & Hauser,H.
(2011) Integrated strategy for the production of therapeutic retroviral
vectors. Human Gene Therapy 22, 370-379.
• Ehlert,N., Badar,M., Christel,A., Lohmeier,S.J., Luessenhop,T.,
Stieve,M., Lenarz,T., Müller,P.P.*., & Behrens,P. (2011) Mesoporous
silica coatings for controlled release of the antibiotic ciprofloxacin
from implants. Journal of Materials Chemistry 21, 752-760.
• Kugel,D., Pulverer,J.E., Koster,M., Hauser,H., & Staeheli,P. (2011)
Novel nonviral bioassays for mouse type I and type III interferon.
Journal of Interferone and Cytokine Research 31, 345-349.
• Lindenmaier,W., Lachmann,K., Meyring,W., Garritsen,H.S.P.,
Thomas,M., & Dittmar,K.J.*. (2011) Innenbeschichtung von Beuteln
für die Kultur adhärenter humaner Zellen. Biospektrum 17, 32-34.
• Lohmeyer,J.A., Liu,F., Kruger,S., Lindenmaier,W., Siemers,F., &
Machens,H.G. (2011) Use of gene-modified keratinocytes and fibroblasts
to enhance regeneration in a full skin defect. Langenbeck’s
Archives of Surgery.
• Lorenz,C., Hoffmann,A., Gross,G., Windhagen,H., Dellinger,P.,
Mohwald,K., Dempwolf,W., & Menzel,H. (2011) Coating of titanium
implant materials with thin polymeric films for binding the signaling
protein BMP2. Macromolecular Bioscience 11, 234-244.
• Lyszkiewicz,M., Zietara,N., Rohde,M., Gekara,N.O., Jablonska,J.,
Dittmar,K.E.*., & Weiss,S. (2011) SIGN-R1+MHC II+ cells of the
splenic marginal zone--a novel type of resident dendritic cells.
Journal of Leukocyte Biology 89, 607-615.
• Thorey,F., Menzel,H., Lorenz,C., Gross,G., Hoffmann,A., &
Windhagen,H. (2011) Osseointegration by bone morphogenetic
protein-2 and transforming growth factor beta2 coated titanium
implants in femora of New Zealand white rabbits. Indian Journal of
Orthopaedics 45, 57-62.
RG Molecular Immunology | Dr. Siegfried Weiss
• Courties,G., Seiffart,V., Presumey,J., Escriou,V., Scherman,D.,
Zwerina,J., Ruiz,G., Zietara,N., Jablonska,J., Weiss,S., Hoffmann,A.,
Jorgensen,C., Apparailly,F., & Gross,G. (2010) In vivo RNAi-mediated
silencing of TAK1 decreases inflammatory Th1 and Th17 cells
through targeting of myeloid cells. Blood 116, 3505-3516.
• Dietrich,N., Rohde,M., Geffers,R., Kröger,A., Hauser,H., Weiss,S., &
Gekara,N.O. (2010) Mast cells elicit proinflammatory but not type I
interferon responses upon activation of TLRs by bacteria. Proceeding
of the National Academy of Sciences of the United States of America
107, 8748-8753.
• Dietrich,N., Lienenklaus,S., Weiss,S., & Gekara,N. (2010) Murine tolllike
receptor 2 activation induces type I interferon responses from
endolysosomal compartments. PLoS ONE 5, e10250.
• Dolowschiak,T., Chassin,C., Ben,M.S., Fuchs,T.M., Weiss,S.,
Vandewalle,A., & Hornef,M.W. (2010) Potentiation of epithelial innate
host responses by intercellular communication. PLoS Pathogens 6,
e1001194.
• Düber,S. & Weiss,S. (2010) Inducible B cell development. G. I. T.
Laboratory Journal Europe 7-8, 28-29.
• Gekara,N.O., Zietara,N., Geffers,R., & Weiss,S. (2010) Listeria monocytogenes
induces T cell receptor unresponsiveness through poreforming
toxin listeriolysin O. Journal of Infectious Diseases 202,
1698-1707.
• Jablonska,J., Leschner,S., Westphal,K., Lienenklaus,S., & Weiss,S.
(2010) Neutrophils responsive to endogenous IFN-beta regulate
tumor angiogenesis and growth in a mouse tumor model. Journal of
Clinical Investigation 120, 1151-1164.
• Jellusova,J., Duber,S., Guckel,E., Binder,C.J., Weiss,S., Voll,R., &
Nitschke,L. (2010) Siglec-G regulates B1 cell survival and selection.
Journal of Immunology 185, 3277-3284.
• Leschner,S. & Weiss,S. (2010) Salmonella-allies in the fight against
cancer. Journal of Molecular Medicine 88, 763-773.
• Leschner,S., Wolf,K., & Weiss,S. (2010) Bakterielle Infektion auf
Rezept. labor&more 5, 14-17.
• Leschner,S. & Weiss,S. (2010) Bacteria - fighters against cancer.
G. I. T. Laboratory Journal Europe 9, 20-21.
• Prajeeth,C.K., Jirmo,A.C., Krishnaswamy,J.K., Ebensen,T.,
Guzmán,C.A.*., Weiss,S., Constabel,H., Schmidt,R.E., & Behrens,G.M.
(2010) The synthetic TLR2 agonist BPPcysMPEG leads to efficient
cross-priming against co-administered and linked antigens. European
Journal of Immunology 40, 1272-1283.
• Pulverer,J.E., Rand,U., Lienenklaus,S., Kugel,D., Zietara,N., Kochs,G.,
Naumann,R., Weiss,S., Staeheli,P., Hauser,H., & Koster,M. (2010)
Temporal and spatial resolution of type I and III interferon responses
in vivo. Journal of Virology 84, 8626-8638.
• Steidle,S., Martinez-Sobrido,L., Mordstein,M., Lienenklaus,S., Garcia-
Sastre,A., Staheli,P., & Kochs,G. (2010) Glycine 184 in nonstructural
protein NS1 determines the virulence of influenza A virus strain PR8
without affecting the host interferon response. Journal of Virology 84,
12761-12770.
• Lyszkiewicz,M., Zietara,N., Rohde,M., Gekara,N.O., Jablonska,J.,
Dittmar,K.E.*., & Weiss,S. (2011) SIGN-R1+MHC II+ cells of the
splenic marginal zone--a novel type of resident dendritic cells. Journal
of Leukocyte Biology 89, 607-615.

SCIENTIFIC REPORTS | Publications
151
• Crull,K., Bumann,D., & Weiss,S. (2011) Influence of infection route
and virulence factors on colonization of solid tumors by Salmonella
enterica serovar Typhimurium. FEMS Immunology and Medical Microbiology,
in press
• Solodova,E., Jablonska,J., Weiss,S., & Lienenklaus,S. (2011) Production
of IFN-beta Listeria monocytogenes infection is restricted to the
monocyte/macrophage lineage. PLOS ONE, in press
• Xu,M.Z., Chan,S.W., Liu,A.M., Wong,K.F., Fan,S.T., Chen,J., Poon,R.T.,
Zender,L., Lowe,S.W., Hong,W., & Luk,J.M. (2011) AXL receptor kinase
is a mediator of YAP-dependent oncogenic functions in hepatocellular
carcinoma. Oncogene 30(10), 1229-1240
• Sawey,E., Chanrion,M., Cai,C., Wu,G., Zhang,J., Zender,L., Zhao,A.,
Busuttil,R., Yee,H., Stein,L., French,D., Finn,R., Lowe,S. & Powers,S.
(2011) Identification of a Therapeutic Strategy Targeting Amplified
FGF19 in Liver Cancer by Oncogenomic Screening. Cancer Cell 19(3),
347-358
RG Model Systems for Infection | Dr. Dagmar Wirth
• Gama-Norton,L., Herrmann,S., Schucht,R., Coroadinha,A.S., Low,R.,
Alves,P.M., Bartholomae,C.C., Schmidt,M., Baum,C., Schambach,A.,
Hauser,H., & Wirth,D. (2010) Retroviral vector performance in defined
chromosomal Loci of modular packaging cell lines. Human
gene therapy 21, 979-991.
• Loew,R., Meyer,Y., Kuehlcke,K., Gama-Norton,L., Wirth,D., Hauser,H.,
Stein,S., Grez,M., Thornhill,S., Thrasher,A., Baum,C., & Schambach,A.
(2010) A new PG13-based packaging cell line for stable production of
clinical-grade self-inactivating gamma-retroviral vectors using targeted
integration. Gene Therapy 17, 272-280.
• May,T., Butueva,M., Bantner,S., Markusic,D., Seppen,J.,
MacLeod,R.A., Weich,H., Hauser,H., & Wirth,D. (2010) Synthetic gene
regulation circuits for control of cell expansion. Tissue Engineering -
Part A 16, 441-452.
• Nehlsen,K., Herrmann,S., Zauers,J., Hauser,H., & Wirth,D. (2010)
Toxin-antitoxin based transgene expression in mammalian cells.
Nucleic Acids Research 38, e32.
• Sandhu,U., Cebula,M., Behme,S., Riemer,P., Wodarczyk,C.,
Metzger,D., Reimann,J., Schirmbeck,R., Hauser,H., & Wirth,D. (2010)
Strict control of transgene expression in a mouse model for sensitive
biological applications based on RMCE compatible ES cells. Nucleic
Acids Research 39, e1.
• Schucht,R., Wirth,D., & May,T. (2010) Precise regulation of transgene
expression level and control of cell physiology. Cell Biology and
Toxicology 26, 29-42.
• Wirth,D., Gama-Norton,L., Schucht,R., Nehlsen,K., & Hauser,H. (2010)
Site-directed engineering of defined chromosomal sites for recombinant
protein and virus expression. BioPharm International 22, 32-39.
• Carrondo,M., Panet,M., Wirth,D., Coroadinha,A.S., Cruz,P., Falk,H.,
Schucht,R., Dupont,F., Geny-Fiamma,C., Merten,O.W., & Hauser,H.
(2011) Integrated strategy for the production of therapeutic retroviral
vectors. Human Gene Therapy 22, 370-379.
JRG Chronic Infections and Cancer | Prof. Dr. Lars Zender
• Gurlevik,E., Woller,N., Struver,N., Schache,P., Kloos,A., Manns,M.P.,
Zender,L., Kuhnel,F., & Kubicka,S. (2010) Selectivity of oncolytic viral
replication prevents antiviral immune response and toxicity, but
does not improve antitumoral immunity. Molecular Therapy 18, 1972-
1982.
• Klawonn,F., Wüstefeld,T., & Zender,L. (2010) Statistical modelling for
data from experiments with short haripin RNAs. LNCS (Lecture Notes
in Computer Science (including subseries Lecture Notes in Artificial
Intelligence and Lecture Notes in Bioinformatics)) 6065, 79-90.
• Wuestefeld,T. & Zender,L. (2010) DLC1 and liver cancer: the Akt
connection. Gastroenterology 139, 1093-1096.
• Zender,L., Villanueva,A., Tovar,V., Sia,D., Chiang,D.Y., & Llovet,J.M.
(2010) Cancer gene discovery in hepatocellular carcinoma. Journal of
Hepatology 52, 921-929.
• Kühnel,F., Gürvelik,E., Wirth,T.C., Strüver,N., Malek,N.P., Müller-
Schilling,M., Manns,M.P., Carnero,A., Zender,L. & Kubicka,S. (2010)
Targeting of p53-transcriptional dysfunction by conditionally replicating
adenovirus is not limited by p53-homologues. Molecular
Therapie 18(5), 936-946
Dept. of Molecular Structural Biology | Prof. Dr. Dirk Heinz
• Brocker,M.J., Schomburg,S., Heinz,D.W.*, Jahn,D., Schubert,W.D., &
Moser,J. (2010) Crystal structure of the nitrogenase-like dark operative
protochlorophyllide oxidoreductase catalytic complex (ChlN/
ChlB)2. Journal of Biological Chemistry 285, 27336-27345.
• Carius,Y., Christian,H., Faust,A., Zander,U., Klink,B.U.*,
Kornberger,P., Kohring,G.W., Giffhorn,F., & Scheidig,A.J. (2010) Structural
insight into substrate differentiation of the sugar-metabolizing
enzyme galactitol dehydrogenase from Rhodobacter sphaeroides D.
Journal of Biological Chemistry 285, 20006-20014.
• Ferraris,D.M., Gherardi,E., Di,Y., Heinz,D.W.*, & Niemann,H. (2010)
Ligand-mediated dimerization of the Met receptor tyrosine kinase by
the bacterial invasion protein InIB. Journal of Molecular Biology 395,
522-532.
• Haffke,M., Menzel,A., Carius,Y., Jahn,D., & Heinz,D.W.* (2010) Structures
of the nucleotide-binding domain of the human ABCB6 transporter
and its complexes with nucleotides. Acta Crystallographica
Section D: Biological Crystallography 66, 979-987.
• Heinemann,I.U., Schulz,C., Schubert,W.D., Heinz,D.W.*, Wang,Y.G.,
Kobayashi,Y., Awa,Y., Wachi,M., Jahn,D., & Jahn,M. (2010) Structure
of the heme biosynthetic Pseudomonas aeruginosa porphobilinogen
synthase in complex with the antibiotic alaremycin. Antimicrobial
Agents and Chemotherapy 54, 267-272.
• Heinz,D.W.*., Betzel,C., & Wilmanns,M. (2010) Highlight: of systems
and structures. Biological Chemistry 391, 717-718.
• Hertiani,T., Edrada-Ebel,R., Ortlepp,S., van Soest,R.W., de Voogd,N.J.,
Wray,V., Hentschel,U., Kozytska,S., Muller,W.E., & Proksch,P. (2010)
From anti-fouling to biofilm inhibition: New cytotoxic secondary
metabolites from two Indonesian Agelas sponges. Bioorganic and
Medicinal Chemistry 18, 1297-1311.
• Ibrahim,S.R., Min,C.C., Teuscher,F., Ebel,R., Kakoschke,C., Lin,W.,
Wray,V., Edrada-Ebel,R., & Proksch,P. (2010) Callyaerins A-F and H,
new cytotoxic cyclic peptides from the Indonesian marine sponge
Callyspongia aerizusa. Bioorganic and Medicinal Chemistry 18, 4947-
4956.
• Kampfer,P., Busse,H.J., Tindall,B.J., Nimtz,M., & Grun-Wollny,I. (2010)
Nonomuraea rosea sp. nov. International Journal of Systematic and
Evolutionary Microbiology 60, 1118-1124.
• Klink,B.U., Barden,S., Heidler,T.V., Borchers,C., Ladwein,M.,
Stradal,T.B.*, Rottner,K., & Heinz,D.W.* (2010) Structure of Shigella
IpgB2 in complex with human RhoA: implications for the mechanism
of bacterial guanine nucleotide exchange factor mimicry. Journal of
Biological Chemistry 285, 17197-17208.
• Klink,B.U.* & Scheidig,A.J. (2010) New insight into the dynamic
properties and the active site architecture of H-Ras p21 revealed by
X-ray crystallography at very high resolution. BMC Structural Biology
10, 38.
• Klodmann,J., Sunderhaus,S., Nimtz,M., Jänsch,L., & Braun,H.P. (2010)
Internal architecture of mitochondrial complex I from Arabidopsis
thaliana. Plant Cell 22, 797-810.
• Layer,G., Reichelt,J., Jahn,D., & Heinz,D.W.* (2010) Structure and
function of enzymes in heme biosynthesis. Protein Science 19, 1137-
1161.

152 SCIENTIFIC REPORTS | Publications
• Marin,M., Perez-Pantoja,D., Donoso,R., Wray,V., Gonzalez,B., &
Pieper,D.H.* (2010) Modified 3-oxoadipate pathway for the biodegradation
of methylaromatics in Pseudomonas reinekei MT1. Journal of
Bacteriology 192, 1543-1552.
• Moghaddam,F.M., Farimani,M.M., Seirafi,M., Taheri,S., Khavasi,H.R.,
Sendker,J., Proksch,P., Wray,V., & Edrada,R. (2010) Sesterterpenoids
and other constituents of Salvia sahendica. Journal of Natural Products
73, 1601-1605.
• Palme,O., Comanescu,G., Stoineva,I., Radel,S., Benes,E., Develter,D.,
Wray,V., & Lang,S. (2010) Sophorolipids from Candida bombicola: Cell
separation by ultrasonic particle manipulation. European Journal of
Lipid Science and Technology 112, 663-673.
• Rand,K., Noll,C., Schiebel,H.M., Kemken,D., Dulcks,T., Kalesse,M.,
Heinz,D.W.*, & Layer,G. (2010) The oxygen-independent coproporphyrinogen
III oxidase HemN utilizes harderoporphyrinogen as a
reaction intermediate during conversion of coproporphyrinogen III to
protoporphyrinogen IX. Biological Chemistry 391, 55-63.
• Solbak,S.M., Reksten,T.R., Wray,V., Bruns,K., Horvli,O., Raae,A.J.,
Henklein,P., Henklein,P., Roder,R., Mitzner,D., Schubert,U., &
Fossen,T. (2010) The intriguing cyclophilin A-HIV-1 Vpr interaction:
prolyl cis/trans isomerisation catalysis and specific binding. BMC
Structural Biology 10, 31.
• Tomala,M., Lavrentieva,A., Moretti,P., Rinas,U., Kasper,C., Stahl,F.,
Schambach,A., Warlich,E., Martin,U., Cantz,T., & Scheper,T. (2010)
Preparation of bioactive soluble human leukemia inhibitory factor
from recombinant Escherichia coli using thioredoxin as fusion partner.
Protein Expression and Purification 73, 51-57.
• Wei,M.X., Hu,P., Wang,P., Naruse,S., Nokihara,K., Wray,V., & Ozaki,T.
(2010) Possible key residues that determine left gastric artery blood
flow response to PACAP in dogs. World Journal of Gastroenterology
16, 4865-4870.
• Wilke,S., Krausze,J., Gossen,M., Gröbe,L., Jager,V., Gherardi,E., van
den Heuvel, J., & Bussow,K. (2010) Glycoprotein production for
structure analysis with stable, glycosylation mutant CHO cell lines
established by fluorescence-activated cell sorting. Protein Science 19,
1264-1271.
• Wilkens,A., Paulsen,J., Wray,V., & Winterhalter,P. (2010) Structures
of two novel trimeric stilbenes obtained by horseradish peroxidase
catalyzed biotransformation of trans-resveratrol and (-)-epsilonviniferin.
Journal of Agricultural and Food Chemistry 58, 6754-6761.
• Wolfram,K., Schmidt,J., Wray,V., Milkowski,C., Schliemann,W., &
Strack,D. (2010) Profiling of phenylpropanoids in transgenic lowsinapine
oilseed rape (Brassica napus). Phytochemistry 71, 1076-1084.
• Xu,J., Aly,A.H., Wray,V., & Proksch,P. (2010) Polyketide derivatives of
endophytic fungus Pestalotiopsis sp. isolated from the Chinese mangrove
plant Rhizophora mucronata. Tetrahedron Letters 52, 21-25.
• Zakikhany,K., Harrington,C.R., Nimtz,M., Hinton,J.C., & Römling,U.
(2010) Unphosphorylated CsgD controls biofilm formation in Salmonella
enterica serovar Typhimurium. Molecular Microbiology 77,
771-786.
• Aly,A.H., Debbab,A., Clements,C., Edrada-Ebel,R., Orlikova,B.,
Diederich,M., Wray,V., Lin,W., & Proksch,P. (2011) NF kappa B inhibitors
and antitrypanosomal metabolites from endophytic fungus
Penicillium sp. isolated from Limonium tubiflorum. Bioorganic and
Medicinal Chemistry 19, 414-421.
• Esatbeyoglu,T., Jaschok-Kentner,B., Wray,V., & Winterhalter,P. (2011)
Structure elucidation of procyanidin oligomers by low-temperature
1H NMR spectroscopy. Journal of Agricultural and Food Chemistry
59, 62-69.
• Quade,N., Dieckmann,M., Haffke,M., Heroven,A.K., Dersch,P., &
Heinz,D.W.* (2011) Structure of the effector-binding domain of the
LysR-type transcription factor RovM from Yersinia pseudotuberculosis.
Acta Crystallographica Section D: Biological Crystallography 67,
81-90.
• Shiloach,J. & Rinas,U. (2011) Bacterial cultivation for producing
of proteins and other biological products. In Manual of industrial
microbiology and biotechnology (Demain,A.L., ed), American Society
for Microbiology (ASM), Washington DC., in press
• Fraatz,M.A., Riemer,S.J.L., Stober,R., Kaspera,R., Nimtz,M.,
Berger,R.G., & Zorn,H. (2011) A novel oxygenase from Pleurotus
sapidus transforms valencene to nootkatone. Journal of Molecular
Catalysis B: Enzymatic, in press
RG Biophysical Analysis | Dr. Victor Wray
• Hertiani,T., Edrada-Ebel,R., Ortlepp,S., van Soest,R.W., de Voogd,N.J.,
Wray,V., Hentschel,U., Kozytska,S., Muller,W.E., & Proksch,P. (2010)
From anti-fouling to biofilm inhibition: New cytotoxic secondary
metabolites from two Indonesian Agelas sponges. Bioorganic and
Medicinal Chemistry 18, 1297-1311.
• Ibrahim,S.R., Min,C.C., Teuscher,F., Ebel,R., Kakoschke,C., Lin,W.,
Wray,V., Edrada-Ebel,R., & Proksch,P. (2010) Callyaerins A-F and H,
new cytotoxic cyclic peptides from the Indonesian marine sponge
Callyspongia aerizusa. Bioorganic and Medicinal Chemistry 18, 4947-
4956.
• Marin,M., Perez-Pantoja,D., Donoso,R., Wray,V., Gonzalez,B., &
Pieper,D.H.*. (2010) Modified 3-oxoadipate pathway for the biodegradation
of methylaromatics in Pseudomonas reinekei MT1. Journal of
Bacteriology 192, 1543-1552.
• Moghaddam,F.M., Farimani,M.M., Seirafi,M., Taheri,S., Khavasi,H.R.,
Sendker,J., Proksch,P., Wray,V., & Edrada,R. (2010) Sesterterpenoids
and other constituents of Salvia sahendica. Journal of Natural
Products 73, 1601-1605.
• Palme,O., Comanescu,G., Stoineva,I., Radel,S., Benes,E., Develter,D.,
Wray,V., & Lang,S. (2010) Sophorolipids from Candida bombicola:
Cell separation by ultrasonic particle manipulation. European Journal
of Lipid Science and Technology 112, 663-673.
• Rand,K., Noll,C., Schiebel,H.M., Kemken,D., Dulcks,T., Kalesse,M.,
Heinz,D.W.*, & Layer,G. (2010) The oxygen-independent coproporphyrinogen
III oxidase HemN utilizes harderoporphyrinogen as a
reaction intermediate during conversion of coproporphyrinogen III to
protoporphyrinogen IX. Biological Chemistry 391, 55-63.
• Solbak,S.M., Reksten,T.R., Wray,V., Bruns,K., Horvli,O., Raae,A.J.,
Henklein,P., Henklein,P., Roder,R., Mitzner,D., Schubert,U., &
Fossen,T. (2010) The intriguing cyclophilin A-HIV-1 Vpr interaction:
prolyl cis/trans isomerisation catalysis and specific binding. BMC
Structural Biology 10, 31.
• Tomala,M., Lavrentieva,A., Moretti,P., Rinas,U., Kasper,C., Stahl,F.,
Schambach,A., Warlich,E., Martin,U., Cantz,T., & Scheper,T. (2010)
Preparation of bioactive soluble human leukemia inhibitory factor
from recombinant Escherichia coli using thioredoxin as fusion partner.
Protein Expression and Purification 73, 51-57.
• Wei,M.X., Hu,P., Wang,P., Naruse,S., Nokihara,K., Wray,V., & Ozaki,T.
(2010) Possible key residues that determine left gastric artery blood
flow response to PACAP in dogs. World Journal of Gastroenterology
16, 4865-4870.
• Wilke,S., Krausze,J., Gossen,M., Gröbe,L., Jager,V., Gherardi,E., van
den Heuvel, J., & Büssow,K. (2010) Glycoprotein production for
structure analysis with stable, glycosylation mutant CHO cell lines
established by fluorescence-activated cell sorting. Protein Science 19,
1264-1271.
• Wilkens,A., Paulsen,J., Wray,V., & Winterhalter,P. (2010) Structures
of two novel trimeric stilbenes obtained by horseradish peroxidase
catalyzed biotransformation of trans-resveratrol and (-)-epsilonviniferin.
Journal of Agricultural and Food Chemistry 58, 6754-6761.
• Wolfram,K., Schmidt,J., Wray,V., Milkowski,C., Schliemann,W., &
Strack,D. (2010) Profiling of phenylpropanoids in transgenic low-sinapine
oilseed rape (Brassica napus). Phytochemistry 71, 1076-1084.

SCIENTIFIC REPORTS | Publications
153
• Xu,J., Aly,A.H., Wray,V., & Proksch,P. (2010) Polyketide derivatives of
endophytic fungus Pestalotiopsis sp. isolated from the Chinese mangrove
plant Rhizophora mucronata. Tetrahedron Letters 52, 21-25.
• Zakikhany,K., Harrington,C.R., Nimtz,M., Hinton,J.C., & Romling,U.
(2010) Unphosphorylated CsgD controls biofilm formation in Salmonella
enterica serovar Typhimurium. Molecular Microbiology 77,
771-786.
• Aly,A.H., Debbab,A., Clements,C., Edrada-Ebel,R., Orlikova,B.,
Diederich,M., Wray,V., Lin,W., & Proksch,P. (2011) NF kappa B inhibitors
and antitrypanosomal metabolites from endophytic fungus
Penicillium sp. isolated from Limonium tubiflorum. Bioorganic and
Medicinal Chemistry 19, 414-421.
• Esatbeyoglu,T., Jaschok-Kentner,B., Wray,V., & Winterhalter,P. (2011)
Structure elucidation of procyanidin oligomers by low-temperature
1H NMR spectroscopy. Journal of Agricultural and Food Chemistry
59, 62-69.
• Quade,N., Dieckmann,M., Haffke,M., Heroven,A.K., Dersch,P., &
Heinz,D.W.* (2011) Structure of the effector-binding domain of the
LysR-type transcription factor RovM from Yersinia pseudotuberculosis.
Acta Crystallographica Section D: Biological Crystallography 67,
81-90.
• Shiloach,J. & Rinas,U. (2011) Bacterial cultivation for producing
of proteins and other biological products. In Manual of industrial
microbiology and biotechnology (Demain,A.L., ed), American Society
for Microbiology (ASM), Washington DC., in press
• Fraatz,M.A., Riemer,S.J.L., Stober,R., Kaspera,R., Nimtz,M.,
Berger,R.G., & Zorn,H. (2011) A novel oxygenase from Pleurotus
sapidus transforms valencene to nootkatone. Journal of Molecular
Catalysis B: Enzymatic, in press
RG Recombinant Protein Expression | Dr. Joop van den Heuvel
• Lu,X., Sun,J., Nimtz,M., Wissing,J., Zeng,A.-P., & Rinas,U. (2010) The
intra- and extracellular proteome of Aspergillus niger growing on defined
medium with xylose or maltose as carbon substrate. Microbial
Cell Factories 9, 1-13.
• Wilke,S., Krausze,J., Gossen,M., Gröbe,L., Jager,V., Gherardi,E., van
den Heuvel, J., & Bussow,K. (2010) Glycoprotein production for
structure analysis with stable, glycosylation mutant CHO cell lines
established by fluorescence-activated cell sorting. Protein Science 19,
1264-1271.
• Bals,C., Schambach,A., Meyer,J., Scheper,T., & Rinas,U. (2011) Expression
and purification of bioactive soluble murine stem cell factor
from recombinant Escherichia coli using thioredoxin as fusion partner.
Journal of Biotechnology 152, 1-8.
• Shiloach,J. & Rinas,U. (2011) Bacterial cultivation for producing
of proteins and other biological products. In: Manual of industrial
microbiology and biotechnology (Demain,A.L., ed.), American
Society for Microbiology (ASM), Washington DC, in press.
RG Cellular Proteome Research | Dr. Lothar Jänsch
• Badar,M., Hemmen,K., Nimtz,M., Stieve,M., Stiesch,M., Lenarz,T.,
Hauser,H., Möllmann,U., Vogt,S., Schnabelrauch,M., & Müller,P.P.*
(2010) Evaluation of madurahydroxylactone as a slow release antibacterial
implant coating. Open Biomedical Engineering Journal 4,
263-270.
• Eisele,N., Linke,D., Bitzer,K., Na’amnieh,S., Nimtz,M., & Berger,R.G.
(2010) The first characterized asparaginase from a basidiomycete,
Flammulina velutipes. Bioresource Technology 102, 3316-3321.
• El-Banna,A., Hajirezaei,M.R., Wissing,J., Ali,Z., Vaas,L., Heine-Dobber
-nack,E., Jacobsen,H.J., Schumacher,H.M., & Kiesecker,H. (2010)
Over-expression of PR-10a leads to increased salt and osmotic tolerance
in potato cell cultures. Journal of Biotechnology 150, 277-287.
• Klodmann,J., Sunderhaus,S., Nimtz,M., Jänsch,L., & Braun,H.P. (2010)
Internal architecture of mitochondrial complex I from Arabidopsis
thaliana. Plant Cell 22, 797-810.
• Lu,X., Sun,J., Nimtz,M., Wissing,J., Zeng,A.-P., & Rinas,U. (2010) The
intra- and extracellular proteome of Aspergillus niger growing on defined
medium with xylose or maltose as carbon substrate. Microbial
Cell Factories 9, 1-13.
• Maahs,D.M., Siwy,J., Argiles,A., Cerna,M., Delles,C., Dominiczak,A.F.,
Gayrard,N., Iphöfer,A., Jänsch,L., Jerums,G., Medek,K., Mischak,H.,
Navis,G.J., Roob,J.M., Rossing,K., Rossing,P., Rychlik,I., Schiffer,E.,
Schmieder,R.E., Wascher,T.C., Winklhofer-Roob,B.M., Zimmerli,L.U.,
Zurbig,P., & Snell-Bergeon,J.K. (2010) Urinary collagen fragments
are significantly altered in diabetes: a link to pathophysiology.
PLoS ONE 5.
• Mischak,H., Kolch,W., Aivaliotis,M., Bouyssie,D., Court,M., Dihazi,H.,
Dihazi,G.H., Franke,J., Garin,J., de Peredo,A.G., Iphöfer,A., Jänsch,L.,
Lacroix,C., Makridakis,M., Masselon,C., Metzger,J., Monsarrat,B.,
Mrug,M., Norling,M., Novak,J., Pich,A., Pitt,A., Bongcam-Rudloff,E.,
Siwy,J., Suzuki,H., Thongboonkerd,V., Wang,L.-S., Zoidakis,J.,
Zurbig,P., Schanstra,J.P., & Vlahou,A. (2010) Comprehensive human
urine standards for comparability and standardization in clinical
proteome analysis. Proteomics - Clinical Applications 4, 464-478.
• Pommerenke,C., Müsken,M., Becker,T., Dötsch,A., Klawonn,F., &
Häußler,S. (2010) Global genotype-phenotype correlations in Pseudomonas
aeruginosa. PLoS Pathogens 6, 89-90.
• Steinke,N., Kaller,M., Nimtz,M., Baro,A., & Laschat,S. (2010) Columnar
liquid crystals derived from crown ethers with two lateral estersubstituted
ortho-terphenyl units: Unexpected destabilisation of the
mesophase by potassium iodide. Liquid Crystals 37, 1139-1149.
• Stepanova,T., Smal,I., van,H.J., Akinci,U., Liu,Z., Miedema,M.,
Limpens,R., van,H.M., van der,R.M., Poot,R., Grosveld,F.,
Mommaas,M., Meijering,E., & Galjart,N. (2010) History-dependent
catastrophes regulate axonal microtubule behavior. Current biology
20, 1023-1028.
• Trunk,K., Benkert,B., Quack,N., Munch,R., Scheer,M., Garbe,J.,
Jänsch,L., Trost,M., Wehland,J., Buer,J., Jahn,M., Schobert,M., &
Jahn,D. (2010) Anaerobic adaptation in Pseudomonas aeruginosa:
definition of the Anr and Dnr regulons. Environmental Microbiology
12, 1719-1733.
• Tschumitschew,K. & Klawonn,F. (2010) Incremental quantile estimation.
Evolving Systems 1, 253-264.
• Tschumitschew,K. & Klawonn,F. (2010) The need for benchmarks
with data from stochastic processes and meta-models in evolving
systems. In Proceedings of the International Symposium on Evolving
Intelligent Systems (Angelov,P., Filev,D., & Kasabov,N., eds), pp. 30-
33. SSAISB, Leicester.
• Tschumitschew,K. & Klawonn,F. (2010) AVEDA: Statistical tests for
finding interesting visulatisations. 5711 LNAI (Part 1) Lecture Notes
in Computer Science (including subseries Lecture Notes in Artificial
Intelligence and Lecture Notes in Bioinformatics), 235-242.
• Krone,M. & Klawonn,F. (2011) Rank correlation coefficient correction
by removing worst cases. In: Information Processing and Management
of Uncertainty in Knowledge-Based Systems: Theory and Methods
(Hüllermeier,E., Kruse,R., & Hoffmann,F., eds.), Springer, Berlin,
pp. 356-364.
• Winkler,R., Klawonn,F., & Kruse,R. (2011) Fuzzy clustering with polynomial
fuzzifier function in connection with m-estimators. Applied
and Computational Mathematics 10, 146-163.
• Eberl,L. & Riedel,K. (2011) Mining quorum sensing regulated proteins
- role of bacterial cell-to-cell communication in global gene
regulation as assessed by proteomics. Proteomics, in press
• Hebecker,S., Arendt,W., Heinemann,I.U., Tiefenau,J.H., Nimtz,M.,
Rohde,M., Soll,D., & Moser,J. (2011) Alanyl-phosphatidylglycerol synthase:
mechanism of substrate recognition during tRNA-dependent
lipid modification in Pseudomonas aeruginosa. Molecular Microbiology,
in press.

154 SCIENTIFIC REPORTS | Publications
• Hemmen,K., Reinl,T., Buttler,K., Behler,F., Dieken,H., Jänsch,L.,
Wilting,J., & Weich,H.A.*. (2011) High-resolution mass spectrometric
analysis of the secretome from mouse lung endothelial progenitor
cells. Angiogenesis, in press.
RG Microbial Proteomics | Prof. Dr. Katharina Riedel
• Bosshard,F., Riedel,K., Schneider,T., Geiser,C., Bucheli,M., & Egli,T.
(2010) Protein oxidation and aggregation in UVA-irradiated
Escherichia coli cells as signs of accelerated cellular senescence.
Environmental Microbiology 12, 2931-2945.
• Carranza,P., Grunau,A., Schneider,T., Hartmann,I., Lehner,A.,
Stephan,R., Gehrig,P., Grossmann,J., Groebel,K., Hoelzle,L.E.,
Eberl,L., & Riedel,K. (2010) A gel-free quantitative proteomics approach
to investigate temperature adaptation of the food-borne pathogen
Cronobacter turicensis 3032. Proteomics 10, 3248-3261.
• Grunau,A. & Riedel,K. (2011) Exoenzymes. In Encyclopaedia of Geobiology
(Reiter,J. & Thiel,V., eds), pp. 355-359. Springer.
• Hartmann,I., Carranza,P., Lehner,A., Stephan,R., Eberl,L., & Riedel,K.
(2010) Genes involved in Cronobacter sakazakii biofilm formation.
Applied and Environmental Microbiology 76, 2251-2261.
• Lumjiaktase,P., Aguilar,C., Battin,T., Riedel,K., & Eberl,L. (2010)
Construction of self-transmissible green fluorescent protein-based biosensor
plasmids and their use for identification of N-acyl homoserine-producing
bacteria in lake sediments. Applied and Environmental
Microbiology 76, 6119-6127.
• Moretti,M., Grunau,A., Minerdi,D., Gehrig,P., Roschitzki,B., Eberl,L.,
Garibaldi,A., Gullino,M.L., & Riedel,K. (2010) A proteomics approach
to study synergistic and antagonistic interactions of the fungal-bacterial
consortium Fusarium oxysporum wild-type MSA 35. Proteomics
10, 3292-3320.
• Schneider,T. & Riedel,K. (2010) Environmental proteomics: analysis
of structure and function of microbial communities. Proteomics 10,
785-798.
• Schneider,T. & Riedel,K. (2010) Environmental proteomics: Studying
structure and function of microbial communities. In Geomicrobiology:
Molecular and Environmental Perspective (Barton,L.L.,
Mandl,M., & Loy,A., eds), pp. 91-108. Springer.
• Schneider,T., Gerrits,B., Gassmann,R., Schmid,E., Gessner,M.O.,
Richter,A., Battin,T., Eberl,L., & Riedel,K. (2010) Proteome analysis of
fungal and bacterial involvement in leaf litter decomposition. Proteomics
10, 1819-1830.
• Srinivas,N., Jetter,P., Ueberbacher,B.J., Werneburg,M., Zerbe,K.,
Steinmann,J., Van der,M.B., Bernardini,F., Lederer,A., Dias,R.L.,
Misson,P.E., Henze,H., Zumbrunn,J., Gombert,F.O., Obrecht,D.,
Hunziker,P., Schauer,S., Ziegler,U., Kach,A., Eberl,L., Riedel,K.,
DeMarco,S.J., & Robinson,J.A. (2010) Peptidomimetic antibiotics target
outer-membrane biogenesis in Pseudomonas aeruginosa. Science
327, 1010-1013.
JRG Macromolecular Interactions | Prof. Dr. Christiane Ritter
• Greenwald,J., Buhtz,C., Ritter,C., Kwiatkowski,W., Choe,S.,
Maddelein,M.L., Ness,F., Cescau,S., Soragni,A., Leitz,D., Saupe,S.J.,
& Riek,R. (2010) The mechanism of prion inhibition by HET-S. Molecules
and Cells 38, 889-899.
• Wasmer,C., Zimmer,A., Sabate,R., Soragni,A., Saupe,S.J., Ritter,C., &
Meier,B.H. (2010) Structural similarity between the prion domain of
HET-s and a homologue can explain amyloid cross-seeding in spite of
limited sequence identity. Journal of Molecular Biology 402, 311-325.
JRG Centre for Structural Systems Biology at DESY |
Prof. Dr. Inari Kursula
• Chen,W.Q., Salmazo,A., Myllykoski,M., Sjoblom,B., Bidlingmaier,M.,
Pollak,A., Baumgartel,P., Djinovic-Carugo,K., Kursula,P.*., & Lubec,G.
(2010) Purification of recombinant growth hormone by clear native
gels for conformational analyses: preservation of conformation and
receptor binding. Amino Acids 39, 859-869.
• Huttu,J., Singh,B.K., Bhargav,S.P., Sattler,J.M., Schuler,H., & Kursula,I.
(2010) Crystallization and preliminary structural characterization of
the two actin-depolymerization factors of the malaria parasite. Acta
Crystallographica Section F: Structural Biology and Crystallisation
Community 66, 583-587.
• Majava,V., Polverini,E., Mazzini,A., Nanekar,R., Knoll,W., Peters,J.,
Natali,F., Baumgartel,P., Kursula,I., & Kursula,P. (2010) Structural
and functional characterization of human peripheral nervous system
myelin protein P2. PLoS ONE 5, e10300.
• Myllykoski,M. & Kursula,P.*. (2010) Expression, purification, and
initial characterization of different domains of recombinant mouse
2’,3’-cyclic nucleotide 3’-phosphodiesterase, an enigmatic enzyme
from the myelin sheath. BMC Research Notes 3, 12.
• Suresh,S., Wang,C., Nanekar,R., Kursula,P.*., & Edwardson,J.M.
(2010) Myelin basic protein and myelin protein 2 act synergistically
to cause stacking of lipid bilayers. Biochemistry 49, 3456-3463.
• Kursula,P.*. (2011) Structural biology insights into the autoantingens
of the myelin sheath. In Autoimmune Diseases: Symptoms, Diagnosis
and Treatment Nova, in press.
Dept. of Medical Microbiology | Prof. Dr. G. Singh Chhatwal
• Abt,B., Foster,B., Lapidus,A., Clum,A., Sun,H., Pukall,R., Lucas,S.,
Glavina Del,R.T., Nolan,M., Tice,H., Cheng,J.F., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Ovchinnikova,G., Pati,A., Goodwin,L.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Göker,M., Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Cellulomonas flavigena type strain (134). Standards in
Genomic Sciences 3, 15-25.
• Anderson,I., Djao,O.D., Misra,M., Chertkov,O., Nolan,M., Lucas,S.,
Lapidus,A., Del Rio,T.G., Tice,H., Cheng,J.F., Tapia,R., Han,C., Good -
win,L., Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Pati,A., Brambilla,E., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Sikorski,J., Spring,S., Rohde,M., Eichinger,K.,
Huber,H., Wirth,R., Göker,M., Detter,J.C., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C.
(2010) Complete genome sequence of Methanothermus fervidus type
strain (V24S). Standards in Genomic Sciences 3, 315-324.
• Aziz,R.K., Kansal,R., Aronow,B.J., Taylor,W.L., Rowe,S.L., Kubal,M.,
Chhatwal,G.S*., Walker,M.J., & Kotb,M. (2010) Microevolution of
group A streptococci in vivo: capturing regulatory networks engaged
in sociomicrobiology, niche adaptation, and hypervirulence. PLoS
ONE 5, e9798.
• Bunk,B., Schulz,A., Stammen,S., Münch,R., Warren,M.J., Rohde,M.,
Jahn,D., & Biedendieck,R. (2010) A short story about a big magic bug.
Bioengineered Bugs 1, 85-91.
• Candela,M., Centanni,M., Fiori,J., Biagi,E., Turroni,S., Orrico,C.,
Bergmann,S., Hammerschmidt,S., & Brigidi,P. (2010) DnaK from
Bifidobacterium animalis subsp. lactis is a surface-exposed human
plasminogen receptor upregulated in response to bile salts. Microbiology
156, 1609-1618.
• Chang,Y.J., Pukall,R., Saunders,E., Lapidus,A., Copeland,A., Nolan,M.,
Glavina Del,R.T., Lucas,S., Chen,F., Tice,H., Cheng,J.F., Han,C.,
Detter,J.C., Bruce,D., Goodwin,L., Pitluck,S., Mikhailova,N., Liolios,K.,
Pati,A., Ivanova,N., Mavromatis,K., Chen,A., Palaniappan,K., Land,M.,
Hauser,L., Jeffries,C.D., Brettin,T., Rohde,M., Göker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P.
(2010) Complete genome sequence of Acidaminococcus fermentans
type strain (VR4). Standards in Genomic Sciences 3, 1-14.

SCIENTIFIC REPORTS | Publications
155
• Chertkov,O., Sikorski,J., Brambilla,E., Lapidus,A., Copeland,A., Glavina
Del,R.T., Nolan,M., Lucas,S., Tice,H., Cheng,J.F., Han,C., Detter,J.C.,
Bruce,D., Tapia,R., Goodwin,L., Pitluck,S., Liolios,K., Ivanova,N.,
Mavromatis,K., Ovchinnikova,G., Pati,A., Chen,A., Palaniappan,K.,
Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Spring,S., Rohde,M.,
Göker,M., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome sequence of
Aminobacterium colombiense type strain (ALA-1). Standards in Genomic
Sciences 2, 280-289.
• Clum,A., Tindall,B.J., Sikorski,J., Ivanova,N., Mavromatis,K.,
Lucas,S., Glavina Del,R.T., Nolan,M., Chen,F., Tice,H., Pitluck,S.,
Cheng,J.F., Chertkov,O., Brettin,T., Han,C., Detter,J.C., Kuske,C.,
Bruce,D., Goodwin,L., Ovchinikova,G., Pati,A., Mikhailova,N.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Chain,P., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Lapidus,A. (2010) Erratum
to: Complete genome sequence of Pirellula staleyi type strain
(ATCC 27377). Standards in Genomic Sciences 2, 228-288.
• Copeland,A., Sikorski,J., Lapidus,A., Nolan,M., Glavina,T., Del,R.,
Lucas,S., Chen,F., Tice,H., Pitluck,S., Cheng,J.F., Pukall,R.,
Chertkov,O., Brettin,T., Han,C., Kuske,C., Bruce,D., Goodwin,L.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Chen,A., Palaniappan,K.,
Chain,P., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Detter,J.C. (2010) Erratum
to: Complete genome sequence of Atopobium parvulum type strain
(IPP 1246). Standards in Genomic Sciences 2, 361-362.
• Del Rio,T.G., Chertkov,O., Yasawong,M., Lucas,S., Deshpande,S.,
Cheng,J.F., Detter,C., Tapia,R., Han,C., Goodwin,L., Pitluck,S.,
Liolios,K., Ivanova,N., Mavromatis,K., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Pukall,R., Sikorski,J., Göker,M., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P.,
& Lapidus,A. (2010) Complete genome sequence of Intrasporangium
calvum type strain (7 KIP). Standards in Genomic Sciences 3, 294-303.
• Dietrich,N., Rohde,M., Geffers,R., Kroger,A., Hauser,H., Weiss,S., &
Gekara,N.O. (2010) Mast cells elicit proinflammatory but not type I
interferon responses upon activation of TLRs by bacteria. Proceeding
of the National Academy of Sciences of the United States of America
107, 8748-8753.
• Djao,O.D., Zhang,X., Lucas,S., Lapidus,A., Del Rio,T.G., Nolan,M.,
Tice,H., Cheng,J.F., Han,C., Tapia,R., Goodwin,L., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Ovchinnikova,G.,
Pati,A., Brambilla,E., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Rohde,M., Sikorski,J., Spring,S., Göker,M.,
Detter,J.C., Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Syntrophothermus lipocalidus type strain (TGB-C1).
Standards in Genomic Sciences 3, 268-275.
• Foster,B., Pukall,R., Abt,B., Nolan,M., Glavina Del,R.T., Chen,F.,
Lucas,S., Tice,H., Pitluck,S., Cheng,J.F., Chertkov,O., Brettin,T.,
Han,C., Detter,J.C., Bruce,D., Goodwin,L., Ivanova,N., Mavromatis,K.,
Pati,A., Mikhailova,N., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Chain,P., Rohde,M., Goker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., &
Lapidus,A. (2010) Complete genome sequence of Xylanimonas cellulosilytica
type strain (XIL07). Standards in Genomic Sciences 2, 1-8.
• Garbe,J., Wesche,A., Bunk,B., Kazmierczak,M., Selezska,K., Rohde,C.,
Sikorski,J., Rohde,M., Jahn,D., & Schobert,M. (2010) Characterization
of JG024, a Pseudomonas aeruginosa PB1-like broad host range
phage under simulated infection conditions. BMC Microbiology 10 ,301.
• Glavina Del,R.T., Abt,B., Spring,S., Lapidus,A., Nolan,M., Tice,H.,
Copeland,A., Cheng,J.F., Chen,F., Bruce,D., Goodwin,L., Pitluck,S.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Chain,P.,
Saunders,E., Detter,J.C., Brettin,T., Rohde,M., Göker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., &
Lucas,S. (2010) Complete genome sequence of Chitinophaga pinensis
type strain (UQM 2034). Standards in Genomic Sciences 2, 87-95.
• Göker,M., Held,B., Lapidus,A., Nolan,M., Spring,S., Yasawong,M.,
Lucas,S., Glavina Del,R.T., Tice,H., Cheng,J.F., Goodwin,L., Tapia,R.,
Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Pati,A., Chen,A., Palaniappan,K., Brambilla,E., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Brettin,T., Detter,J.C., Han,C., Rohde,M.,
Sikorski,J., Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Ignisphaera aggregans type strain (AQ1.S1). Standards in
Genomic Sciences 3, 66-75.
• Göker,M., Held,B., Lucas,S., Nolan,M., Yasawong,M., Glavina Del,R.T.,
Tice,H., Cheng,J.F., Bruce,D., Detter,J.C., Tapia,R., Han,C., Goodwin,L.,
Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Pati,A., Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J.,
Jeffries,C.D., Rohde,M., Sikorski,J., Pukall,R., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., &
Lapidus,A. (2010) Complete genome sequence of Olsenella uli type
strain (VPI D76D-27C). Standards in Genomic Sciences 3, 76-84.
• Gronow,S., Welnitz,S., Lapidus,A., Nolan,M., Ivanova,N., Glavina
Del,R.T., Copeland,A., Chen,F., Tice,H., Pitluck,S., Cheng,J.F.,
Saunders,E., Brettin,T., Han,C., Detter,J.C., Bruce,D., Goodwin,L.,
Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Pati,A., Mavromatis,K.,
Mikhailova,N., Chen,A., Palaniappan,K., Chain,P., Rohde,M.,
Göker,M., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., Klenk,H.P., & Lucas,S. (2010) Complete genome sequence
of Veillonella parvula type strain (Te3). Standards in Genomic
Sciences 2, 57-65.
• Han,C., Gu,W., Zhang,X., Lapidus,A., Nolan,M., Copeland,A., Lucas,S.,
Del Rio,T.G., Tice,H., Cheng,J.F., Tapia,R., Goodwin,L., Pitluck,S.,
Pagani,I., Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Schneider,S., Rohde,M., Göker,M., Pukall,R., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P.,
& Detter,J.C. (2010) Complete genome sequence of Thermaerobacter
marianensis type strain (7p75a). Standards in Genomic Sciences 3,
337-345.
• Hänisch,J., Ehinger,J., Ladwein,M., Rohde,M., Derivery,E., Bosse,T.,
Steffen,A., Bumann,D., Misselwitz,B., Hardt,W.D., Gautreau,A.,
Stradal,T.B.*., & Rottner,K. (2010) Molecular dissection of Salmonella-induced
membrane ruffling versus invasion. Cellular Microbiology
12, 84-98.
• Hoffmann,C., Berking,A., Agerer,F., Buntru,A., Neske,F.,
Chhatwal,G.S*., Ohlsen,K., & Hauck,C.R. (2010) Caveolin limits
membrane microdomain mobility and integrin-mediated uptake of
fibronectin-binding pathogens. Journal of Cell Science 123, 4280-4291.
• Hu,Y., van der,G.R., Besra,G.S., Gurcha,S.S., Liu,A., Rohde,M.,
Singh,M., & Coates,A. (2010) 3-Ketosteroid 9alpha-hydroxylase is an
essential factor in the pathogenesis of Mycobacterium tuberculosis.
Molecular Microbiology 75, 107-121.
• Itzek,A., Gillen,C.M., Fulde,M., Friedrichs,C., Rodloff,A.C.,
Chhatwal,G.S*., & Nitsche-Schmitz,D.P. (2010) Contribution of plasminogen
activation towards the pathogenic potential of oral streptococci.
PLoS ONE 5, e13826.
• Ivanova,N., Sikorski,J., Jando,M., Munk,C., Lapidus,A., Glavina
Del,R.T., Copeland,A., Tice,H., Cheng,J.F., Lucas,S., Chen,F., Nolan,M.,
Bruce,D., Goodwin,L., Pitluck,S., Mavromatis,K., Mikhailova,N.,
Pati,A., Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J.,
Jeffries,C.D., Meincke,L., Brettin,T., Detter,J.C., Rohde,M., Göker,M.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., &
Klenk,H.P. (2010) Complete genome sequence of Geodermatophilus
obscurus type strain (G-20). Standards in Genomic Sciences 2, 158-167.
• Ivanova,N., Sikorski,J., Jando,M., Lapidus,A., Nolan,M., Lucas,S.,
Del Rio,T.G., Tice,H., Copeland,A., Cheng,J.F., Chen,F., Bruce,D.,
Goodwin,L., Pitluck,S., Mavromatis,K., Ovchinnikova,G., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Chain,P., Saunders,E., Han,C., Detter,J.C., Brettin,T., Rohde,M.,
Göker,M., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Klenk,H.P., & Kyrpides,N.C. (2010) Complete genome sequence
of Gordonia bronchialis type strain (3410). Standards in Genomic
Sciences 2, 19-28.

156 SCIENTIFIC REPORTS | Publications
• Ivanova,N., Daum,C., Lang,E., Abt,B., Kopitz,M., Saunders,E.,
Lapidus,A., Lucas,S., Glavina Del,R.T., Nolan,M., Tice,H., Copeland,A.,
Cheng,J.F., Chen,F., Bruce,D., Goodwin,L., Pitluck,S., Mavromatis,K.,
Pati,A., Mikhailova,N., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Detter,J.C., Brettin,T., Rohde,M., Göker,M.,
Bristow,J., Markowitz,V., Eisen,J.A., Hugenholtz,P., Kyrpides,N.C., &
Klenk,H.P. (2010) Complete genome sequence of Haliangium ochraceum
type strain (SMP-2). Standards in Genomic Sciences 2, 96-106.
• Jensch,I., Gamez,G., Rothe,M., Ebert,S., Fulde,M., Somplatzki,D.,
Bergmann,S., Petruschka,L., Rohde,M., Nau,R., & Hammerschmidt,S.
(2010) PavB is a surface-exposed adhesin of Streptococcus pneumoniae
contributing to nasopharyngeal colonization and airways infections.
Molecular Microbiology 77, 22-43.
• Kaur,S.J., Nerlich,A., Bergmann,S., Rohde,M., Fulde,M., Zahner,D.,
Hanski,E., Zinkernagel,A., Nizet,V., Chhatwal,G.S.*., & Talay,S.R.
(2010) The CXC chemokine-degrading protease SpyCep of Streptococcus
pyogenes promotes its uptake into endothelial cells. Journal of
Biological Chemistry 285, 27798-27805.
• Kiss,H., Lang,E., Lapidus,A., Copeland,A., Nolan,M., Glavina
Del,R.T., Chen,F., Lucas,S., Tice,H., Cheng,J.F., Han,C., Goodwin,L.,
Pitluck,S., Liolios,K., Pati,A., Ivanova,N., Mavromatis,K., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Detter,J.C., Brettin,T., Spring,S., Rohde,M., Göker,M., Woyke,T.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C.,
& Klenk,H.P. (2010) Complete genome sequence of Denitrovibrio
acetiphilus type strain (N2460). Standards in Genomic Sciences 2,
270-279.
• Kotz,A., Wagener,J., Engel,J., Routier,F., Echtenacher,B., Pich,A.,
Rohde,M., Hoffmann,P., Heesemann,J., & Ebel,F. (2010) The mitA
gene of Aspergillus fumigatus is required for mannosylation of inositol-phosphorylceramide,
but is dispensable for pathogenicity. Fungal
Genetics and Biology 47, 169-178.
• Labutti,K., Sikorski,J., Schneider,S., Nolan,M., Lucas,S., Glavina
Del,R.T., Tice,H., Cheng,J.F., Goodwin,L., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Tindall,B.J., Rohde,M., Göker,M., Woyke,T., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Lapidus,A.
(2010) Complete genome sequence of Planctomyces limnophilus type
strain (Mu 290). Standards in Genomic Sciences 3, 47-56.
• Labutti,K., Mayilraj,S., Clum,A., Lucas,S., Glavina Del,R.T., Nolan,M.,
Tice,H., Cheng,J.F., Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K.,
Mikhailova,N., Pati,A., Goodwin,L., Chen,A., Palaniappan,K., Land,M.,
Hauser,L., Chang,Y.J., Jeffries,C.D., Rohde,M., Spring,S., Göker,M.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., Klenk,H.P., & Lapidus,A. (2010) Permanent draft genome
sequence of Dethiosulfovibrio peptidovorans type strain (SEBR
4207). Standards in Genomic Sciences 3, 85-92.
• Lail,K., Sikorski,J., Saunders,E., Lapidus,A., Glavina Del,R.T.,
Copeland,A., Tice,H., Cheng,J.F., Lucas,S., Nolan,M., Bruce,D.,
Goodwin,L., Pitluck,S., Ivanova,N., Mavromatis,K., Ovchinnikova,G.,
Pati,A., Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J.,
Jeffries,C.D., Chain,P., Brettin,T., Detter,J.C., Schutze,A., Rohde,M.,
Tindall,B.J., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Chen,F. (2010) Complete
genome sequence of Spirosoma linguale type strain (1). Standards in
Genomic Sciences 2, 176-185.
• Liolios,K., Sikorski,J., Jando,M., Lapidus,A., Copeland,A., Glavina,T.,
Del,R., Nolan,M., Lucas,S., Tice,H., Cheng,J.F., Han,C., Woyke,T.,
Goodwin,L., Pitluck,S., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Chertkov,O., Kuske,C., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Detter,J.C., Brettin,T., Rohde,M., Göker,M.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Klenk,H.P., &
Kyrpides,N.C. (2010) Complete genome sequence of Thermobispora
bispora type strain (R51). Standards in Genomic Sciences 2, 318-326.
• Maamary,P.G., Sanderson-Smith,M.L., Aziz,R.K., Hollands,A.,
Cole,J.N., McKay,F.C., Mcarthur,J.D., Kirk,J.K., Cork,A.J., Keefe,R.J.,
Kansal,R.G., Sun,H., Taylor,W.L., Chhatwal,G.S*., Ginsburg,D.,
Nizet,V., Kotb,M., & Walker,M.J. (2010) Parameters governing invasive
disease propensity of non-M1 serotype group A streptococci.
Journal of Innate Immunology 2, 596-606.
• Macke,L., Garritsen,H.S., Meyring,W., Hannig,H., Pagelow,U.,
Wormann,B., Piechaczek,C., Geffers,R., Rohde,M., Lindenmaier,W.,
& Dittmar,K.E.*. (2010) Evaluating maturation and genetic modification
of human dendritic cells in a new polyolefin cell culture bag
system. Transfusion 50, 843-855.
• Marjenberg,Z.R., Ellis,I.R., Hagan,R.M., Prabhakaran,S., Hook,M.,
Talay,S.R.*., Potts,J.R., Staunton,D., & Schwarz-Linek,U. (2010) Cooperative
binding and activation of fibronectin by a bacterial surface
protein. Journal of Biological Chemistry 286, 1884-1894.
• Mavromatis,K., Sikorski,J., Pabst,E., Teshima,H., Lapidus,A., Lucas,S.,
Nolan,M., Glavina Del,R.T., Cheng,J.F., Bruce,D., Goodwin,L.,
Pitluck,S., Liolios,K., Ivanova,N., Mikhailova,N., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Spring,S., Göker,M., Wirth,R., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C.
(2010) Complete genome sequence of Vulcanisaeta distributa type
strain (IC-017). Standards in Genomic Sciences 3, 117-125.
• Mavromatis,K., Abt,B., Brambilla,E., Lapidus,A., Copeland,A.,
Deshpande,S., Nolan,M., Lucas,S., Tice,H., Cheng,J.F., Han,C.,
Detter,J.C., Woyke,T., Goodwin,L., Pitluck,S., Held,B., Brettin,T.,
Tapia,R., Ivanova,N., Mikhailova,N., Pati,A., Liolios,K., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C. (2010) Complete genome
sequence of Coraliomargarita akajimensis type strain (04OKA010-
24). Standards in Genomic Sciences 2, 290-299.
• Mavromatis,K., Sikorski,J., Lapidus,A., Glavina Del,R.T., Copeland,A.,
Tice,H., Cheng,J.F., Lucas,S., Chen,F., Nolan,M., Bruce,D.,
Goodwin,L., Pitluck,S., Ivanova,N., Ovchinnikova,G., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Chain,P.,
Meincke,L., Sims,D., Chertkov,O., Han,C., Brettin,T., Detter,J.C.,
Wahrenburg,C., Rohde,M., Pukall,R., Göker,M., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C. (2010) Complete
genome sequence of Alicyclobacillus acidocaldarius type strain
(104-IA). Standards in Genomic Sciences 2, 9-18.
• Mavromatis,K., Yasawong,M., Chertkov,O., Lapidus,A., Lucas,S.,
Nolan,M., Del Rio,T.G., Tice,H., Cheng,J.F., Pitluck,S., Liolios,K.,
Ivanova,N., Tapia,R., Han,C., Bruce,D., Goodwin,L., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Detter,J.C., Rohde,M., Brambilla,E., Spring,S., Göker,M., Sikorski,J.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Klenk,H.P., & Kyrpides,N.C. (2010) Complete genome sequence of Spirochaeta
smaragdinae type strain (SEBR 4228). Standards in Genomic
Sciences 3, 136-144.
• Moeller,R., Rohde,M., & Reitz,G. (2010) Effects of ionizing radioation
on the survival of bacterial spores in artificial martian regolith.
Icarus - International Journal of Solar System Studies 206, 783-786.
• Nolan,M., Sikorski,J., Davenport,K., Lucas,S., Del Rio,T.G.,
Tice,H., Cheng,J.F., Goodwin,L., Pitluck,S., Liolios,K., Ivanova,N.,
Mavromatis,K., Ovchinnikova,G., Pati,A., Chen,A., Palaniappan,K.,
Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Tapia,R., Brettin,T.,
Detter,J.C., Han,C., Yasawong,M., Rohde,M., Tindall,B.J., Göker,M.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., Klenk,H.P., & Lapidus,A. (2010) Complete genome
sequence of Ferrimonas balearica type strain (PAT). Standards in
Genomic Sciences 3, 174-182.
• Nolan,M., Sikorski,J., Jando,M., Lucas,S., Lapidus,A., Glavina Del,R.T.,
Chen,F., Tice,H., Pitluck,S., Cheng,J.F., Chertkov,O., Sims,D., Meincke,L.,
Brettin,T., Han,C., Detter,J.C., Bruce,D., Goodwin,L., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Chen,A., Palaniappan,K., Chain,P., Rohde,M., Göker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P.
(2010) Complete genome sequence of Streptosporangium roseum type
strain (NI 9100). Standard in Genomic Sciences 2, 29-37.
• Pati,A., Gronow,S., Lapidus,A., Copeland,A., Glavina Del,R.T.,
Nolan,M., Lucas,S., Tice,H., Cheng,J.F., Han,C., Chertkov,O.,
Bruce,D., Tapia,R., Goodwin,L., Pitluck,S., Liolios,K., Ivanova,N.,
Mavromatis,K., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Detter,J.C., Rohde,M., Göker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C.
(2010) Complete genome sequence of Arcobacter nitrofigilis type
strain (CI). Standards in Genomic Sciences 2, 300-308.

SCIENTIFIC REPORTS | Publications
157
• Pati,A., Sikorski,J., Gronow,S., Munk,C., Lapidus,A., Copeland,A.,
Glavina Del,T.T., Nolan,M., Lucas,S., Chen,F., Tice,H., Cheng,J.F.,
Han,C., Detter,J.C., Bruce,D., Tapia,R., Goodwin,L., Pitluck,S.,
Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Spring,S., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Brachyspira murdochii type strain (56-150). Standards in
Genomic Sciences 2, 260-269.
• Pati,A., Labutti,K., Pukall,R., Nolan,M., Glavina Del,R.T., Tice,H.,
Cheng,J.F., Lucas,S., Chen,F., Copeland,A., Ivanova,N., Mavromatis,K.,
Mikhailova,N., Pitluck,S., Bruce,D., Goodwin,L., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Chen,A., Palaniappan,K., Chain,P., Brettin,T.,
Sikorski,J., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Lapidus,A. (2010) Complete
genome sequence of Sphaerobacter thermophilus type strain (S
6022). Standards in Genomic Sciences 2, 49-56.
• Pitluck,S., Yasawong,M., Munk,C., Nolan,M., Lapidus,A., Lucas,S.,
Glavina Del,R.T., Tice,H., Cheng,J.F., Bruce,D., Detter,C., Tapia,R.,
Han,C., Goodwin,L., Liolios,K., Ivanova,N., Mavromatis,K.,
Mikhailova,N., Pati,A., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Rohde,M., Spring,S., Sikorski,J., Göker,M.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome sequence of
Thermosediminibacter oceani type strain (JW/IW-1228P). Standards
in Genomic Sciences 3, 108-116.
• Pitluck,S., Yasawong,M., Held,B., Lapidus,A., Nolan,M.,
Copeland,A., Lucas,S., Del Rio,T.G., Tice,H., Cheng,J.F., Chertkov,O.,
Goodwin,L., Tapia,R., Han,C., Liolios,K., Ivanova,N., Mavromatis,K.,
Ovchinnikova,G., Pati,A., Chen,A., Palaniappan,K., Land,M.,
Hauser,L., Chang,Y.J., Jeffries,C.D., Pukall,R., Spring,S., Rohde,M.,
Sikorski,J., Göker,M., Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Non-contiguous
finished genome sequence of Aminomonas paucivorans type strain
(GLU-3). Standards in Genomic Sciences 3, 285-293.
• Pukall,R., Lapidus,A., Glavina Del,R.T., Copeland,A., Tice,H.,
Cheng,J.F., Lucas,S., Chen,F., Nolan,M., Labutti,K., Pati,A., Ivanova,N.,
Mavromatis,K., Mikhailova,N., Pitluck,S., Bruce,D., Goodwin,L.,
Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D., Chen,A., Palaniappan,K.,
Chain,P., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Brettin,T. (2010) Complete
genome sequence of Kribbella flavida type strain (IFO 14399).
Standards in Genomic Sciences 2, 186-193.
• Pukall,R., Lapidus,A., Glavina Del,R.T., Copeland,A., Tice,H.,
Cheng,J.F., Lucas,S., Chen,F., Nolan,M., Bruce,D., Goodwin,L.,
Pitluck,S., Mavromatis,K., Ivanova,N., Ovchinnikova,G., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Chain,P., Meincke,L., Sims,D., Brettin,T., Detter,J.C., Rohde,M.,
Göker,M., Bristow,J., Eisen,J.A., Markowitz,V., Kyrpides,N.C.,
Klenk,H.P., & Hugenholtz,P. (2010) Complete genome sequence of
Conexibacter woesei type strain (ID131577). Standards in Genomic
Sciences 2, 212-219.
• Reißmann,S., Friedrichs,C., Rajkumari,R., Itzek,A., Fulde,M.,
Rodloff,A.C., Brahmadathan,K.N., Chhatwal,G.S.*., & Nitsche-
Schmitz,D.P. (2010) Contribution of Streptococcus anginosus to
infections caused by groups C and G Streptococci, Southern India.
Emerging Infectious Diseases 16, 656-663.
• Rohde,M., Graham,R.M., Branitzki-Heinemann,K., Borchers,P.,
Preuss,C., Schleicher,I., Zahner,D., Talay,S.R., Fulde,M., Dinkla,K., &
Chhatwal,G.S.*. (2010) Differences in the aromatic domain of homologous
streptococcal fibronectin-binding proteins trigger different
cell invasion mechanisms and survival rates. Cellular Microbiology
13, 450-468.
• Saunders,E., Tindall,B.J., Fahnrich,R., Lapidus,A., Copeland,A., Del
Rio,T.G., Lucas,S., Chen,F., Tice,H., Cheng,J.F., Han,C., Detter,J.C.,
Bruce,D., Goodwin,L., Chain,P., Pitluck,S., Pati,A., Ivanova,N.,
Mavromatis,K., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Brettin,T., Rohde,M., Göker,M., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Klenk,H.P., & Kyrpides,N.C.
(2010) Complete genome sequence of Haloterrigena turkmenica type
strain (4k). Standards in Genomic Sciences 2, 107-116.
• Sikorski,J., Chertkov,O., Lapidus,A., Nolan,M., Lucas,S., Del Rio,T.G.,
Tice,H., Cheng,J.F., Tapia,R., Han,C., Goodwin,L., Pitluck,S.,
Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Brambilla,E., Yasawong,M., Rohde,M., Pukall,R., Spring,S., Göker,M.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome sequence of
Ilyobacter polytropus type strain (CuHbu1). Standards in Genomic
Sciences 3, 304-314.
• Sikorski,J., Lapidus,A., Chertkov,O., Lucas,S., Copeland,A., Glavina
Del,R.T., Nolan,M., Tice,H., Cheng,J.F., Han,C., Brambilla,E.,
Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K., Mikhailova,N.,
Pati,A., Bruce,D., Detter,C., Tapia,R., Goodwin,L., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Göker,M., Spring,S., Woyke,T., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010)
Complete genome sequence of Acetohalobium arabaticum type strain
(Z-7288). Standards in Genomic Sciences 3, 57-65.
• Sikorski,J., Lapidus,A., Copeland,A., Glavina Del,R.T., Nolan,M.,
Lucas,S., Chen,F., Tice,H., Cheng,J.F., Saunders,E., Bruce,D.,
Goodwin,L., Pitluck,S., Ovchinnikova,G., Pati,A., Ivanova,N.,
Mavromatis,K., Chen,A., Palaniappan,K., Chain,P., Land,M.,
Hauser,L., Chang,Y.J., Jeffries,C.D., Brettin,T., Detter,J.C., Han,C.,
Rohde,M., Lang,E., Spring,S., Göker,M., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete
genome sequence of Sulfurospirillum deleyianum type strain
(5175). Standards in Genomic Sciences 2, 149-157.
• Sikorski,J., Munk,C., Lapidus,A., Ngatchou Djao,O.D., Lucas,S.,
Glavina Del,R.T., Nolan,M., Tice,H., Han,C., Cheng,J.F., Tapia,R.,
Goodwin,L., Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K.,
Mikhailova,N., Pati,A., Sims,D., Meincke,L., Brettin,T., Detter,J.C.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Lang,E., Spring,S., Göker,M., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P.
(2010) Complete genome sequence of Sulfurimonas autotrophica type
strain (OK10). Standards in Genomic Sciences 3, 194-202.
• Sikorski,J., Tindall,B.J., Lowry,S., Lucas,S., Nolan,M., Copeland,A.,
Glavina Del,R.T., Tice,H., Cheng,J.F., Han,C., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A., Goodwin,L.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Rohde,M., Göker,M., Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Lapidus,A. (2010) Complete
genome sequence of Meiothermus silvanus type strain (VI-R2).
Standards in Genomic Sciences 3, 37-46.
• Sikorski,J., Lapidus,A., Copeland,A., Misra,M., Glavina Del,R.T.,
Nolan,M., Lucas,S., Chen,F., Tice,H., Cheng,J.F., Jando,M.,
Schneider,S., Bruce,D., Goodwin,L., Pitluck,S., Liolios,K.,
Mikhailova,N., Pati,A., Ivanova,N., Mavromatis,K., Chen,A.,
Palaniappan,K., Chertkov,O., Land,M., Hauser,L., Chang,Y.J.,
Jeffries,C.D., Brettin,T., Detter,J.C., Han,C., Rohde,M., Göker,M.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., &
Klenk,H.P. (2010) Complete genome sequence of Segniliparus rotundus
type strain (CDC 1076). Standards in Genomic Sciences 2, 203-211.
• Spring,S., Nolan,M., Lapidus,A., Glavina Del,R.T., Copeland,A.,
Tice,H., Cheng,J.F., Lucas,S., Land,M., Chen,F., Bruce,D., Goodwin,L.,
Pitluck,S., Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A.,
Chen,A., Palaniappan,K., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Munk,C., Kiss,H., Chain,P., Han,C., Brettin,T., Detter,J.C.,
Schuler,E., Göker,M., Rohde,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Desulfohalobium retbaense type strain (HR(100)). Standards
in Genomic Siences 2, 38-48.
• Spring,S., Scheuner,C., Lapidus,A., Lucas,S., Glavina Del,R.T.,
Tice,H., Copeland,A., Cheng,J.F., Chen,F., Nolan,M., Saunders,E.,
Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K., Lykidis,A., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Goodwin,L., Detter,J.C., Brettin,T., Rohde,M., Göker,M., Woyke,T.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C.,
& Klenk,H.P. (2010) The Genome Sequence of Methanohalophilus
mahii SLP Reveals Differences in the Energy Metabolism among
Members of the Methanosarcinaceae Inhabiting Freshwater and
Saline Environments. Archaea 2010, 690737.

158 SCIENTIFIC REPORTS | Publications
• Sun,H., Spring,S., Lapidus,A., Davenport,K., Del Rio,T.G., Tice,H.,
Nolan,M., Copeland,A., Cheng,J.F., Lucas,S., Tapia,R., Goodwin,L.,
Pitluck,S., Ivanova,N., Pagani,I., Mavromatis,K., Ovchinnikova,G.,
Pati,A., Chen,A., Palaniappan,K., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Detter,J.C., Han,C., Rohde,M., Brambilla,E., Göker,M., Woyke,T.,
Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C.,
Klenk,H.P., & Land,M. (2010) Complete genome sequence of Desulfarculus
baarsii type strain (2st14). Standards in Genomic Sciences 3,
276-284.
• Sun,H., Lapidus,A., Nolan,M., Lucas,S., Del Rio,T.G., Tice,H.,
Cheng,J.F., Tapia,R., Han,C., Goodwin,L., Pitluck,S., Pagani,I.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Pati,A., Chen,A.,
Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Djao,O.D., Rohde,M., Sikorski,J., Göker,M., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P.
(2010) Complete genome sequence of Nocardiopsis dassonvillei type
strain (IMRU 509). Standards in Genomic Sciences 3, 325-336.
• Tegtmeyer,N., Hartig,R., Delahay,R.M., Rohde,M., Brandt,S.,
Conradi,J., Takahashi,S., Smolka,A.J., Sewald,N., & Backert,S. (2010)
A small fibronectin-mimicking protein from bacteria induces cell
spreading and focal adhesion formation. Journal of Biological
Chemistry 285, 23515-23526.
• Tice,H., Mayilraj,S., Sims,D., Lapidus,A., Nolan,M., Lucas,S., Glavina
Del,R.T., Copeland,A., Cheng,J.F., Meincke,L., Bruce,D., Goodwin,L.,
Pitluck,S., Ivanova,N., Mavromatis,K., Ovchinnikova,G., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Detter,J.C., Brettin,T., Rohde,M., Goker,M., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Chen,F.
(2010) Complete genome sequence of Nakamurella multipartita type
strain (Y-104). Standards in Genomic Sciences 2, 168-175.
• Tindall,B.J., Sikorski,J., Lucas,S., Goltsman,E., Copeland,A., Glavina
Del,R.T., Nolan,M., Tice,H., Cheng,J.F., Han,C., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Ovchinnikova,G., Pati,A., Fahnrich,R.,
Goodwin,L., Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J.,
Jeffries,C.D., Rohde,M., Göker,M., Woyke,T., Bristow,J., Eisen,J.A.,
Markowitz,V., Hugenholtz,P., Kyrpides,N.C., Klenk,H.P., & Lapidus,A.
(2010) Complete genome sequence of Meiothermus ruber type strain
(21). Standards in Genomic Sciences 3, 26-36.
• Traverso,F.R., Bohr,U.R.M., Oyarzabal,O.A., Rohde,M., Clarici,A.,
Wex,T., Kuester,D., Malfertheiner,P., Fox,J.G., & Backert,S. (2010) Morphologic,
genetic, and biochemical characterization of Helicobacter
magdeburgensis, a novel species isolated from the intestine of laboratory
mice. Helicobacter 15, 403-415.
• von Jan,M., Lapidus,A., Del Rio,T.G., Copeland,A., Tice,H., Cheng,J.F.,
Lucas,S., Chen,F., Nolan,M., Goodwin,L., Han,C., Pitluck,S., Liolios,K.,
Ivanova,N., Mavromatis,K., Ovchinnikova,G., Chertkov,O., Pati,A.,
Chen,A., Palaniappan,K., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Saunders,E., Brettin,T., Detter,J.C., Chain,P., Eichinger,K., Huber,H.,
Spring,S., Rohde,M., Göker,M., Wirth,R., Woyke,T., Bristow,J.,
Eisen,J.A., Markowitz,V., Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P.
(2010) Complete genome sequence of Archaeoglobus profundus type
strain (AV18). Standards in Genomic Sciences 2, 327-346.
• Wirth,R., Sikorski,J., Brambilla,E., Misra,M., Lapidus,A., Copeland,A.,
Nolan,M., Lucas,S., Chen,F., Tice,H., Cheng,J.F., Han,C., Detter,J.C.,
Tapia,R., Bruce,D., Goodwin,L., Pitluck,S., Pati,A., Anderson,I.,
Ivanova,N., Mavromatis,K., Mikhailova,N., Chen,A., Palaniappan,K.,
Bilek,Y., Hader,T., Land,M., Hauser,L., Chang,Y.J., Jeffries,C.D.,
Tindall,B.J., Rohde,M., Göker,M., Bristow,J., Eisen,J.A., Markowitz,V.,
Hugenholtz,P., Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome
sequence of Thermocrinis albus type strain (HI 11/12). Standards in
Genomic Sciences 2, 194-202.
• Yasawong,M., Teshima,H., Lapidus,A., Nolan,M., Lucas,S., Glavina
Del,R.T., Tice,H., Cheng,J.F., Bruce,D., Detter,C., Tapia,R., Han,C.,
Goodwin,L., Pitluck,S., Liolios,K., Ivanova,N., Mavromatis,K.,
Mikhailova,N., Pati,A., Chen,A., Palaniappan,K., Land,M., Hauser,L.,
Chang,Y.J., Jeffries,C.D., Rohde,M., Sikorski,J., Pukall,R., Göker,M.,
Woyke,T., Bristow,J., Eisen,J.A., Markowitz,V., Hugenholtz,P.,
Kyrpides,N.C., & Klenk,H.P. (2010) Complete genome sequence of
Arcanobacterium haemolyticum type strain (11018). Standards in
Genomic Sciences 3, 126-135.
• Bergmann,R., Dinkla,K., Nitsche-Schmitz,D.P., Graham,R.M.,
Luttge,M., Sanderson-Smith,M.L., Nerlich,A., Rohde,M., &
Chhatwal,G.S.*. (2011) Biological functions of GCS3, a novel plasminogen-binding
protein of Streptococcus dysgalactiae ssp. equisimilis.
International Journal of Medical Microbiology 301, 157-164.
• Fulde,M., Rohde,M., Hitzmann,A., Preissner,K.T., Nitsche-
Schmitz,D.P., Nerlich,A., Chhatwal,G.S.*., & Bergmann,S. (2011)
SCM, a novel M-like protein from Streptococcus canis, binds (mini)-
plasminogen with high affinity and facilitates bacterial transmigration.
Biochemical Journal 434, 523-535.
• Härtel,T., Klein,M., Koedel,U., Rohde,M., Petruschka,L., &
Hammerschmidt,S. (2011) Impact of glutamine transporters on pneumococcal
fitness under infection-related conditions. Infection and
Immunity 79, 44-58.
• Lyszkiewicz,M., Zietara,N., Rohde,M., Gekara,N.O., Jablonska,J.,
Dittmar,K.E.*., & Weiss,S. (2011) SIGN-R1+MHC II+ cells of the
splenic marginal zone--a novel type of resident dendritic cells.
Journal of Leukocyte Biology 89, 607-615.
• Gupta,A.K., Dharne,M.S., Rangrez,A.Y., Verma,P., Ghate,H.V.,
Rohde,M., Patole,M.S., & Shouche,Y.S. (2010) Ignatzschineria indica
sp. nov. and Ignatzschineria ureaclastica sp. nov., isolated from adult
flesh fly (Diptera: Sarcophagidae). International Journal of Systematic
and Evolutionary Microbiology, in press.
• von Jan,M., Riegger,N., Potter,G., Schumann,P., Verbarg,S., Sproer,C.,
Rohde,M., Lauer,B., Labeda,D.P., & Klenk,H.P. (2010) Kroppenstedtia
eburnea gen. nov., sp. nov., a novel thermoactinomycete isolated by
environmental screening, and emended description of the family
Thermoactinomycetaceae Matsuo et al. 2006 emend. Yassin et al.
2009. International Journal of Systematic and Evolutionary Microbiology,
in press.
• Abel,J., Goldmann,O., Ziegler,C., Höltje,C., Smeltzer,M.S.,
Cheung,A.L., Bruhn,D., Rohde,M., & Medina,E. (2011) Staphylococcus
aureus evades the extracellular antimicrobial activity of mast cells
by promoting its own uptake. Journal of Innate Immunity, in press.
• Hebecker,S., Arendt,W., Heinemann,I.U., Tiefenau,J.H., Nimtz,M.,
Rohde,M., Soll,D., & Moser,J. (2011) Alanyl-phosphatidylglycerol synthase:
mechanism of substrate recognition during tRNA-dependent
lipid modification in Pseudomonas aeruginosa. Molecular Microbiology,
in press.
• Willenborg,J., Fulde,M., de,G.A., Rohde,M., Smith,H.E., Valentin-
Weigand,P., & Goethe,R. (2011) Role of glucose and CcpA in capsule
expression and virulence of Streptococcus suis. Microbiology, in press.
RG Infection Immunology | Priv.-Doz. Dr. Eva Medina
• Goldmann,O., Lehne,S., & Medina,E. (2010) Age-related susceptibility
to Streptococcus pyogenes infection in mice: underlying immune
dysfunction and strategy to enhance immunity. Journal of Pathology
220, 521-529.
• Goldmann,O., Hertzen,E., Hecht,A., Schmidt,H., Lehne,S., Norrby-
Teglund,A., & Medina,E. (2010) Inducible cyclooxygenase released
prostaglandin E2 modulates the severity of infection caused by
Streptococcus pyogenes. Journal of Immunology 185, 2372-2381.
• Loof,T.G., Goldmann,O., Gessner,A., Herwald,H., & Medina,E. (2010)
Aberrant inflammatory response to Streptococcus pyogenes in mice
lacking myeloid differentiation factor 88. American Journal of Pathology
176, 754-763.
• Medina,E. & Goldmann,O. (2011) In vivo and ex vivo protocols for
measuring the killing of extracellular pathogens by macrophages.
Current Protocols in Immunology Chapter 14, Unit14.19.
• Medina,E. (2010) Murine model of cutaneous infection with Streptococcus
pyogenes. Methods in Molecular Biology 602, 395-403.
• Medina,E. (2010) Murine model of pneumococcal pneumonia. Methods
in Molecular Biology 602, 405-410.

SCIENTIFIC REPORTS | Publications
159
• Medina,E. (2010) Murine model of polymicrobial septic peritonitis
using cecal ligation and puncture (CLP). Methods in Molecular Biology
602, 411-415.
• Nippe,N., Varga,G., Holzinger,D., Loffler,B., Medina,E., Becker,K.,
Roth,J., Ehrchen,J.M., & Sunderkotter,C. (2010) Subcutaneous infection
with S. aureus in mice reveals association of resistance with
influx of neutrophils and Th2 response. Journal of Investigative Dermatology
131, 125-132.
• Tuchscherr,L., Medina,E., Hussain,M., Volker,W., Heitmann,V.,
Niemann,S., Holzinger,D., Roth,J., Proctor,R.A., Becker,K., Peters,G.,
& Loffler,B. (2011) Staphylococcus aureus phenotype switching: an
effective bacterial strategy to escape host immune response and
establish a chronic infection. EMBO Molecular Medicine 3, 129-141.
• Medina,E. (2011) Novel experimental models for dissecting genetic
susceptibility of superantigen-mediated diseases. In: Superantigens:
Molecular Basis for Their Role in Human Disease (Kotb,M. &
Frase,J.D., eds.), ASM Press, Washington, D.C., in press.
• Abel,J., Goldmann,O., Ziegler,C., Höltje,C., Smeltzer,M.S.,
Cheung,A.L., Bruhn,D., Rohde,M., & Medina,E. (2011) Staphylococcus
aureus evades the extracellular antimicrobial activity of mast cells
by promoting its own uptake. Journal of Innate Immunity, in press.
RG Microbial Interactions and Processes |
Prof. Dr. Dietmar Pieper
• Gomes,N.C., Flocco,C.G., Costa,R., Junca,H., Vilchez,R., Pieper,D.H.*.,
Krogerrecklenfort,E., Paranhos,R., Mendonca-Hagler,L.C., & Smalla,K.
(2010) Mangrove microniches determine the structural and functional
diversity of enriched petroleum hydrocarbon-degrading consortia.
FEMS Microbiology Ecology 74, 276-290.
• Marin,M., Perez-Pantoja,D., Donoso,R., Wray,V., Gonzalez,B., &
Pieper,D.H.*. (2010) Modified 3-oxoadipate pathway for the biodegradation
of methylaromatics in Pseudomonas reinekei MT1. Journal of
Bacteriology 192, 1543-1552.
• Vilchez-Vargas,R., Junca,H., & Pieper,D.H.*. (2010) Metabolic networks,
microbial ecology and ‘omics’ technologies: towards understanding
in situ biodegradation processes. Environental Microbiology
12, 3089-3104.
• Wos-Oxley,M.L., Plumeier,I., von,E.C., Taudien,S., Platzer,M., Vilchez-
Vargas,R., Becker,K., & Pieper,D.H.*. (2010) A poke into the diversity
and associations within human anterior nare microbial communities.
ISME Journal 4, 839-851.
• Junca,H. & Pieper,D.H.* (2011) Functional marker gene assays for hydrocarbon
degrading microbial communities - aerobic. In: Handbook
of Hydrocarbon Microbiology (Timmis,K.N., ed.), Springer, in press.
• Palleroni,N., Pieper,D.H.*, & Moore,E.R.B. (2011) Microbiology of
hydrocarbon-degrading Pseudomonas. In: Handbook of Hydrocarbon
Microbiology (Timmis,K.N., ed.), Springer, in press.
• Perez-Pantoja,D., Gonzalez,B., & Pieper,D.H.* (2011) Aerobic degradation
of aromatic hydrocarbons. In: Handbook of Hydrocarbon Microbiology
(Timmis,K.N., ed.), Springer, in press.
• Pieper,D.H.*., Gonzalez,B., Camara,B., Perez-Pantoja,D., & Reineke,W.
(2011) Aerobic degradation of chloroaromatics. In: Handbook of
Hydrocarbon Microbiology (Timmis,K.N., ed.), Springer, in press.
• Seeger,M. & Pieper,D.H.* (2011) Genetics of biphenyl biodegradation
and co-metabolism of PCBs. In: Handbook of Hydrocarbon Microbiology
(Timmis,K.N., ed.), Springer, in press.
• Becker,K., Mutter,W., Daniel,R., Rudack,C., Peschel,A., & Pieper,D.
(2011) Bakterielle Untermieter in der menschlichen Nase. GenomX-
Press, in press.
• Pfingsten-Würzburg,S., Pieper,D., Bautsch,W., & Probst-Kepper,M.
(2011) Prevelence and molecular epidemiology of meticillin-resistant
Staphylococcus aureus in nursing home residents in northern Germany.
Journal of Hospital Infection, in press.
• Vilchez,R., Gomez-Silvan,C., Purswani,J., Gonzalez-Lopez,J., &
Rodelas,B. (2011) Characterization of bacterial communities exposed
to Cr(III) and Pb(II) in submerged fixed-bed biofilms for groundwater
treatment. Ecotoxicology, in press.
Dept. of Vaccinology and Applied Microbiology |
Prof. Dr. Dr. Carlos Guzmán
• Becker,P.D., Royo,J.L., & Guzmán,C.A.* (2010) Exploitation of
prokaryotic expression systems based on the salicylate-dependent
control circuit encompassing nahRPsal::xylS2 for biotechnological
applications. Bioengineered Bugs 1, 246-253.
• Becker,P.D., Legrand,N., van Geelen,C.M., Noerder,M.,
Huntington,N.D., Lim,A., Yasuda,E., Diehl,S.A., Scheeren,F.A., Ott,M.,
Weijer,K., Wedemeyer,H., Di Santo,J.P., Beaumont,T., Guzmán,C.A.*,
& Spits,H. (2010) Generation of human antigen-specific monoclonal
IgM antibodies using vaccinated “human immune system” mice.
PLoS ONE 5.
• Bjorkstrom,N.K., Riese,P., Heuts,F., Andersson,S., Fauriat,C.,
Ivarsson,M.A., Bjorklund,A.T., Flodstrom-Tullberg,M., Michaelsson,J.,
Rottenberg,M.E., Guzmán,C.A.*, Ljunggren,H.G., & Malmberg,K.J.
(2010) Expression patterns of NKG2A, KIR, and CD57 define a process
of CD56dim NK-cell differentiation uncoupled from NK-cell
education. Blood 116, 3853-3864.
• Caro-Quintero,A., Deng,J., Auchtung,J., Brettar,I., Höfle,M.G.*,
Klappenbach,J., & Konstantinidis,K.T. (2010) Unprecedented levels
of horizontal gene transfer among spatially co-occurring Shewanella
bacteria from the Baltic Sea. ISME Journal 5, 140.
• Cazorla,S.I., Frank,F.M., Becker,P.D., Arnaiz,M., Mirkin,G.A.,
Corral,R.S., Guzmán,C.A.*, & Malchiodi,E.L. (2010) Redirection of the
immune response to the functional catalytic domain of the cystein
proteinase cruzipain improves protective immunity against Trypanosoma
cruzi infection. Journal of Infectious Diseases 202, 136-144.
• Chen,H.L., Lünsdorf,H., Hecht,H.J., & Tsai,H. (2010) Porcine pulmonary
angiotensin I-converting enzyme-biochemical characterization
and spatial arrangement of the N- and C-domains by three-dimensional
electron microscopic reconstruction. Micron 41, 674-685.
• Ebensen,T., Libanova,R., & Guzman,C.A.* (2010) Bis-(3’,5’)-cyclic
di-GMP: promising adjuvant for vaccine design. In The Second Messenger
Cyclic di-GMP (Wolf,A.J. & Visick,K.L.I., eds), pp. 311-319.
ASM Press, Washington, D.C., USA.
• Fritzer,A., Senn,B.M., Minh,D.B., Hanner,M., Gelbmann,D., Noiges,B.,
Henics,T., Schulze,K., Guzmán,C.A.*, Goodacre,J., von,G.A., Nagy,E.,
& Meinke,A.L. (2010) Novel conserved group A streptococcal proteins
identified by the antigenome technology as vaccine candidates for a
non-M protein-based vaccine. Infection and Immunity 78, 4051-4067.
• Guzmán,C.A.*. (2010) Research needs in vaccinology. In: Handbook
of Hydrocarbon and Lipid Microbiology (Timmis,K.N., ed.), Springer-
Verlag, Berlin Heidelberg, pp. 3383-3385.
• Guzmán,C.A.* (2010) Optimizing rational vaccine design. Microbial
Biotechnology 2, 136-138.
• Hahn,M.W., Lang,E., Brandt,U., Lünsdorf,H., Wu,Q.L., &
Stackebrandt,E. (2010) Polynucleobacter cosmopolitanus sp. nov., freeliving
planktonic bacteria inhabiting freshwater lakes and rivers.
International Journal of Systematic and Evolutionary Microbiology 60,
166-173.
• Kahlisch,L., Henne,K., Draheim,J., Brettar,I., & Höfle,M.G.* (2010)
High-resolution in situ genotyping of Legionella pneumophila populations
in drinking water by multiple-locus variable-number tandemrepeat
analysis using environmental DNA. Applied and Environmental
Microbiology 76, 6186-6195.
• Kahlisch,L., Henne,K., Gröbe,L., Draheim,J., Höfle,M.G.*, & Brettar,I.
(2010) Molecular analysis of the bacterial drinking water community
with respect to live/dead status. Water Science and Technology 61,
9-14.

160 SCIENTIFIC REPORTS | Publications
• Knothe,S., Mutschler,V., Rochlitzer,S., Winkler,C., Ebensen,T.,
Guzmán,C.A.*, Hohlfeld,J., Braun,A., & Muller,M. (2010) Local treatment
with BPPcysMPEG reduces allergic airway inflammation in
sensitized mice. Immunobiology 216, 110-117.
• Libanova,R., Ebensen,T., Schulze,K., Bruhn,D., Nörder,M., Yevsa,T.,
Morr,M., & Guzmán,C.A.* (2010) Corrigendum to “The member of
the cyclic di-nucleotide family bis- (3’,5’)-cyclic dimeric inosine
monophosphate exerts potent activity as mucosal adjuvant [Vaccine
28 (2010) 2249-2258]. Vaccine 28, 3625.
• Libanova,R., Ebensen,T., Schulze,K., Bruhn,D., Norder,M., Yevsa,T.,
Morr,M., & Guzmán,C.A.* (2010) The member of the cyclic di-nucleotide
family bis-(3’, 5’)-cyclic dimeric inosine monophosphate exerts
potent activity as mucosal adjuvant. Vaccine 28, 2249-2258.
• Nörder,M., Becker,P.D., Drexler,I., Link,C., Erfle,V., & Guzmán,C.A.*
(2010) Modified vaccinia virus Ankara exerts potent immune modulatory
activities in a murine model. PLoS ONE 5, e11400.
• Prajeeth,C.K., Jirmo,A.C., Krishnaswamy,J.K., Ebensen,T.,
Guzmán,C.A.*, Weiss,S., Constabel,H., Schmidt,R.E., & Behrens,G.M.
(2010) The synthetic TLR2 agonist BPPcysMPEG leads to efficient
cross-priming against co-administered and linked antigens. European
Journal of Immunology 40, 1272-1283.
• Roming,M., Lünsdorf,H., Dittmar,K.E.*, & Feldmann,C. (2010)
ZrO(HPO(4))(1-x)(FMN)(x): quick and easy synthesis of a nanoscale
luminescent biomarker. Angewandte Chemie International Edition
49, 632-637.
• Stegmann,K.A., Björkström,N.K., Liermann,H., Ciesek,S., Riese,P.,
Wiegand,J., Hadem,J., Suneetha,P.V., Jaroszewicz,J., Wang,C.,
Schlaphoff,V., Fytili,P., Cornberg,M., Manns,M.P., Geffers,R.,
Pietschmann,T., Guzmán,C.A.*, Ljunggren,H.-G., & Wedemeyer,H.
(2010) IFN alpha-induced TRAIL on human NK cells is associated
with control of hepatitis C virus infection. Gastroenterology 138,
1885-1897.
• Switalla,S., Lauenstein,L., Prenzler,F., Knothe,S., Forster,C.,
Fieguth,H.G., Pfennig,O., Schaumann,F., Martin,C., Guzmán,C.A.*,
Ebensen,T., Muller,M., Hohlfeld,J.M., Krug,N., Braun,A., & Sewald,K.
(2010) Natural innate cytokine response to immunomodulators and
adjuvants in human precision-cut lung slices. Toxicology and Applied
Pharmacology 246, 107-115.
• Ebensen,T., Libanova,R., & Guzmán,C.A.* (2011) Infection prevention:
oil- and lipid-containing products in vaccinology. In Handbook
of Hydrocarbon and Lipid Microbiology (Timmis,K.N., ed), pp. 3311-
3331. Springer-Verlag, Berlin, Heidelberg.
• Knothe,S., Mutschler,V., Rochlitzer,S., Winkler,C., Ebensen,T.,
Guzmán,C.A.*, Hohlfeld,J., Braun,A., & Muller,M. (2011) Local treatment
with BPPcysMPEG reduces allergic airway inflammation in
sensitized mice. Immunobiology 216, 110-117.
• Guzmán,C.A.* (2009) Research needs in vaccinology. In: Handbook
of hydrocarbon and lipid microbiology (Timmis,K.N., ed.), Springer,
in press.
• Guzmán,C.A.* & Timmis,K.N.*. (2011) Towards intelligent vaccines:
the VAC-CHIP. In the “Crystal ball - 2011” section. Microbial Biotechnology,
in press.
• Huntington,N.D., Alves,N.L., Legrand,N., Lim,A., Strick-Marchand,H.,
Mention,J.J., Plet,A., Weijer,K., Jacques,Y., Becker,P.D., Guzmán,C.
A.*, Soussan,P., Kremsdorf,D., Spits,H., & Di Santo,J.P. (2011) IL-15
transpresentation promotes both human T-cell reconstitution and T-
cell-dependent antibody responses in vivo. Proceeding of the National
Academy of Sciences USA, in press.
• TrehanPati,N., Sukriti,S., Geffers,R., Hissar,S., Riese,P., Toepfer,T.,
Buer,J., Adams,D.H., Guzmán,C.A.*, & Sarin,S.K. (2011) Acute and
resolving phase of HEV infected patients and its cellular immune
and global gene expression patterns. Journal of Clinical Immunology,
in press.
RG Chemical Microbiology | Dr. Wolf-Rainer Abraham
• Abraham,W.-R., Estrela,A.B., Nikitin,D.I., Smit,J., & Vancanneyt,M.
(2010) Brevundimonas halotolerans sp. nov., Brevundimonas poindexterae
sp. nov. and Brevundimonas staleyi sp. nov., prosthecate bacteria
from aquatic habitats. International Journal of Systematic and
Evolutionary Microbiology 60, 1837-1843.
• Abraham,W.-R., Macedo,A.J., Lünsdorf,H., & Nikitin,D. (2011) Genus
V. Arcicella Nikitin, Strömpl, Oranskaya and Abraham. 2004, 683VP.
(Arcocella Nitikin, Oranskaya, Pitryuk, Chernykh and Lysenko. 1994,
152). In: Bergey’s Manual of Systematic Bacteriology (Garrity,G.M., &
Ed.of Vol.4: Hedlund,B., Krieg,N.R., Ludwig,W., Paster,B.J., Staley,J.T.,
Ward,N., & Whitman,W.B., eds.), pp. 377-380.
• Estrela,A.B. & Abraham,W.-R. (2010) Brevundimonas vancanneytii sp.
nov., isolated from blood of a patient with endocarditis. International
Journal of Systematic and Evolutionary Microbiology 60, 2129-2134.
• Estrela,A.B. & Abraham,W.-R. (2010) Combining biofilm-controlling
compounds and antibiotics as a promising new way to control biofilm
infections. Pharmaceuticals 3, 1374.
• Estrela,A.B.*., Seixas,A., Teixeira,V.O., Pinto,A.F., & Termignoni,C.
(2010) Vitellin- and hemoglobin-digesting enzymes in Rhipicephalus
(Boophilus) microplus larvae and females. Comparative Biochemistry
and Physiology - B Biochemistry and Molecular Biology 157, 326-335.
• Jakobs-Schonwandt,D., Mathies,H., Abraham,W.-R., Pritzkow,W.,
Stephan,I., & Noll,M. (2010) Biodegradation of a biocide (Cu-N-cyclohexyldiazenium
dioxide) component of a wood preservative by a
defined soil bacterial community. Applied and Environment Microbiology
76, 8076-8083.
• Menyailo,O.V., Abraham,W.-R., & Conrad,R. (2010) Tree species affect
atmospheric CH4 oxidation without community composition of soil
methanotrophs. Soil Biology and Biochemistry 42, 101-107.
• Stiesch,M., Heuer,W., Kohorst,P., Winkel,A., Stumpp,S.N., Menzel,H.,
Pfaffenroth,C., Behrens,P., Chichkov,B., & Abraham,W.-R. (2010) Biofilmbildung
auf dentalen Implantaten - Biofilm formation on dental
implants. Biomedizinische Technik - Biomedical Engineering 55.
• Heuer,W., Stiesch,M., & Abraham,W.-R. (2011) Microbial diversity
of supra- and subgingival biofilms on freshly colonized titanium
implant abutments in the human mouth. European Journal of Clinical
Microbiology and Infectious Diseases, in press.
• Abraham,W.-R. (2011) Megacities as sources for pathogenic bacteria
in rivers and their fate downstream. International Journal of Microbiology,
in press.
• Peres de Carvalo,M. & Abraham,W.-R. (2011) Antimicrobial secondary
metabolites from fungi for food safety. In: Natural Antimicrobials
for Biosafety and Quality (Rai,M., ed.), CABI, UK, in press.
JRG Phagosome Biology | Dr. Maximiliano Gutierrez
• Bleck,C.K., Merz,A., Gutierrez,M.G.*., Walther,P., Dubochet,J.,
Zuber,B., & Griffiths,G. (2010) Comparison of different methods for
thin section EM analysis of Mycobacterium smegmatis. Journal of
Microscopy 237, 23-38.
• Gutierrez,M.G.*. (2010) Salmonella vacuole maturation: PIKfyve
leads the way. Embo Journal 29, 1316-1317.
• Wahe,A., Kasmapour,B., Schmaderer,C., Liebl,D., Sandhoff,K.,
Nykjaer,A., Griffiths,G., & Gutierrez,M.G.*. (2010) Golgi-to-phagosome
transport of acid sphingomyelinase and prosaposin is mediated
by sortilin. Journal of Cell Science 123, 2502-2511.
• Zygmunt,B.M., Gröbe,L., & Guzmán,C.A.* (2011) Peritoneal cavity is
dominated by IFNgamma-secreting CXCR3+ Th1 cells. PLOS ONE, in
press.

SCIENTIFIC REPORTS | Publications
161
JRG Immune Aging and Chronic Infections |
Dr. Dr. Luka Cicin-Sain
• Cicin-Sain,L., Smyk-Pearson,S., Currier,N., Byrd,L., Koudelka,C.,
Robinson,T., Swarbrick,G., Tackitt,S., Legasse,A., Fischer,M.,
Nikolich-Zugich,D., Park,B., Hobbs,T., Doane,C.J., Mori,M.,
Axthelm,M.K., Lewinsohn,D.A., & Nikolich-Zugich,J. (2010) Loss of
naive T cells and repertoire constriction predict poor response to vaccination
in old primates. Journal of Immunology 184, 6739-6745.
• Voss,A., Gescher,K., Hensel,A., Nacken,W.& Kerkhoff,C. (2011) Double-
Stranded RNA InducesMMP-9 Gene Expression in HaCaT Keratinocytes
by Tumor Necrosis Factor-α. Inflamm Allergy Drug Targets, in press.
Dept. Infection Genetics | Prof. Dr. Klaus Schughart
• Alberts,R. & Schughart,K. (2010) QTLminer: identifying genes regulating
quantitative traits. BMC Bioinformatics 11, 516.
• Alberts,R., Srivastava,B., Wu,H., Viegas,N., Geffers,R., Klawonn,F.,
Novoselova,N., Zaverucha,d., V, Panthier,J.J. & Schughart,K. (2010)
Gene expression changes in the host response between resistant
and susceptible inbred mouse strains after influenza A infection.
Microbes and Infection 12, 309-318.
• Bertram,S., Glowacka,I., Blazejewska,P., Soilleux,E., Allen,P.,
Danisch,S., Steffen,I., Choi,S.Y., Park,Y., Schneider,H., Schughart,K. &
Pohlmann,S. (2010) TMPRSS2 and TMPRSS4 facilitate trypsin-independent
spread of influenza virus in Caco-2 cells. Journal of Virology
84, 10016-10025.
• Bruck,N., Gahr,M. & Pessler,F. (2010) Transient oligoarthritis of the
lower extremity following influenza B virus infection: Case report.
Pediatric Rheumatology 8, 4.
• Bucher,K., Dietz,K., Lackner,P., Pasche,B., Fendel,R., Mordmuller,B.
Ben-Smith,A., & Hoffmann,W.H. (2010) Schistosoma co-infection
protects against brain pathology but does not prevent severe disease
and death in a murine model of cerebral malaria. International Journal
for Parasitology 41, 21-31.
• Dai,L., Zhu,L.J., Zheng,D.H., Mo,Y.Q., Wei,X.N., Su,J.H., Pessler,F. &
Zhang,B.Y. (2010) Elevated serum glucose-6-phosphate isomerase
correlates with histological disease activity and clinical improvement
after initiation of therapy in patients with rheumatoid arthritis.
Journal of Rheumatology 37, 2452-2461.
• do Valle,T.Z., Billecocq,A., Guillemot,L., Alberts,R., Gommet,C.,
Geffers,R., Calabrese,K., Schughart,K., Bouloy,M., Montagutelli,X., &
Panthier,J.J. (2010) A new mouse model reveals a critical role for host
innate immunity in resistance to Rift Valley fever. Journal of Immunology
185, 6146-6156.
• Durzynska,J., Blazejewska,P., Szydlowski,J. & Gozdzicka-Jozefiak,A.
(2010) Detection of anti-HPV11-L1 antibodies in immune sera from
patients suffering from recurrent respiratory papillomatosis using
ELISA. Viral Immunology 23, 415-423.
• Faller,G., Mikolajczyk,R.T., Akmatov,M.K.*., Meier,S. & Kramer,A.
(2010) Accidents in the context of study among university students-
-a multicentre cross-sectional study in North Rhine-Westphalia,
Germany. Accidents Analysis Prevention 42, 487-491.
• Gruenberger,M., Alberts,R., Smedley,D., Swertz,M., Schofield,P. &
Schughart,K. (2010) Towards the integration of mouse databases -
definition and implementation of solutions to two use-cases in mouse
functional genomics. BMC Research Notes 3, 16.
• Hahn,P., Wegener,I., Burrells,A., Bose,J., Wolf,A., Erck,C., Butler,D.,
Schofield,C.J., Bottger,A. & Lengeling,A. (2010) Analysis of Jmjd6 cellular
localization and testing for its involvement in histone demethylation.
PLoS ONE 5, e13769.
• Hauck,F., Lee-Kirsch,M.A., Aust,D., Roesler,J. & Pessler,F. (2010) Complement
C2 deficiency disarranging innate and adaptive humoral
immune responses in a pediatric patient: Treatment with rituximab.
Arthritis Care Res (Hoboken.) 63, 454-459.
• Holz,A., Kollmus,H., Ryge,J., Niederkofler,V., Dias,J., Ericson,J.,
Stoeckli,E.T., Kiehn,O. & Arnold,H.H. (2010) The transcription factors
Nkx2.2 and Nkx2.9 play a novel role in floor plate development and
commissural axon guidance. Development 137, 4249-4260.
• Kim,J.K., Kayali,G., Walker,D., Forrest,H.L., Ellebedy,A.H., Griffin,Y.S.,
Rubrum,A., Bahgat,M.M.*., Kutkat,M.A., Ali,M.A., Aldridge,J.R.,
Negovetich,N.J., Krauss,S., Webby,R.J. & Webster,R.G. (2010) Puzzling
inefficiency of H5N1 influenza vaccines in Egyptian poultry.
Proceeding of the National Academy of Sciences of the United States of
America 107, 11044-11049.
• Meier,S., Mikolajczyk,R.T., Helmer,S., Akmatov,M., Steinke,B. &
Krämer,A. (2010) Health status of students - Results from a multicentre
cross-sectional study in North Rhine-Westphalia, Germany
[Prävalenz von Erkrankungen und Beschwerden bei Studierenden
in NRW - Ergebnisse des Gesundheitssurveys NRW]. Prävention und
Gesundheitsforderung 5, 257-264.
• Michaelson,J.J., Alberts,R., Schughart,K. & Beyer,A. (2010) Data-driven
assessment of eQTL mapping methods. BMC GENOMICS 11, 502.
• Moller,J., Girschick,H.J., Hahn,G. & Pessler,F. (2010) [Steroid-induced
spinal epidural lipomatosis in pediatric patients]. Zeitschrift Rheumatologie
69, 447-449.
• Ogdie,A., Schumacher,H.R., Jr., Dai,L., Chen,L.X., Einhorn,E. &
Pessler,F. (2010) Synovial biopsy findings in arthritis associated with
hepatitis C virus infection. Journal of Rheumatology 37, 1361-1363.
• Ogdie,A., Li,J., Dai,L., Paessler,M.E., Yu,X., az-Torne,C., Akmatov,M.,
Schumacher,H.R. & Pessler,F. (2010) Identification of broadly discriminatory
tissue biomarkers of synovitis with binary and multicategory
receiver operating characteristic analysis. Biomarkers 15,
183-190.
• Schofield,P.N., Eppig,J., Huala,E., De Angelis,M.H., Harvey,M.,
Davidson,D., Weaver,T., Brown,S., Smedley,D., Rosenthal,N.,
Schughart,K., Aidinis,V., Tocchini-Valentini,G. & Hancock,J.M. (2010)
Research funding. Sustaining the data and bioresource commons.
Science 330, 592-593.
• Schughart,K. (2010) SYSGENET: a meeting report from a new European
network for systems genetics. Mammalian Genome 21, 331-336.
• Slansky,E., Li,J., Haupl,T., Morawietz,L., Krenn,V. & Pessler,F. (2010)
Quantitative determination of the diagnostic accuracy of the synovitis
score and its components. Histopathology 57, 436-443.
• Smedley,D., Schofield,P., Chen,C.K., Aidinis,V., Ainali,C., Bard,J.,
Balling,R., Birney,E., Blake,A., Bongcam-Rudloff,E., Brookes,A.J.,
Cesareni,G., Chandras,C., Eppig,J., Flicek,P., Gkoutos,G.,
Greenaway,S., Gruenberger,M., Heriche,J.K., Lyall,A., Mallon,A.M.,
Muddyman,D., Reisinger,F., Ringwald,M., Rosenthal,N., Schughart,K.,
Swertz,M., Thorisson,G.A., Zouberakis,M. & Hancock,J.M. (2010)
Finding and sharing: new approaches to registries of databases and
services for the biomedical sciences. Database (Oxford) 2010, baq014.
• Swertz,M.A., Velde,K.J., Tesson,B.M., Scheltema,R.A., Arends,D.,
Vera,G., Alberts,R., Dijkstra,M., Schofield,P., Schughart,K.,
Hancock,J.M., Smedley,D., Wolstencroft,K., Goble,C., de Brock,E.O.,
Jones,A.R., Parkinson,H.E. & Jansen,R.C. (2010) XGAP: a uniform and
extensible data model and software platform for genotype and phenotype
experiments. Genome Biology 11, R27.
• Wu,H., Haist,V., Baumgartner,W., & Schughart,K. (2010) Sustained
viral load and late death in Rag2-/- mice after influenza A virus
infection. Virology Journal 7, 172.
• Zouberakis,M., Chandras,C., Swertz,M., Smedley,D., Gruenberger,M.,
Bard,J., Schughart,K., Rosenthal,N., Hancock,J.M., Schofield,P.N.,
Kollias,G. & Aidinis,V. (2010) Mouse Resource Browser – a database
of mouse databases. Database (Oxford) 2010, baq010.
• Akmatov,M.K.*. (2011) Child abuse in 28 developing and transitional
countries – results from the Multiple Indicator Cluster Surveys.
International Journal of Epidemiology 40, 219-227.
• Bahgat,M.M., Blazejewska,P. & Schughart,K. (2011) Inhibition of lung
serine proteases in mice: a potentially new approach to control influenza
infection. Virology Journal 8, 27.

162 SCIENTIFIC REPORTS | Publications
• Blazejewska,P., Koscinski,L., Viegas,N., Anhlan,D., Ludwig,S. &
Schughart,K. (2011) Pathogenicity of different PR8 influenza A virus
variants in mice is determined by both viral and host factors. Virology
412, 36-45.
• Bruck,N., Suttorp,M., Kabus,M., Heubner,G., Gahr,M. & Pessler,F.
(2011) Rapid and sustained remission of systemic juvenile idiopathic
arthritis-associated macrophage activation syndrome through treatment
with anakinra and corticosteroids. Journal of Clinical Rheumatology
17, 23-27.
• Gemulla,G. & Pessler,F. (2011) Can norovirus infection lead to a
postinfectious arthritis? Report of 2 possible cases. Klinische Pädiatrie
223, 43-44.
• Schneider,J., Fricke,C., Overwin,H. & Hofer,B. (2011) High level expression
of a recombinant amylosucrase gene and selected properties
of the enzyme. Applied Microbiology and Biotechnology, in press.
• Akmatov,M., Mikolajczyk,R.T., Meier,S. & Krämer,A. (2011) Alcohol
consumption among university students in North Rhine-Westphalia,
Germany - results from a multicentre cross-sectional study. Journal
of American College Health, in press.
• Alberts,R. & Schughart,K. (2011) High throughput gene expression
analysis and the identification of expression QTLs. In Gene Discovery
for Disease Models John Wiley & Sons, Inc., Hoboken, New Jersey,
USA, in press.
• Dai,L., Wei,X.N., Zheng,D.H., Mo,Y.Q., Pessler,F. & Zhang,B.Y. (2011)
Effective treatment of Kimura’s disease with leflunomide in combination
with glucocorticoids. Clinical Rheumatology, in press.
• Hauck,F., Lee-Kirsch,M.A., Aust,D., Roesler,J. & Pessler,F. (2011)
Disarranged innate and adaptive humoral immune responses in
complement 2 deficiency: treatment with rituximab. Arthritis Care
and Research (Hoboken), in press.
• Pessler,F. & Sherry,D.D. (2011) Rheumatic Fever. In The Merck
Manual, in press.
• von Jagwitz,M., Pessler,F. & Akmatov,M. (2011) Exhaled breath condensate
pH as risk factor for asthma in asymptomatic children with
prior wheezing. Journal of Allergy and Clinical Immunology, in press.
AG Systemorientierte Immunologie und Entzündungsforschung |
Prof. Dr. Ingo Schmitz
• Brandt,S., Ellwanger,K., Beuter-Gunia,C., Schuster,M., Hausser,A.,
Schmitz,I., & Beer-Hammer,S. (2010) SLy2 targets the nuclear
SAP30/HDAC1 complex. International Journal of Biochemistry and
Cell Biology 42, 1472-1481.
• Emadi,B.M., Soheili,Z.S., Schmitz,I., Sameie,S., & Schulz,W.A. (2010)
Snail regulates cell survival and inhibits cellular senescence in human
metastatic prostate cancer cell lines. Cell Biology and Toxicology
26, 553-567.
• Smith,A.J., Dai,H., Correia,C., Takahashi,R., Lee,S.H., Schmitz,I., &
Kaufmann,S.H. (2011) Noxa/Bcl-2 interactions contribute to bortezomib
resistance in human lymphoid cells. Journal of Biological
Chemistry, in press.
Abt. für Experimentelle Immunologie | Prof. Dr. Jochen Hühn
• Hubert,S., Rissiek,B., Klages,K., Hühn,J., Sparwasser,T., Haag,F., Koch-
Nolte,F., Boyer,O., Seman,M., & Adriouch,S. (2010) Extracellular NAD+
shapes the Foxp3+ regulatory T cell compartment through the ART2-
P2X7 pathway. Journal of Experimental Medicine 207, 2561-2568.
• Humrich,J.Y., Morbach,H., Undeutsch,R., Enghard,P., Rosenberger,S.,
Weigert,O., Kloke,L., Heimann,J., Gaber,T., Brandenburg,S.,
Scheffold,A., Hühn,J., Radbruch,A., Burmester,G.R., & Riemekasten,G.
(2010) Homeostatic imbalance of regulatory and effector T cells due
to IL-2 deprivation amplifies murine lupus. Proceedings of the National
Academy of Sciences of the United States of America 107, 204-209.
• Klages,K., Mayer,C.T., Lahl,K., Loddenkemper,C., Teng,M.W.,
Ngiow,S.F., Smyth,M.J., Hamann,A., Hühn,J., & Sparwasser,T. (2010)
Selective depletion of Foxp3+ regulatory T cells improves effective
therapeutic vaccination against established melanoma. Cancer
Research 70, 7788-7799.
• Mayer,C.T., Floess,S., Baru,A.M., Lahl,K., Hühn,J., & Sparwasser,T.
(2011) CD8(+) Foxp3(+) T cells share developmental and phenotypic
features with classical CD4(+) Foxp3(+) regulatory T cells but lack
potent suppressive activity. European Journal of Immunology 41, 716-
725.
• Menning,A., Loddenkemper,C., Westendorf,A.M., Szilagyi,B., Buer,J.,
Siewert,C., Hamann,A., & Hühn,J. (2010) Retinoic acid-induced gut
tropism improves the protective capacity of Treg in acute but not
in chronic gut inflammation. European Journal of Immunology 40,
2539-2548.
• Polansky,J.K., Schreiber,L., Thelemann,C., Ludwig,L., Kruger,M.,
Baumgrass,R., Cording,S., Floess,S., Hamann,A., & Hühn,J. (2010)
Methylation matters: binding of Ets-1 to the demethylated Foxp3
gene contributes to the stabilization of Foxp3 expression in regulatory
T cells. Journal of Molecular Medicine 88, 1029-1040.
• Toker,A. & Hühn,J. (2011) To be or not to be a Treg cell: lineage decisions
controlled by epigenetic mechanisms. Science Signaling 4, e4.
RG Immunoregulation | Priv.-Doz. Dr. Dunja Bruder
• Bar-On,L., Birnberg,T., Lewis,K.L., Edelson,B.T., Bruder,D., Hildner,K.,
Buer,J., Murphy,K.M., Reizis,B., & Jung,S. (2010) CX3CR1+ CD8alpha+
dendritic cells are a steady-state population related to plasmacytoid
dendritic cells. Proceedings of the National Acadamy of
Sciences of the United States of America 107, 14745-14750.
• Bruns,S., Kniemeyer,O., Hasenberg,M., Aimanianda,V., Nietzsche,S.,
Thywissen,A., Jeron,A., Latge,J.P., Brakhage,A.A., & Gunzer,M. (2010)
Production of extracellular traps against Aspergillus fumigatus in vitro
and in infected lung tissue is dependent on invading neutrophils
and influenced by hydrophobin RodA. PLoS Pathogens 6, e1000873.
• Liesz,A., Zhou,W., Mracskó,E., Karcher,S., Doerr,H., Schwarting,S.,
Sun,L., Bruder,D., Stegemann,S., Cerwenka,A., Sommer,C., Dalpke,A.,
& Veltkamp,R. (2011) Inhibition of lymphocyte trafficking shields the
brain against deleterious neuroinflammation after stroke. Brain 134,
704-720.
• Hasenberg,M., Köhler,A., Jeron,A., Bonifatius,S., & Gunzer,M. (2011)
Direct observation of phagocytosis and NET formation by neutrophils
in infected lungs using 2-photon microscopy. Journal of Visualized
Experiments, in press.
• Tosiek,M.J., Gruber,A.D., Bader,S., Hoymann,H.G., Mauel,S.,
Tschwernig,T., Buer,J., Gereke,M., & Bruder,D. (2011) CD4+CD25+Foxp3+
regulatory T cells are dispensable for controlling CD8+ T cellmediated
lung inflammation. Journal of Immunology, in press.
Dept. of Systems Immunology | Prof. Dr. Michael Meyer-Hermann
• Garin,A., Meyer-Hermann,M., Contie,M., Figge,M.T., Buatois,V.,
Gunzer,M., Toellner,K.M., Elson,G., & Kosco-Vilbois,M.H. (2010)
Toll-like receptor 4 signaling by follicular dendritic cells is pivotal for
germinal center onset and affinity maturation. Immunity 33, 84-95.
• Jain,H.V. & Meyer-Hermann,M. (2010) The molecular asis of synergism
between carboplatin and ABT-737 therapy targeting ovarian
carcinomas. Cancer Research 71, 705-715.
• Kempf,H., Bleicher,M., & Meyer-Hermann,M. (2010) Spatio-temporal
cell dynamics in tumour spheroid irradiation. European Physical
Journal D 60, 177-193.
• Kobayashi,Y. & Telschow,A. (2010) Cytoplasmic feminizing elements
in a two-population model: infection dynamics, gene flow modification,
and the spread of autosomal suppressors. Journal of Evolutionary
Biology 23, 2558-2568.

SCIENTIFIC REPORTS | Publications
163
• Meyer-Hermann,M. & Benninger,R.K. (2010) A mathematical model
of beta-cells in an islet of Langerhans sensing a glucose gradient.
HFSP Journal 4, 61-71.
• Victora,G.D., Schwickert,T.A., Fooksman,D.R., Kamphorst,A.O., Meyer-Hermann,M.,
Dustin,M.L., & Nussenzweig,M.C. (2010) Germinal
center dynamics revealed by multiphoton microscopy with a photoactivatable
fluorescent reporter. Cell 143, 592-605.
• Grise,G. & Meyer-Hermann,M. (2011) Surface reconstruction using
Delaunay triangulation for applications in life sciences. Computer
Physics Communications 182, 967-977.
• Kobayashi,Y. & Telschow,A. (2011) The concept of effective recombination
rate and its application in speciation theory. Evolution 65,
617-628.
Abt. für Chemische Biologie | Dr. Ronald Frank, Dr. Florenz Sasse
• Andreetto,E., Yan,L.M., Tatarek-Nossol,M., Velkova,A., Frank,R., &
Kapurniotu,A. (2010) Identification of hot regions of the abeta-IAPP
interaction interface as high-affinity binding sites in both cross- and
self-association. Angewandte Chemie International Edition 49, 3081-
• Barnickel,B., Bayliffe,F., Diestel,R., Kempf,K., Laschat,S., Pachali,S.,
Sasse,F., Schlenk,A., & Schobert,R. (2010) Structure-activity relationships
of precursors and analogs of natural 3-enoyl-tetramic acids.
Chemistry & Biodiversity 7, 2830-2845.
• Beutling,U. & Frank,R. (2010) Epitope analysis using synthetic peptide
repertoires prepared by SPOT synthesis technology. In: Antibody
Engineering (Kontermann,R. & Dübel,S., eds.), Springer-Verlag,
Berlin, Heidelberg.
• Bourbeillon,J., Orchard,S., Benhar,I., Borrebaeck,C., de,D.A.,
Dubel,S., Frank,R., Gibson,F., Gloriam,D., Haslam,N., Hiltker,T.,
Humphrey-Smith,I., Hust,M., Juncker,D., Koegl,M., Konthur,Z.,
Korn,B., Krobitsch,S., Muyldermans,S., Nygren,P.A., Palcy,S., Polic,B.,
Rodriguez,H., Sawyer,A., Schlapshy,M., Snyder,M., Stoevesandt,O.,
Taussig,M.J., Templin,M., Uhlen,M., van der,M.S., Wingren,C.,
Hermjakob,H., & Sherman,D. (2010) Minimum information about a
protein affinity reagent (MIAPAR). Nature Biotechnology 28, 650-653.
• Brodmann,T., Janssen,D., Sasse,F., Irschik,H., Jansen,R., Müller,R., &
Kalesse,M. (2010) Isolation and synthesis of chivotriene, a chivosazole
shunt product from Sorangium cellulosum. European Journal of
Organic Chemistry 27, 5155-5159.
• Buchner,A., Pohla,H., Willimsky,G., Frankenberger,B., Frank,R., Baur-
Melnyk,A., Siebels,M., Stief,C.G., Hofstetter,A., Kopp,J., Pezzutto,A.,
Blankenstein,T., Oberneder,R., & Schendel,D.J. (2010) Phase 1 trial of
allogeneic gene-modified tumor cell vaccine RCC-26/CD80/IL-2 in
patients with metastatic renal cell carcinoma. Human Gene Therapy
21, 285-297.
• Bulow,L., Nickeleit,I., Girbig,A.K., Brodmann,T., Rentsch,A.,
Eggert,U., Sasse,F., Steinmetz,H., Frank,R., Carlomagno,T.,
Malek,N.P., & Kalesse,M. (2010) Synthesis and biological characterization
of argyrin F. ChemMedChem 5, 832-836.
• Frank,R., Theis,F.J., & Klamt,S. (2010) From binary to multivalued to
continuous models: the lac operon as a case study. Journal of Integrative
Bioinformatics 7, 151-169.
• Gloriam,D.E., Orchard,S., Bertinetti,D., Bjorling,E., Bongcam-
Rudloff,E., Borrebaeck,C.A., Bourbeillon,J., Bradbury,A.R., de,D.A.,
Dubel,S., Frank,R., Gibson,T.J., Gold,L., Haslam,N., Herberg,F.W.,
Hiltke,T., Hoheisel,J.D., Kerrien,S., Koegl,M., Konthur,Z., Korn,B.,
Landegren,U., Montecchi-Palazzi,L., Palcy,S., Rodriguez,H., Schweinsberg,S.,
Sievert,V., Stoevesandt,O., Taussig,M.J., Ueffing,M., Uhlen,M.,
van der,M.S., Wingren,C., Woollard,P., Sherman,D.J., & Hermjakob,H.
(2010) A community standard format for the representation of protein
affinity reagents. Molecular and Cellular Proteomics 9, 1-10.
• Harmrolfs,K., Brunjes,M., Drager,G., Floss,H.G., Sasse,F., Taft,F., &
Kirschning,A. (2010) Cyclization of synthetic seco-proansamitocins
to ansamitocin macrolactams by Actinosynnema pretiosum as biocatalyst.
ChemBioChem 11, 2517-2520.
• Heinzelmann,K., Scholz,B.A., Nowak,A., Fossum,E., Kremmer,E.,
Haas,J., Frank,R., & Kempkes,B. (2010) Kaposi’s sarcoma-associated
herpesvirus viral interferon regulatory factor 4 (vIRF4/K10) is a
novel interaction partner of CSL/CBF1, the major downstream effector
of Notch signaling. Journal of Virology 84, 12255-12264.
• Kubicek,K., Grimm,S.K., Orts,J., Sasse,F., & Carlomagno,T. (2010)
The tubulin-bound structure of the antimitotic drug tubulysin. Angewandte
Chemie International Edition 49, 4809-4812.
• Meens,J., Bolotin,V., Frank,R., Bohmer,J., & Gerlach,G.F. (2010) Characterization
of a highly immunogenic Mycoplasma hyopneumoniae
lipoprotein Mhp366 identified by peptide-spot array. Veterinary
Microbiology 142, 293-302.
• Meyer,T., Stratmann-Selke,J., Meens,J., Schirrmann,T., Gerlach,G.F.,
Frank,R., Dubel,S., Strutzberg-Minder,K., & Hust,M. (2010) Isolation
of scFv fragments specific to OmpD of Salmonella typhimurium.
Veterinary Microbiology 147, 162-169.
• Micklinghoff,J.C., Schmidt,M., Geffers,R., Tegge,W., & Bange,F.C.
(2010) Analysis of expression and regulatory functions of the
ribosome-binding protein TypA in Mycobacterium tuberculosis under
stress conditions. Archives of Microbiology 192, 499-504.
• Nickl,C.K., Raidas,S.K., Zhao,H., Sausbier,M., Ruth,P., Tegge,W.,
Brayden,J.E., & Dostmann,W.R. (2010) (d)-Amino acid analogues of
DT-2 as highly selective and superior inhibitors of cGMP-dependent
protein kinase Ialpha. Biochimica et Biophysica Acta 1804, 524-532.
• Nicolas,L., Anderl,T., Sasse,F., Steinmetz,H., Jansen,R., Hofle,G.,
Laschat,S., & Taylor,R.E. (2010) Gephyronic acid, a missing link
between polyketide inhibitors of eukaryotic protein synthesis (Part I):
Structural revision and stereochemical assignment of gephyronic
acid. Angewandte Chemie International Edition 50, 938-941.
• Rink,C., Sasse,F., Zubriene,A., Matulis,D., & Maier,M.E. (2010) Probing
the influence of an allylic methyl group in zearalenone analogues
on binding to Hsp90. Chemistry 16, 14469-14478.
• Schackel,R., Hinkelmann,B., Sasse,F., & Kalesse,M. (2010) The synthesis
of novel disorazoles. Angewandte Chemie International Edition
49, 1619-1622.
• Schlenk,A., Diestel,R., Sasse,F., & Schobert,R. (2010) A selective
3-acylation of tetramic acids and the first synthesis of ravenic acid.
Chemistry 16, 2599-2604.
• Schobert,R., Sasse,F., Biersack,B., Effenberger,K., Breyer,S., &
Diestel,R. (2010) Tumor-selective amphiphilic para-quinones and
tetramic acids. International Journal of Clinical Pharmacology and
Therapeutics 48, 459-461.
• Schulz,S., Dickschat,J.S., Kunze,B., Wagner-Döbler,I., Diestel,R., &
Sasse,F. (2010) Biological activity of volatiles from marine and terrestrial
bacteria. Marine Drugs 8, 2976-2987.
• Sharma,R.K., Sundriyal,S., Wangoo,N., Tegge,W., & Jain,R. (2010)
New antimicrobial hexapeptides: synthesis, antimicrobial activities,
cytotoxicity, and mechanistic studies. ChemMedChem 5, 86-95.
• Voigt,J., Kiess,M., Getzlaff,R., Wostemeyer,J., & Frank,R. (2010) Generation
of the heterodimeric precursor GP3 of the Chlamydomonas
cell wall. Molecular Microbiology 77, 1512-1526.
• Capell,A., Liebscher,S., Fellerer,K., Brouwers,N., Willem,M.,
Lammich,S., Gijselinck,I., Bittner,T., Carlson,A.M., Sasse,F., Kunze,B.,
Steinmetz,H., Jansen,R., Dormann,D., Sleegers,K., Cruts,M., Herms,J.,
Van,B.C., & Haass,C. (2011) Rescue of progranulin deficiency associated
with frontotemporal lobar degeneration by alkalizing reagents
and inhibition of vacuolar ATPase. Journal of Neuroscience 31, 1885-
1894.
• Elsebai,M.F., Kehraus,S., Lindequist,U., Sasse,F., Shaaban,S.,
Gutschow,M., Josten,M., Sahl,H.G., & Konig,G.M. (2011) Antimicrobial
phenalenone derivatives from the marine-derived fungus Coniothyrium
cereale. Organic and Biomolecular Chemistry 9, 802-808.

164 SCIENTIFIC REPORTS | Publications
• Knobloch,T., Harmrolfs,K., Taft,F., Thomaszewski,B., Sasse,F., &
Kirschning,A. (2011) Mutational biosynthesis of ansamitocin antibiotics:
A diversity-oriented approach to exploit biosynthetic flexibility.
ChemBioChem 13, 540-547.
• Omnus,D.J., Mehrtens,S., Ritter,B., Resch,K., Yamada,M., Frank,R.,
Nourbakhsh,M., & Reboll,M.R. (2011) JKTBP1 is involved in stabilization
and IRES-dependent translation of NRF mRNAs by binding to 5’
and 3’ untranslated regions. Journal of Molecular Biology 407, 492-
504.
• Wunderlich,K., Juozapaitis,M., Ranadheera,C., Kessler,U., Martin,A.,
Eisel,J., Beutling,U., Frank,R., & Schwemmle,M. (2011) Identification
of high-affinity PB1-derived peptides with enhanced affinity to the
PA protein of Influenza A virus polymerase. Antimicrobial Agents and
Chemotherapy 55, 696-702.
• Anderl,T., Nicolas,L., Munkemer,J., Baro,A., Sasse,F., Steinmetz,H.,
Jansen,R., Hofle,G., Taylor,R.E., & Laschat,S. (2011) Gephyronic acid,
a missing link between polyketide inhibitors of eukaryotic protein
synthesis (Part II): Total synthesis of gephyronic acid. Angewandte
Chemie International Edition, in press.
• Bremer,C.M., Sominskaya,I., Skrastina,D., Pumpens,P., Wahed,A.A.,
Beutling,U., Frank,R., Fritz,H.J., Hunsmann,G., Gerlich,W.H., &
Glebe,D. (2011) N-terminal myristoylation-dependent masking of
neutralizing epitopes in the preS1 attachment site of hepatitis B
virus. Journal of Hepatology, in press.
• Kraemer,S., Lue,H., Zernecke,A., Kapurniotu,A., Andreetto,E.,
Frank,R., Lennartz,B., Weber,C., & Bernhagen,J. (2011) MIF-chemokine
receptor interactions in atherogenesis are dependent on an
N-loop-based 2-site binding mechanism. FASEB Journal, in press.
• Maenz,B., Goetz,V., Wunderlich,K., Eisel,J., Kirchmair,J., Stech,J.,
Stech,O., Chase,G., Frank,R., & Schwemmle,M. (2011) Disruption of
the viral polymerase complex assembly as a novel approach to attenuate
influenza a virus. Journal of Biological Chemistry, in press.
• Ritter,B., Kilian,P., Reboll,M.R., Resch,K., Distefano,J.K., Frank,R.,
Beil,W., & Nourbakhsh,M. (2011) Differential effects of multiplicity
of infection on Helicobacter pylori-induced signaling pathways and
interleukin-8 gene transcription. Journal of Clinical Immunology, in
press.
• Schneider,J., Fricke,C., Overwin,H., & Hofer,B. (2011) High level
expression of a recombinant amylosucrase gene and selected properties
of the enzyme. Applied Microbiology and Biotechnology, in press.
• Westermann,J., Florcken,A., Willimsky,G., van,L.A., Kopp,J.,
Takvorian,A., Johrens,K., Lukowsky,A., Schonemann,C., Sawitzki,B.,
Pohla,H., Frank,R., Dorken,B., Schendel,D.J., Blankenstein,T., &
Pezzutto,A. (2011) Allogeneic gene-modified tumor cells (RCC-26/
IL-7/CD80) as a vaccine in patients with metastatic renal cell cancer:
a clinical phase-I study. Gene Therapy, in press.
RG Biological Systems Analysis | Prof. Dr. Ursula Bilitewski
• Klippel,N., Cui,S., Groebe,L., & Bilitewski,U. (2010) Deletion of the
Candida albicans histidine kinase gene CHK1 improves recognition
by phagocytes through an increased exposure of cell wall beta-1,3-
glucans. Microbiology 156, 3432-3444.
Dept. of Medicinal Chemistry | Prof. Dr. Markus Kalesse
• Brodmann,T., Janssen,D., Sasse,F., Irschik,H., Jansen,R., Müller,R., &
Kalesse,M. (2010) Isolation and synthesis of chivotriene, a chivosazole
shunt product from Sorangium cellulosum. European Journal of
Organic Chemistry 27, 5155-5159.
• Brodmann,T., Janssen,D., & Kalesse,M. (2010) Total synthesis of
chivosazole F. Journal of the American Chemical Society 132, 13610-
13611.
• Bulow,L., Nickeleit,I., Girbig,A.K., Brodmann,T., Rentsch,A.,
Eggert,U., Sasse,F., Steinmetz,H., Frank,R., Carlomagno,T.,
Malek,N.P., & Kalesse,M. (2010) Synthesis and biological characterization
of argyrin F. ChemMedChem 5, 832-836.
• Kena,D.A., Noll,C., Richter,M., Gieseler,M.T., & Kalesse,M. (2010)
Intramolecular stereoselective protonation of aldehyde-derived enolates.
Angewandte Chemie International Edition 49, 8367-8369.
• Li,P., Li,J., Arikan,F.*., Ahlbrecht,W., Dieckmann,M., & Menche,D.
(2010) Stereoselective total synthesis of etnangien and etnangien
methyl ester. Journal of Organic Chemistry 75, 2429-2444.
• Menche,D., Li,P., & Irschik,H. (2010) Design, synthesis and biological
evaluation of simplified analogues of the RNA polymerase inhibitor
etnangien. Bioorganic and Medicinal Chemistry Letters 20, 939-941.
• Mohamed,I.E., Kehraus,S., Krick,A., Konig,G.M., Kelter,G., Maier,A.,
Fiebig,H.H., Kalesse,M., Malek,N.P., & Gross,H. (2010) Mode of action
of epoxyphomalins A and B and characterization of related metabolites
from the marine-derived fungus Paraconiothyrium sp. Journal of
Naturald Products 73, 2053-2056.
• Rand,K., Noll,C., Schiebel,H.M., Kemken,D., Dulcks,T., Kalesse,M.,
Heinz,D.W.*., & Layer,G. (2010) The oxygen-independent coproporphyrinogen
III oxidase HemN utilizes harderoporphyrinogen as a
reaction intermediate during conversion of coproporphyrinogen III to
protoporphyrinogen IX. Biological Chemistry 391, 55-63.
• Schackel,R., Hinkelmann,B., Sasse,F., & Kalesse,M. (2010) The synthesis
of novel disorazoles. Angewandte Chemie International Edition
49, 1619-1622.
• Stauch,B., Simon,B., Basile,T., Schneider,G., Malek,N.P., Kalesse,M., &
Carlomagno,T. (2010) Elucidation of the structure and intermolecular
interactions of a reversible cyclic-peptide inhibitor of the proteasome
by NMR spectroscopy and molecular modeling. Angewandte Chemie
International Edition 49, 3934-3938.
RG Environmental Microbiology | Prof. Dr. Ken N. Timmis
• Beloqui,A., Nechitaylo,T.Y., Lopez-Cortes,N., Ghazi,A.,
Guazzaroni,M.E., Polaina,J., Strittmatter,A.W., Reva,O., Waliczek,A.,
Yakimov,M.M., Golyshina,O.V., Ferrer,M., & Golyshin,P.N. (2010) Diversity
of glycosyl hydrolases from cellulose-depleting communities
enriched from casts of two earthworm species. Applied and Environmental
Microbiology 76, 5934-5946.
• Beloqui,A., Polaina,J., Vieites,J.M., Reyes-Duarte,D., Torres,R.,
Golyshina,O.V., Chernikova,T.N., Waliczek,A., Aharoni,A.,
Yakimov,M.M., Timmis,K.N.*., Golyshin,P.N., & Ferrer,M. (2010)
Novel hybrid esterase-haloacid dehalogenase enzyme. ChemBioChem
11, 1975-1978.
• Fazzini,R.A., Preto,M.J., Quintas,A.C., Bielecka,A., Timmis,K.N.*.,
& dos Santos,V.A.P.M.*. (2010) Consortia modulation of the stress
response: proteomic analysis of single strain versus mixed culture.
Environmental Microbiology 12, 2436-2449.
• Gertler,C., Näther,D.J., Gerdts,G., Malpass,M.C., & Golyshin,P.N.*.
(2010) A mesocosm study of the changes in marine flagellate and
ciliate communities in a crude oil bioremediation trial. Microbial
Ecology 60, 180-191.
• Martin-Arjol,I., Bassas-Galia,M.*., Bermudo,E., Garcia,F., &
Manresa,A. (2010) Identification of oxylipins with antifungal activity
by LC-MS/MS from the supernatant of Pseudomonas 42A2. Chemistry
and Physics of Lipids 163, 341-346.
• Nechitaylo,T.Y., Yakimov,M.M., Godinho,M., Timmis,K.N.*.,
Belogolova,E., Byzov,B.A., Kurakov,A.V., Jones,D.L., & Golyshin,P.N.
(2010) Effect of the earthworms Lumbricus terrestris and Aporrectodea
caliginosa on bacterial diversity in soil. Microbial Ecology 59,
574-587.

SCIENTIFIC REPORTS | Publications
165
• Nechitaylo,T.Y., Timmis,K.N.*., Byzov,B.A., Kurakov,A.V., Jones,D.L.,
Ferrer,M., Belogolova,E., & Golyshin,P.N. (2010) Fate of prions in soil:
Degradation of recombinant prion in aqueous extracts from soil and
casts of two earthworm species. Soil Biology and Biochemistry 42,
1168-1171.
• Oxley,A.P., Lanfranconi,M.P., Wurdemann,D., Ott,S., Schreiber,S.,
McGenity,T.J., Timmis,K.N.*., & Nogales,B. (2010) Halophilic archaea
in the human intestinal mucosa. Environmental Microbiology 12,
2398-2410.
• Timmis,K.N.*. (2010) Human biome biotechnology and the personalization
of odour profiles. Microbial Biotechnology 2, 150-152.
• Wiethaus,J., Busch,A.W., Dammeyer,T., & Frankenberg-Dinkel,N.
(2010) Phycobiliproteins in Prochlorococcus marinus: biosynthesis of
pigments and their assembly into proteins. European Journal of Cell
Biology 89, 1005-1010.
• Ferrer,M., Beloqui,A., Zumárraga,M., Alcalde,M., & Golyshin,P.N.*
(2011) Microbes and enzymes: recent trends and new directions to
explore protein diversity space. In: Protein Engineering Handbook
(Bornscheuer,U., ed.), Wiley, in press
• Gertler,C., Gerdts,G., Yakimov,M.M., Timmis,K.N.*., & Golyshin,P.N.*.
(2011) Populations of heavy fuel oil-degrading marine microbial community
on oil-degrading marine microbial community on oil sorbent
material surface. Journal of Applied Microbiology, in press.
• Guzmán,C.A.*. & Timmis,K.N.* (2011) Towards intelligent vaccines:
the VAC-CHIP. In the “Crystal ball - 2011” section. Microbial Biotechnology,
in press
• Sabirova,J.S., Haddouche,R., Van Bogaert,I.N., Mulaa,F., Verstraete,W.,
Timmis,K.N.*, Schmid-Dannert,C., Nicaud,J.M., & Soetaert,W. (2011)
The Lipo Yeast project: using the oleaginous yeast Yarrowia lipolytica
in combination with specific bacterial genes for the bioconversion of
lipids, fats and oils into high-value products. Microbial Biotechnology,
in press.
Dept. Genome Analysis | Dr. Helmut Blöcker
• Buntin,K., Irschik,H., Weissman,K.J., Luxenburger,E., Blöcker,H., &
Müller,R. (2010) Biosynthesis of thuggacins in myxobacteria: comparative
cluster analysis reveals basis for natural product structural
diversity. Chemistry and Biology 17, 342-356.
• Danilowicz,E., Martinez-Arias,R., Dolf,G., Singh,M., Probst,I.,
Tummler,B., Holtig,D., Waldmann,K.H., Gerlach,G.F., Stanke,F., &
Leeb,T. (2010) Characterization of the porcine transferrin gene (TF)
and its association with disease severity following an experimental
Actinobacillus pleuropneumoniae infection. Animal Genetics 41, 424-
427.
• Deyneko,I.V., Kalybaeva,Y.M., Kel,A.E., & Blöcker,H. (2010) Humanchimpanzee
promoter comparisons: Property-conserved evolution?
Genomics 96, 129-133.
• Dötsch,A., Klawonn,F., Jarek,M., Scharfe,M., Blöcker,H., & Häußler,S.
(2010) Evolutionary conservation of essential and highly expressed
genes in Pseudomonas aeruginosa. BMC GENOMICS 11, 234.
• Hu,Y., van der,G.R., Besra,G.S., Gurcha,S.S., Liu,A., Rohde,M.,
Singh,M., & Coates,A. (2010) 3-Ketosteroid 9alpha-hydroxylase is an
essential factor in the pathogenesis of Mycobacterium tuberculosis.
Molecular Microbiology 75, 107-121.
• Lin,L., Flisikowski,K., Schwarzenbacher,H., Scharfe,M., Severitt,S.,
Blöcker,H., & Fries,R. (2010) Characterization of the procine AMPK
alpha 2 catalytic subunitgene (PRKAA2): genome structure, polymorphism
detection and association study. Animal Genetics 41(2), 203-
207
• Scholler,J., Singh,M., Bergmeier,L., Brunstedt,K., Wang,Y., Whittall,T.,
Rahman,D., Pido-Lopez,J., & Lehner,T. (2010) A recombinant human
HLA-class I antigen linked to dextran elicits innate and adaptive immune
responses. Journal of Immunological Methods 360, 1-9.
• Varbiro,S., Biro,A., Cervenak,J., Cervenak,L., Singh,M., Banhidy,F.,
Sebestyen,A., Fust,G., & Prohaszka,Z. (2010) Human anti-60 kD heat
shock protein autoantibodies are characterized by basic features of
natural autoantibodies. Acta Physiologica Hungarica 97, 1-10.
• von Groll,A., Martin,A., Stehr,M., Singh,M., Portaels,F., da Silva,P.E.,
& Palomino,J.C. (2010) Fitness of Mycobacterium tuberculosis strains
of the W-Beijing and Non-W-Beijing genotype. PLoS ONE 5, e10191.
• Adhikary,T., Kaddatz,K., Finkernagel,F., Schonbauer,A., Meissner,W.,
Scharfe,M., Jarek,M., Blöcker,H., Muller-Brusselbach,S., & Müller,R.
(2011) Genomewide analyses define different modes of transcriptional
regulation by peroxisome proliferator-activated receptor-beta/
delta (PPARbeta/delta). PLoS ONE 6, e16344.
Former Dept. of Cell Biology | Prof. Dr. Jürgen Wehland
• Alberts,R., Srivastava,B., Wu,H., Viegas,N., Geffers,R., Klawonn,F.,
Novoselova,N., Zaverucha,d., V, Panthier,J.J., & Schughart,K. (2010)
Gene expression changes in the host response between resistant
and susceptible inbred mouse strains after influenza A infection.
Microbes and Infection 12, 309-318.
• Bruns,S., Kniemeyer,O., Hasenberg,M., Aimanianda,V., Nietzsche,S.,
Thywissen,A., Jeron,A., Latge,J.P., Brakhage,A.A., & Gunzer,M. (2010)
Production of extracellular traps against Aspergillus fumigatus in vitro
and in infected lung tissue is dependent on invading neutrophils
and influenced by hydrophobin RodA. PLoS Pathogens 6, e1000873.
• Christgen,M., Geffers,R., Ballmaier,M., Christgen,H., Poczkaj,J.,
Krech,T., Kreipe,H., & Lehmann,U. (2010) Down-regulation of the
fetal stem cell factor SOX17 by H33342: a mechanism responsible for
differential gene expression in breast cancer side population cells.
Journal of Biological Chemistry 285, 6412-6418.
• Dietrich,N., Rohde,M., Geffers,R., Kroger,A., Hauser,H., Weiss,S., &
Gekara,N.O. (2010) Mast cells elicit proinflammatory but not type I
interferon responses upon activation of TLRs by bacteria. Proceeding
of the National Academy of Sciences of the United States of America
107, 8748-8753.
• do Valle,T.Z., Billecocq,A., Guillemot,L., Alberts,R., Gommet,C.,
Geffers,R., Calabrese,K., Schughart,K., Bouloy,M., Montagutelli,X., &
Panthier,J.J. (2010) A new mouse model reveals a critical role for host
innate immunity in resistance to Rift Valley fever. Journal of Immunology
185, 6146-6156.
• Fleissner,D., Hansen,W., Geffers,R., Buer,J., & Westendorf,A.M. (2010)
Local induction of immunosuppressive CD8+ T cells in the gut-associated
lymphoid tissues. PLoS ONE 5, e15373.
• Garin,A., Meyer-Hermann,M., Contie,M., Figge,M.T., Buatois,V.,
Gunzer,M., Toellner,K.M., Elson,G., & Kosco-Vilbois,M.H. (2010)
Toll-like receptor 4 signaling by follicular dendritic cells is pivotal for
germinal center onset and affinity maturation. Immunity 33, 84-95.
• Garritsen,H.S., Macke,L., Meyring,W., Hannig,H., Pagelow,U.,
Wormann,B., Geffers,R., Dittmar,K.E.*., & Lindenmaier,W. (2010)
Efficient generation of clinical-grade genetically modified dendritic
cells for presentation of multiple tumor-associated proteins. Transfusion
50, 831-842.
• Gekara,N.O., Zietara,N., Geffers,R., & Weiss,S. (2010) Listeria monocytogenes
induces T cell receptor unresponsiveness through poreforming
toxin listeriolysin O. Journal of Infectious Diseases 202,
1698-1707.
• Hansen,W., Westendorf,A.M., Toepfer,T., Mauel,S., Geffers,R.,
Gruber,A.D., & Buer,J. (2010) Inflammation in vivo is modulated by
GPR83 isoform-4 but not GPR83 isoform-1 expression in regulatory
T cells. Genes and Immunity 11, 357-361.
• Kahlisch,L., Henne,K., Gröbe,L., Draheim,J., Höfle,M.G.*., & Brettar,I.
(2010) Molecular analysis of the bacterial drinking water community
with respect to live/dead status. Water Science and Technology 61,
9-14.

166 SCIENTIFIC REPORTS | Publications
• Lefever,T., Pedersen,E., Basse,A., Paus,R., Quondamatteo,F.,
Stanley,A.C., Langbein,L., Wu,X., Wehland,J., Lommel,S., &
Brakebusch,C. (2010) N-WASP is a novel regulator of hair-follicle
cycling that controls antiproliferative TGF{beta} pathways. Journal of
Cell Science 123, 128-140.
• Leonhardt,J., Kuebler,J.F., Turowski,C., Tschernig,T., Geffers,R., &
Petersen,C. (2010) Susceptibility to experimental biliary atresia
linked to different hepatic gene expression profiles in two mouse
strains. Hepatology Research 40, 196-203.
• Macke,L., Garritsen,H.S., Meyring,W., Hannig,H., Pagelow,U.,
Wormann,B., Piechaczek,C., Geffers,R., Rohde,M., Lindenmaier,W.,
& Dittmar,K.E.*. (2010) Evaluating maturation and genetic modification
of human dendritic cells in a new polyolefin cell culture bag
system. Transfusion 50, 843-855.
• Micklinghoff,J.C., Schmidt,M., Geffers,R., Tegge,W., & Bange,F.C.
(2010) Analysis of expression and regulatory functions of the
ribosome-binding protein TypA in Mycobacterium tuberculosis under
stress conditions. Archives of Microbiology 192, 499-504.
• Pils,M.C., Pisano,F., Fasnacht,N., Heinrich,J.M., Gröbe,L.,
Schippers,A., Rozell,B., Jack,R.S., & Muller,W. (2010) Monocytes/
macrophages and/or neutrophils are the target of IL-10 in the LPS
endotoxemia model. European Journal of Immunology 40, 443-448.
• Quiel,A., Jurgen,B., Piechotta,G., Le Foll,A.P., Ziebandt,A.K., Kohler,C.,
Koster,D., Engelmann,S., Erck,C., Hintsche,R., Wehland,J., Hecker,M.,
& Schweder,T. (2010) Electrical protein array chips for the detection
of staphylococcal virulence factors. Applied Microbiology and Biotechnology
85, 1619-1627.
• Stegmann,K.A., Björkström,N.K., Liermann,H., Ciesek,S., Riese,P.,
Wiegand,J., Hadem,J., Suneetha,P.V., Jaroszewicz,J., Wang,C.,
Schlaphoff,V., Fytili,P., Cornberg,M., Manns,M.P., Geffers,R.,
Pietschmann,T., Guzmán,C.A.*., Ljunggren,H.-G., & Wedemeyer,H.
(2010) IFN alpha-induced TRAIL on human NK cells is associated
with control of hepatitis C virus infection. Gastroenterology 138,
1885-1897.
• Steinweg,C., Kuenne,C.T., Billion,A., Mraheil,M.A., Domann,E.,
Ghai,R., Barbuddhe,S.B., Karst,U., Goesmann,A., Puhler,A.,
Weisshaar,B., Wehland,J., Lampidis,R., Kreft,J., Goebel,W.,
Chakraborty,T., & Hain,T. (2010) Complete genome sequence of Listeria
seeligeri, a nonpathogenic member of the genus Listeria. Journal
of Bacteriology 192, 1473-1474.
• Trunk,K., Benkert,B., Quack,N., Munch,R., Scheer,M., Garbe,J.,
Jänsch,L., Trost,M., Wehland,J., Buer,J., Jahn,M., Schobert,M., &
Jahn,D. (2010) Anaerobic adaptation in Pseudomonas aeruginosa:
definition of the Anr and Dnr regulons. Environmental Microbiology
12, 1719-1733.
• Wilke,S., Krausze,J., Gossen,M., Gröbe,L., Jager,V., Gherardi,E., van
den,H.J., & Bussow,K. (2010) Glycoprotein production for structure
analysis with stable, glycosylation mutant CHO cell lines established
by fluorescence-activated cell sorting. Protein Science 19, 1264-1271.
• Froese,N., Kattih,B., Breitbart,A., Grund,A., Geffers,R., Molkentin,J.D.,
Kispert,A., Wollert,K.C., Drexler,H., & Heineke,J. (2011) GATA6
promotes angiogenic function and survival in endothelial cells by
suppression of autocrine transforming growth factor beta/activin
receptor-like kinase 5 signaling. Journal of Biological Chemistry
286(7), 5680-5690
• Lorenz,U., Lorenz,B., Schmitter,T., Streker,K., Erck,C., Wehland,J.,
Nickel,J., Zimmermann,B., & Ohlsen,K. (2011) Functional antibodies
targeting IsaA of Staphylococcus aureus augment host immune
response and open new perspectives for antibacterial therapy. Antimicrobial
Agents and Chemotherapy 55, 165-173.
• Schenk,U., Frascoli,M., Proietti,M., Geffers,R., Traggiai,E., Buer,J.,
Ricordi,C., Westendorf,A.M., & Grassi,F. (2011) ATP inhibits the
generation and function of regulatory T cells through the activation
of purinergic P2X receptors. Science Signaling 4, ra12.
• Kusch,K., Hanke,K., Holtfreter,S., Schmudde,M., Kohler,C., Erck,C.,
Wehland,J., Hecker,M., Ohlsen,K., Bröker,B. & Engelmann,S. (2011)
The influence of SaeRS and σ B on the expression of superantigens
in different Staphylococcus aureus isolates. International Journal of
Microbiology, in press
• TrehanPati,N., Sukriti,S., Geffers,R., Hissar,S., Riese,P., Toepfer,T.,
Buer,J., Adams,D.H., Guzmán,C.A.*, & Sarin,S.K. (2011) Acute and
resolving phase of HEV infected patients and its cellular immune
and global gene expression patterns. Journal of Clinical Immunology,
in press
Former RG Cytoskeleton Dynamics | Prof. Dr. Klemens Rottner
• Hanisch,J., Ehinger,J., Ladwein,M., Rohde,M., Derivery,E., Bosse,T.,
Steffen,A., Bumann,D., Misselwitz,B., Hardt,W.D., Gautreau,A.,
Stradal,T.B.*., & Rottner,K. (2010) Molecular dissection of Salmonella-induced
membrane ruffling versus invasion. Cellular Microbiology
12, 84-98
• Hertzog,M., Milanesi,F., Hazelwood,L., Disanza,A., Liu,H., Perlade,E.,
Malabarba,M.G., Pasqualato,S., Maiolica,A., Confalonieri,S., Le,C.C.,
Offenhauser,N., Block,J., Rottner,K., Di Fiore,P.P., Carlier,M.F.,
Volkmann,N., Hanein,D., & Scita,G. (2010) Molecular basis for the
dual function of Eps8 on actin dynamics: bundling and capping.
PLoS Biology 8, e1000387.
• Klink,B.U., Barden,S., Heidler,T.V., Borchers,C., Ladwein,M.,
Stradal,T.B.*., Rottner,K., & Heinz,D.W.*. (2010) Structure of Shigella
IpgB2 in complex with human RhoA: implications for the mechanism
of bacterial guanine nucleotide exchange factor mimicry. Journal of
Biological Chemistry 285, 17197-17208.
• Marg,S., Winkler,U., Sestu,M., Himmel,M., Schonherr,M., Bar,J.,
Mann,A., Moser,M., Mierke,C.T., Rottner,K., Blessing,M., Hirrlinger,J.,
& Ziegler,W.H. (2010) The vinculin-DeltaIn20/21 mouse: characteristics
of a constitutive, actin-binding deficient splice variant of vinculin.
PLoS ONE 5, e11530.
• Rottner,K., Hanisch,J., & Campellone,K.G. (2010) WASH, WHAMM
and JMY: regulation of Arp2/3 complex and beyond. Trends in Cell
Biology 20, 650-661.
• Jackson,B., Peyrollier,K., Pedersen,E., Basse,A., Karlsson,R., Wang,Z.,
Lefever,T., Ochsenbein,A.M., Schmidt,G., Aktories,K., Stanley,A.,
Quondamatteo,F., Ladwein,M., Rottner,K., van,H.J., & Brakebusch,C.
(2011) RhoA is dispensable for skin development, but crucial for contraction
and directed migration of keratinocytes. Molecular Biology of
the Cell 22, 593-605.
Former RG Signalling and Motility | Prof. Dr. Theresia Stradal
• Hanisch,J., Ehinger,J., Ladwein,M., Rohde,M., Derivery,E., Bosse,T.,
Steffen,A., Bumann,D., Misselwitz,B., Hardt,W.D., Gautreau,A.,
Stradal,T.B.*., & Rottner,K. (2010) Molecular dissection of Salmonella-induced
membrane ruffling versus invasion. Cellular Microbiology
12, 84-98.
• Klink,B.U., Barden,S., Heidler,T.V., Borchers,C., Ladwein,M.,
Stradal,T.B.*., Rottner,K., & Heinz,D.W.*. (2010) Structure of Shigella
IpgB2 in complex with human RhoA: implications for the mechanism
of bacterial guanine nucleotide exchange factor mimicry. Journal of
Biological Chemistry 285, 17197-17208.
• Stradal,T.B.*. & Backert,S. (2010) Host-pathogen interaction: How
EHEC influence the actin cytoskeleton of the host cell [Wirt-Pathogen-Interaktion
wie EHEC das Aktinzytoskelett der Wirtszelle beeinflussen].
Biospektrum 16, 624-627.
• Tahirovic,S., Hellal,F., Neukirchen,D., Hindges,R., Garvalov,B.K.,
Flynn,K.C., Stradal,T.E.*., Chrostek-Grashoff,A., Brakebusch,C., &
Bradke,F. (2010) Rac1 regulates neuronal polarization through the
WAVE complex. Journal of Neuroscience 30, 6930-6943.

SCIENTIFIC REPORTS | Publications
167
• Breitsprecher,D., Kiesewetter,A.K., Linkner,J., Vinzenz,M.,
Stradal,T.E.*., Small,J.V., Curth,U., Dickinson,R.B., & Faix,J. (2011)
Molecular mechanism of Ena/VASP-mediated actin-filament elongation.
Embo Journal 30, 456-467.
Former RG Systems and Synthetic Biology | Prof. Dr. Vitor
Martins dos Santos
• Fazzini,R.A., Preto,M.J., Quintas,A.C., Bielecka,A., Timmis,K.N.*.,
& dos Santos,V.A.P.M.*. (2010) Consortia modulation of the stress
response: proteomic analysis of single strain versus mixed culture.
Environmental Microbiology 12, 2436-2449.
• Koutinas,M., Kiparissides,A., Lam,M.-C., Silva-Rocha,R., de
Lorenzo,V., dos Santos,V.A.P.M.*., Pistikopoulos,E.N., & Mantalaris,A.
(2010) Combining genetic circuit and microbial growth kinetic models:
A challenge for biological modelling. Computer Aided Chemical
Engineering 28, 301-306.
• Koutinas,M., Lam,M.C., Kiparissides,A., Silva-Rocha,R., Godinho,M.,
Livingston,A.G., Pistikopoulos,E.N., de,L., V, Dos,S.V., & Mantalaris,A.
(2010) The regulatory logic of m-xylene biodegradation by Pseudomonas
putida mt-2 exposed by dynamic modelling of the principal
node Ps/Pr of the TOL plasmid. Environmental Microbiology 12, 1705-
1718.
• Lam,C.M.C., Puchalka,J., Diez,M.S., & dos Santos,V.A.P.M.*. (2010)
Exploring networks at the genome scale. European Biotechnology
Science and Industry News 9, 40-42.
Dept. of Microbial Natural Products – HIPS & RG Microbial
Drugs – HZI | Prof. Dr. Rolf Müller
• Arp,G., Bissett,A., Brinkmann,N., Cousin,S., de Beer,D., Friedl,T.,
Mohr,K.I.*., Neu,T.R., Reimer,A., Shiraishi,F., Stackebrandt,E.,
& Zippel,B. (2010) Tufa-forming biofilms of German karstwater
streams: Microorganisms, exopolymers, hydrochemistry and calcification.
Geological Society Stecial Publication 336, 83-118.
• Binz,T.M., Maffioli,S.I., Sosio,M., Donadio,S., & Müller,R. (2010) Insights
into an unusual nonribosomal peptide synthetase biosynthesis:
identification and characterization of the GE81112 biosynthetic
gene cluster. Journal of Biological Chemistry 285, 32710-32719.
• Brodmann,T., Janssen,D., Sasse,F., Irschik,H., Jansen,R., Müller,R., &
Kalesse,M. (2010) Isolation and synthesis of chivotriene, a chivosazole
shunt product from Sorangium cellulosum. European Journal of
Organic Chemistry 27, 5155-5159.
• Bulow,L., Nickeleit,I., Girbig,A.K., Brodmann,T., Rentsch,A.,
Eggert,U., Sasse,F., Steinmetz,H., Frank,R., Carlomagno,T.,
Malek,N.P., & Kalesse,M. (2010) Synthesis and biological characterization
of argyrin F. ChemMedChem 5, 832-836.
• Buntin,K., Irschik,H., Weissman,K.J., Luxenburger,E., Blöcker,H., &
Müller,R. (2010) Biosynthesis of thuggacins in myxobacteria: comparative
cluster analysis reveals basis for natural product structural
diversity. Chemistry and Biology 17, 342-356.
• Buntin,K., Weissman,K.J., & Müller,R. (2010) An unusual thioesterase
promotes isochromanone ring formation in ajudazol biosynthesis.
ChemBioChem 11, 1137-1146.
• Chai,Y., Pistorius,D., Ullrich,A., Weissman,K.J., Kazmaier,U., &
Müller,R. (2010) Discovery of 23 natural tubulysins from Angiococcus
disciformis An d48 and Cystobacter SBCb004. Chemistry and Biology
17, 296-309.
• Daum,M., Schnell,H.J., Herrmann,S., Gunther,A., Murillo,R.,
Müller,R., Bisel,P., Muller,M., & Bechthold,A. (2010) Functions of
genes and enzymes involved in phenalinolactone biosynthesis.
ChemBioChem 11, 1383-1391.
• Erol,O., Schaberle,T.F., Schmitz,A., Rachid,S., Gurgui,C., El,O.M.,
Lohr,F., Kehraus,S., Piel,J., Müller,R., & Konig,G.M. (2010) Biosynthesis
of the myxobacterial antibiotic corallopyronin A. ChemBioChem
11, 1253-1265.
• Garcia,R., Gerth,K., Stadler,M., Dogma,I.J., Jr., & Müller,R. (2010)
Expanded phylogeny of myxobacteria and evidence for cultivation of
the ‘unculturables’. Molecular Phylogenetics and Evolution 57, 878-
887.
• Irschik,H., Kopp,M., Weissman,K.J., Buntin,K., Piel,J., & Müller,R.
(2010) Analysis of the sorangicin gene cluster reinforces the utility of
a combined phylogenetic/retrobiosynthetic analysis for deciphering
natural product assembly by trans-AT PKS. ChemBioChem 11, 1840-
1849.
• Khatri,Y., Hannemann,F., Ewen,K.M., Pistorius,D., Perlova,O.,
Kagawa,N., Brachmann,A.O., Müller,R., & Bernhardt,R. (2010) The
CYPome of Sorangium cellulosum So ce56 and identification of
CYP109D1 as a new fatty acid hydroxylase. Chemistry and Biology 17,
1295-1305.
• Kunze,B., Reck,M., Dotsch,A., Lemme,A., Schummer,D., Irschik,H.,
Steinmetz,H., & Wagner-Döbler,I. (2010) Damage of Streptococcus
mutans biofilms by carolacton, a secondary metabolite from the
myxobacterium Sorangium cellulosum. BMC Microbiology 10, 199.
• Li,Y., Weissman,K.J., & Müller,R. (2010) Insights into multienzyme
docking in hybrid PKS-NRPS megasynthetases revealed by heterologous
expression and genetic engineering. ChemBioChem 11, 1069-
1075.
• Menche,D., Li,P., & Irschik,H. (2010) Design, synthesis and biological
evaluation of simplified analogues of the RNA polymerase inhibitor
etnangien. Bioorganic and Medicinal Chemistry Letters 20, 939-941.
• Nawrath,T., Gerth,K., Müller,R., & Schulz,S. (2010) The biosynthesis
of the aroma volatile 2-methyltetrahydrothiophen-3-one in the bacterium
Chitinophaga Fx7914. ChemBioChem 11, 1914-1919.
• Nawrath,T., Gerth,K., Müller,R., & Schulz,S. (2010) Volatile methyl
esters of medium chain length from the bacterium Chitinophaga
Fx7914. Chemistry and Biodiversity 7, 2228-2253.
• Nicolas,L., Anderl,T., Sasse,F., Steinmetz,H., Jansen,R., Höfle,G.,
Laschat,S., & Taylor,R.E. (2010) Gephyronic Acid, a Missing Link between
Polyketide Inhibitors of Eukaryotic Protein Synthesis (Part I):
Structural Revision and Stereochemical Assignment of Gephyronic
Acid. Angewandte Chemie International Edition 50, 938-941.
• Rachid,S., Revermann,O., Dauth,C., Kazmaier,U., & Müller,R. (2010)
Characterization of a novel type of oxidative decarboxylase involved
in the biosynthesis of the styryl moiety of chondrochloren from an
acylated tyrosine. Journal of Biological Chemistry 285, 12482-12489.
• Vilchez,R., Lemme,A., Ballhausen,B., Thiel,V., Schulz,S., Jansen,R.,
Sztajer,H., & Wagner-Döbler,I. (2010) Streptococcus mutans inhibits
Candida albicans hyphal formation by the fatty acid signaling molecule
trans-2-decenoic acid (SDSF). ChemBioChem 11, 1552-1562.
• Weissmann,K.J. & Müller,R. (2010) Myxobacterial secondary metabolites:
Bioactivities and modes-of-action. Natural Product Reports 27,
1276-1295.
• Adhikary,T., Kaddatz,K., Finkernagel,F., Schonbauer,A., Meissner,W.,
Scharfe,M., Jarek,M., Blöcker,H., Muller-Brusselbach,S., & Müller,R.
(2011) Genomewide analyses define different modes of transcriptional
regulation by peroxisome proliferator-activated receptor-beta/
delta (PPARbeta/delta). PLoS ONE 6, e16344.
• Capell,A., Liebscher,S., Fellerer,K., Brouwers,N., Willem,M.,
Lammich,S., Gijselinck,I., Bittner,T., Carlson,A.M., Sasse,F., Kunze,B.,
Steinmetz,H., Jansen,R., Dormann,D., Sleegers,K., Cruts,M., Herms,J.,
Van,B.C., & Haass,C. (2011) Rescue of progranulin deficiency associated
with frontotemporal lobar degeneration by alkalizing reagents
and inhibition of vacuolar ATPase. Journal of Neuroscience 31, 1885-
1894.

168 SCIENTIFIC REPORTS | Publications
• Dehn,R., Katsuyama,Y., Weber,A., Gerth,K., Jansen,R., Steinmetz,H.,
Höfle,G., Müller,R., & Kirschning,A. (2011) Molecular basis of elansolid
biosynthesis: evidence for an unprecedented quinone methide
inititiated intramolecular Diels-Alder cycloaddition / macrolactonization.
Angewandte Chemie 50, 532-536.
• Huntley,S., Hamann,N., Wegener-Feldbrugge,S., Treuner-Lange,A.,
Kube,M., Reinhardt,R., Klages,S., Müller,R., Ronning,C.M.,
Nierman,W.C., & Sogaard-Andersen,L. (2011) Comparative genomic
analysis of fruiting body formation in Myxococcales. Molecular
Biology and Evolution 28, 1083-1097.
• Steinmetz,H., Gerth,K., Jansen,R., Schlager,N., Dehn,R., Reinecke,S.,
Kirschning,A., & Müller,R. (2011) Elansolid A, a unique macrolide antibiotic
from Chitinophaga sancti isolated as two stable atropisomers.
Angewandte Chemie International Edtion 50, 532-536.
• Wenzel,S.C. & Müller,R. (2011) Myxobacteria - Unique microbial
secondary metabolic factories. In: Comprehensive Natural Products
II: Chemistry and Biology, Elsevier Science & Technology, Oxford, pp.
189-222.
• Anderl,T., Nicolas,L., Munkemer,J., Baro,A., Sasse,F., Steinmetz,H.,
Jansen,R., Hofle,G., Taylor,R.E., & Laschat,S. (2011) Gephyronic Acid,
a Missing Link between Polyketide Inhibitors of Eukaryotic Protein
Synthesis (Part II): Total Synthesis of Gephyronic Acid. Angewandte
Chemie International Edition, in press.
• Garcia,R., Pistorius,D., Stadler,M., & Müller,R. (2011) Fatty acid Related
Phylogeny of Myxobacteria as an Approach to Discover Polyunsaturated
Omega-3/6 Fatty Acids. Journal of Bacteriology, in press.
• Okanya,P.W., Mohr,K.I., Gerth,K., Jansen,R., & Müller,R. (2011) Marinoquinolines
A-F, Pyrroloquinolines from Ohtaekwangia kribbensis
(Bacteroidetes). Journal of Natural Products, in press.
• Pistorius,D., Ullrich,A., Lucas,S., Hartmann,R.W., Kazmaier,U., &
Müller,R. (2011) Biosynthesis of 2-Alkyl-4(1H)-Quinolones in Pseudomonas
aeruginosa: Potential for Therapeutic Interference with
Pathogenicity. ChemBioChem, in press.
• Rachid,S., Huo,L., Herrmann,J., Stadler,M., Kopcke,B., Bitzer,J., &
Müller,R. (2011) Mining the cinnabaramide biosynthetic pathway to
generate novel proteasome inhibitors. ChemBioChem, in press.
• Zander,W., Gerth,K., Mohr,K.I., Kessler,W., Jansen,R., & Müller,R.
(2011) Roimatacene, an antibiotic against gram-negative bacteria
isolated from cystobacter Cb G35 (myxobacteria). Chemistry - A European
Journal, in press.
Dept. of Drug Delivery – HIPS | Prof. Dr. Claus-Michael Lehr
• Beisner,J., Dong,M., Taetz,S., Nafee,N., Griese,E.U., Schaefer,U.,
Lehr,C.-M., Klotz,U., & Murdter,T.E. (2010) Nanoparticle mediated
delivery of 2’-O-methyl-RNA leads to efficient telomerase inhibition
and telomere shortening in human lung cancer cells. Lung Cancer
68, 346-354.
• Bur,M., Huwer,H., Muys,L., & Lehr,C.-M. (2010) Drug transport across
pulmonary epithelial cell monolayers: effects of particle size, apical
liquid volume, and deposition technique. Journal of Aerosol Medicine
and Pulmonary Drug Delivery 23, 119-127.
• Ciana,A., Meier,K., Daum,N., Gerbes,S., Veith,M., Lehr,C.-M., &
Minetti,G. (2010) A dynamic ratio of the alpha+ and alpha- isoforms
of the tight junction protein ZO-1 is characteristic of Caco-2 cells and
correlates with their degree of differentiation. Cell Biology International
34, 669-678.
• Collnot,E.M., Baldes,C., Schaefer,U.F., Edgar,K.J., Wempe,M.F., &
Lehr,C.-M. (2010) Vitamin E TPGS P-glycoprotein inhibition mechanism:
influence on conformational flexibility, intracellular ATP levels,
and role of time and site of access. Molecular Pharmaceutics 7,
642-651.
• Gaschen,A., Lang,D., Kalberer,M., Savi,M., Geiser,T., Gazdhar,A.,
Lehr,C.-M., Bur,M., Dommen,J., Baltensperger,U., & Geiser,M. (2010)
Cellular responses after exposure of lung cell cultures to secondary
organic aerosol particles. Environmental Science and Technology 44,
1424-1430.
• Henning,A., Schneider,M., Nafee,N., Muijs,L., Rytting,E., Wang,X.,
Kissel,T., Grafahrend,D., Klee,D., & Lehr,C.-M. (2010) Influence of
particle size and material properties on mucociliary clearance from
the airways. Journal of Aerosol Medicine and Pulmonary Drug Delivery
23, 233-241.
• Henning,A., Hein,S., Schneider,M., Bur,M., & Lehr,C.-M. (2010) Pulmonary
drug delivery: medicines for inhalation. Handbook of Experimental
Pharmacology 171-192.
• Lehmann,A.D., Daum,N., Bur,M., Lehr,C.-M., Gehr,P., & Rothen-
Rutishauser,B.M. (2010) An in vitro triple cell co-culture model with
primary cells mimicking the human alveolar epithelial barrier. European
Journal of Pharmaceutics and Biopharmaceutics 77, 398-406.
• Leonard,F., Collnot,E.M., & Lehr,C.-M. (2010) A three-dimensional
coculture of enterocytes, monocytes and dendritic cells to model
inflamed intestinal mucosa in vitro. Molecular Pharmaceutics 7,
2103-2119.
• Melero,A., Lehr,C.-M., Schafer,U.F., & Garrigues,T.M. (2010) Wistar
rat skin as surrogate for human skin in nortriptyline hydrochloride
patch studies. International Journal of Pharmaceutics 384, 137-139.
• Muendoerfer,M., Schaefer,U.F., Koenig,P., Walk,J.S., Loos,P.,
Balbach,S., Eichinger,T., & Lehr,C.-M. (2010) Online monitoring of
transepithelial electrical resistance (TEER) in an apparatus for combined
dissolution and permeation testing. International Journal of
Pharmaceutics 392, 134-140.
• Neumeyer,A., Bukowski,M., Veith,M., Lehr,C.-M., & Daum,N. (2010)
Interaction of nanoparticles and cells - cellular uptake and cytotoxic
effects in vitro. In Modern Polymeric Materials for Environmental
Applications (Pielichowski,K., ed), pp. 239-246. Cracow.
• Philippi,C., Loretz,B., Schaefer,U.F., & Lehr,C.-M. (2010) Telomerase
as an emerging target to fight cancer - Opportunities and challenges
for nanomedicine. Journal of Controlled Release 146, 228-240.
• Reum,N., Fink-Straube,C., Klein,T., Hartmann,R.W.*., Lehr,C.-M., &
Schneider,M. (2010) Multilayer coating of gold nanoparticles with
drug-polymer coadsorbates. Langmuir 26, 16901-16908.
• Rytting,E., Bur,M., Cartier,R., Bouyssou,T., Wang,X., Kruger,M.,
Lehr,C.-M., & Kissel,T. (2010) In vitro and in vivo performance of
biocompatible negatively-charged salbutamol-loaded nanoparticles.
Journal of Controlled Release 141, 101-107.
• Santander-Ortega,M.J., Stauner,T., Loretz,B., Ortega-Vinuesa,J.L.,
Bastos-Gonzalez,D., Wenz,G., Schaefer,U.F., & Lehr,C.-M. (2010) Nanoparticles
made from novel starch derivatives for transdermal drug
delivery. Journal of Controlled Release 141, 85-92.
• Hahn,T., Hansen,S., Neumann,D., Kostka,K.-H., Lehr,C.-M., Muys,L.,
& Schaefer,U.F. (2011) Infrared densitometry: A fast and non-destructive
method for exact stratum corneum depth calculation for in vitro
tape-stripping. Skin Pharmacology and Physiology 23, 183-192.
• Hahn,T., Winkler,K., Lehr,C.-M., & Schäfer,U.F. (2011) Ointment bases
and the rate of water loss of the skin [Salbengrundlagen und die
Wasserabgaberate der Haut]. Deutsche Apotheker Zeitung 150, 59-62.
• Hansen,S., Selzer,D., Schaefer,U.F., & Kasting,G.B. (2011) An extended
database of keratin binding. Journal of Pharmaceutical Sciences
100, 1712-1726.
• Hein,S., Bur,M., Schaefer,U.F., & Lehr,C.-M. (2011) A new Pharmaceutical
Aerosol Deposition Device on Cell Cultures (PADDOCC) to
evaluate pulmonary drug absorption for metered dose dry powder
formulations. European Journal of Pharmacology and Biopharmacology
77, 132-138.

SCIENTIFIC REPORTS | Publications
169
• Khvedelidze,M., Mdzinarashvili,T., Partskhaladze,T., Nafee,N.,
Schaefer,U.F., Lehr,C.-M., & Schneider,M. (2011) Calorimetric and
spectrophotometric investigation of PLGA nanoparticles and their
complex with DNA. Journal of Thermal Analysis and Calorimetry 99,
337-348.
• Maurer,F., Daum,N., Schaefer,U.F., Lehr,C.-M., & Bauer,P. (2011) Plant
genetic factors for iron homeostasis affect iron bioavailability in
Caco-2 cells. Food Research International 43, 1661-1665.
• Patzelt,A., Richter,H., Knorr,F., Schafer,U., Lehr,C.-M., Dahne,L.,
Sterry,W., & Lademann,J. (2011) Selective follicular targeting by
modification of the particle sizes. Journal of Control Release 150,
45-48.
• Schulze,C., Schaefer,U.F., Ruge,C.A., Wohlleben,W., & Lehr,C.-M.
(2011) Interaction of metal oxide nanoparticles with lung surfactant
protein A. European Journal of Pharmaceutics and Biopharmaceutics
77, 376-383.
Dept. of Drug Design and Optimisation – HIPS |
Prof. Dr. Rolf Hartmann
• Gobbi,S., Zimmer,C.*., Belluti,F., Rampa,A., Hartmann,R.W.*.,
Recanatini,M., & Bisi,A. (2010) Novel highly potent and selective
nonsteroidal aromatase inhibitors: synthesis, biological evaluation
and structure-activity relationships investigation. Journal of
Medicinal Chemistry 53, 5347-5351.
• Haller,F., Moman,E., Hartmann,R.W.*., Adamski,J., & Mindnich,R.
(2010) Molecular framework of steroid/retinoid discrimination in
17beta-hydroxysteroid dehydrogenase type 1 and photoreceptorassociated
retinol dehydrogenase. Journal of Molecular Biology 399,
255-267.
• Heinzerling,L., Hartmann,R.W.*., Frotscher,M., & Neumann,D. (2011)
Predicting putative inhibitors of 17beta-HSD1. Molecular Informatics
29, 695-705.
• Hille,U.E., Zimmer,C., Vock,C.A., & Hartmann,R.W.*. (2011) First
selective CYP11B1 inhibitors for the treatment of cortisol-dependent
diseases. ACS Medicinal Chemistry Letters 2, 2-6.
• Marchais-Oberwinkler,S., Wetzel,M., Ziegler,E., Kruchten,P.,
Werth,R., Henn,C., Hartmann,R.W.*., & Frotscher,M. (2011) New
drug-like hydroxyphenylnaphthol steroidomimetics as potent and
selective 17beta-hydroxysteroid dehydrogenase type 1 inhibitors for
the treatment of estrogen-dependent diseases. Journal of Medicinal
Chemistry 54, 534-547.
• Oster,A., Klein,T., Henn,C., Werth,R., Marchais-Oberwinkler,S.,
Frotscher,M., & Hartmann,R.W.*. (2011) Bicyclic substituted hydroxyphenylmethanone
type inhibitors of 17 beta-hydroxysteroid dehydrogenase
Type 1 (17 beta-HSD1): the role of the bicyclic moiety.
ChemMedChem 6, 476-487.
• Stefanachi,A., Favia,A.D., Nicolotti,O., Leonetti,F., Pisani,L., Catto,M.,
Zimmer,C., Hartmann,R.W.*., & Carotti,A. (2011) Design, Synthesis,
and Biological Evaluation of Imidazolyl Derivatives of 4,7-Disubstituted
Coumarins as Aromatase Inhibitors Selective over 17-alpha-
Hydroxylase/C17-20 Lyase. Journal of Medicinal Chemistry 54, 1613-
1625.
• Wetzel,M., Marchais-Oberwinkler,S., & Hartmann,R.W.*. (2011)
17beta-HSD2 inhibitors for the treatment of osteoporosis: Identification
of a promising scaffold. Bioorganic and Medicinal Chemistry 19,
807-815.
• Yadav,M.R., Sabale,P.M., Giridhar,R., Zimmer,C., Haupenthal,J., &
Hartmann,R.W.*. (2011) Synthesis of some novel androstanes as
potential aromatase inhibitors. Steroids 76, 464-470.
• Hu,Q., Yin,L., Jagusch,C., Hille,U.E., & Hartmann,R.W.*. (2010) Isopropylidene
substitution increases activity and selectivity of biphenylmethylene
4-pyridine type CYP17 inhibitors. Journal of Medicinal
Chemistry 53, 5049-5053.
• Hu,Q., Jagusch,C., Hille,U.E., Haupenthal,J., & Hartmann,R.W.*.
(2010) Replacement of imidazolyl by pyridyl in biphenylmethylenes
results in selective CYP17 and dual CYP17/CYP11B1 inhibitors for
the treatment of prostate cancer. Journal of Medicinal Chemistry 53,
5749-5758.
• Hu,Q., Negri,M., Olgen,S., & Hartmann,R.W. (2010) The role of
fluorine substitution in biphenyl methylene imidazole-type CYP17
inhibitors for the treatment of prostate carcinoma. ChemMedChem 5,
899-910.
• Negri,M., Recanatini,M., & Hartmann,R.W.*. (2010) Insights in
17beta-HSD1 enzyme kinetics and ligand binding by dynamic
motion investigation. PLoS ONE 5, e12026.
• Oster,A., Hinsberger,S., Werth,R., Marchais-Oberwinkler,S.,
Frotscher,M., & Hartmann,R.W.*. (2010) Bicyclic substituted hydroxyphenylmethanones
as novel inhibitors of 17beta-hydroxysteroid
dehydrogenase type 1 (17beta-HSD1) for the treatment of estrogendependent
diseases. Journal of Medicinal Chemistry 53, 8176-8186.
• Oster,A., Klein,T., Werth,R., Kruchten,P., Bey,E., Negri,M., Marchais-
Oberwinkler,S., Frotscher,M., & Hartmann,R.W.*. (2010) Novel
estrone mimetics with high 17beta-HSD1 inhibitory activity. Bioorganic
and Medicinal Chemistry 18, 3494-3505.
• Reum,N., Fink-Straube,C., Klein,T., Hartmann,R.W.*., Lehr,C.-M., &
Schneider,M. (2010) Multilayer coating of gold nanoparticles with
drug-polymer coadsorbates. Langmuir 26, 16901-16908.
• Zimmer,C., Hafner,M., Zender,M., Ammann,D., Hartmann,R.W.*., &
Vock,C.A. (2010) N-(Pyridin-3-yl)benzamides as selective inhibitors
of human aldosterone synthase (CYP11B2). Bioorganic and Medicinal
Chemistry Letters 21, 186-190.

FOCUS RESEARCH REVIEWS SPECIAL FEATURES
Photos from left to right: Kurt Dittmar and Werner Lindenmaier investigate cells for immunotherapy using confocal
laser scanning microscopy. | The BioTechnikum bus of the BMBF during the Biotech-Fair organized by OMNILAB
at the HZI Campus. | During the opening ceremony of the symposium “Networking and Technology Transfer”, Bangkok,
Thailand. In the 1st row (from left) Dr. Peter Winter (InWEnt/GIZ), Prof. Dr. Visith Sitprija (Director of QSMI, Bangkok),
Mr. Stefan Duppel (Deputy Ambassador, German Embassy, Bangkok), in the 2nd row on the left: Dr. Suparp Artjariyasripong
(Deputy Governor of TISTR). (right) Photos: HZI, Scheibe (le) | OMNILAB (ce) | QSMI, Thai Red Cross (ri)

SCIENTIFIC REPORTS
FACTS AND FIGURES

172 FACTS AND FIGURES
Facts and Figures
Prof. Dr. Rainer Jonas | Department of Scientific Information and Internationalization | rjo@helmholtz-hzi.de
In 1965 the HZI was founded as “Centre for Molecular Biological Research” (GMBF) with fi nancial support by the Volkswagen
Foundation. In 1976 the Federal Government through the Ministry for Research and Technology (BMFT) together with the State of
Niedersachsen took over the Centre, now called “German Research Centre for Biotechnology” (GBF). Since then the BMFT/BMBF
as well as the State of Niedersachsen have jointly fi nanced the GBF/HZI. In 2006 it was the fi rst research centre of the Helmholtz
Association to change its name into a Helmholtz Centre institution: the Helmholtz Centre for Infection Research – HZI.
Research Financing In 2010 the total costs of the HZI
amounted to 62.9 Mio. €
Costs per programme (in T€)
Research Area Programme Full Cost
Health
Infection and
Immunity 56 460
Technology Transfer 580
Special Tasks 3 901
Others 1 955
Total Sum 62 896
Full Costs 2010
Infection and Immunity 89.8 %
External Funding for Research More than 75% of the
external funding came from national research programmes.
About 14% and 5% were from EU programmes and industry,
respectively.
External financing of research (in T€)
Source
Full Cost
BMBF 7 041.96
DFG 2 761.78
EU 2 465.56
Industry 898.9
HGF 3 133.37
DAAD 3.33
State of Niedersachsen 128.33
Others 1 105.28
Total Sum 17 538.60
External funding 2010 – by source
BMBF 40.2 %
Special Tasks 6.2 %
Others 3.1 %
Technology Transfer 0.9 %
HGF 17.9 %
DFG 15.8 %
EU 14.1 %
Others 6.3 %
Industry 5.1 %
State of Niedersachsen 0.7 %
DAAD

FACTS AND FIGURES
173
Property Rights / Licences In 2010, six patents were applied
for, five of them outside of Germany.
Patents and property rights, licences, year 2010
Total number Germany
Priority based applications 6 1 5
(2010)
Priority based applications, 102 35 67
total number
Granted patents (2010) 16 1 15
Total number of held 553 46 507
property rights
Licence agreements, 41 32 9
total number
Licence proceeds* (in T€) 631 161 470
* Including revenues from other “know-how” transfer agreements
Publications, Professorships, DFG-Programmes, and
Guest Scientists The HZI has further increased the impact
of its scientific output in recent years. Several articles have
been published in highly renowned journals of the Nature
or Cell Press groups.
Many HZI scientists are participating in important national
and international research programmes.
Participation of HZI scientists in national and international
research programmes
Quantitative Category 2008 2009 2010
Zoonosis / Systemsbiology PBA-Zoo ZooMAP Influenza
Parameters
Publications ISI-listed Papers pub- 205 229 281
lished by the Centre
Peer-reviewed non-ISI 5 11 66
publications in journals
Total number 210 240 347
Habilitations 2 1 4
Dissertations 40 34 40
PhD Students 219 223 291
Full Professor- Calls for professorship 4 7 2
ship Offerings
(W2/W3)
Special DFG- DFG-SFBs, Transregios,
Programmes Excellence clusters 8 8 10
DFG-Research Focus 2 3 3
Graduiertenkollegs 3 4 4
Total number 13 15 17
Guest Scientists 98 87 123
BMBF
AiF-Dechema
Cellular Screening Systems
Basic Innovation Genome Research Candida Therapy
Basic Innovation Genome Research Tuberculosis Therapy
Bioprofile
TransMedLab
ERA-Net
SPATELIS
ERA-Net
METAGUT
ERA-Net
LISTRESS
FORSYS
Metabilic Balance in E. coli
FORSYS
T cell Talk
FORSYS
SysLogics
GenoMik
ProTumor
GenoMik – Transfer
MiPro
GenoMik – Transfer
ExpressSys
GerontoSys
GerontoMitoSys
GerontoSys
GerontoSHIELD
InnoNet
InnoSurf
Innovative Therapies
Armed Bacteria
KMU innovative
De novo – Gencluster-Synthese
KMU innovative
Dis-Z-Konjugate
Medicinal Infection Genomic Nasal Metagenomic
Medicinal Infection Genomic Metagenome and Interactome
Medicinal Infection Genomic LegioProtect
MedSys
Jekyll & Hyde
MedSys
BioInSys
NGFN Plus
German Mouse Clinic
NGFN Plus
Adipositas Network
Excellence Research New States “Taschentuchlabor”
Susceptbility & Resistance against SkinStaph
Infections
Susceptbility & Resistance against PROGRESS
Infections
Susceptbility & Resistance against Resistance Susceptibility
Infections
Susceptbility & Resistance against SkinStaph2
Infections
SysMO
PSYSMO
Zoonosis
Zoo MAP
Zoonosis
PBA-Zoo
Zoonosis
Influenza
Zoonosis
ZooMAP2
Zoonosis Influenza 2
Zoonosis
FBI-Zoo II
Graduate Schools
STREP
Fastest TB
Helmholtz Graduate School for
STREP
PANFLUVAC
Infection Research
HIGS
Helmholtz-Kolleg for Infection Biology H-IRISIB
DFG-Graduate School GRK 653
“Pseudomonas”
DFG-Graduate School GRK1273
“Chronical Infections”

174 FACTS AND FIGURES
EU Frame Programmes
CA
CASIMIR
COST
Sysgenet
CP
FAST-XDR-DETECT
CP
FLUINHIBIT
CP
BACSIN
CP
HEPTROMIC
CP
OPTISTEM
CP
MagicPAH
CP
CAREPNEUMO
CSA
TARPOL
ERC StG RESISTOME
ERC StG CMVAgSTIMULUS
IMI
EMTRAIN
Infrastructures
Infrastructures
Infrastructures
Infrastructures
Infrastructures
Infrastructures
IP
Marie Curie IOF
NoE
NoE
SME driven
EATRIS
Infrafrontier
ProteomeBinders
Instruct – Associated
TRANSVAC
EU-Openscreen
EUMODIC
APPI
EuroPathoGenomics EPG
Clinigene
NOPERSIST
STREP
Healthy-Water
STREP
ASSIST
STREP
Fastest TB
STREP PANFLUVAC Intellectual Property Since 2002 the Ascenion Ltd. Co.
offers services principally for the four Helmholtz Research
Centres in the area of health care: GSF, HZI, MDC, and
Technology Transfer The HZI has a great potential for the
development of innovative products, processes and services,
especially in cooperation with industrial partners. Therefore,
an important goal is to foster the transfer of research
results into industrial applications through technology
transfer. Thus, the establishment of spin-off and start-up
biotech companies, licence agreements as well as service
contracts with industrial partners are important elements
for the transfer of R&D results. In order to further support
technology transfer activities, the HZI is a member of
BioRegioN and the “Transferkolleg Biotechnologie e.V.”.
DFG (German Research Foundation)
SFB 566 Cytokin-Receptors
SFB 578 From Gene to Product
SFB 587 Lung Immunity
SFB 599 Permanent Implantates
SFB 621 Pathobiology of the Intestinal Mucosa
SFB 738 Optimization of Conventional and Innovative
Transplants
SFB 854 Molecular Organisation Cellular
Communication in the Immune System
SFB 900
SFB/TR 51
SPP 1258
SPP 1316
SPP 1394
EXC 62
FOR 629
FOR 1220
FOR 1406
KFO 250
Chronic Infections: Microbial Persistence
and its Control
Ecology, Physiology and Molecular Biology
of the Roseobacter-Group
Sensory and regulatory RNAs in Prokaryotes
Host-adapted Metabolism of Bacterial
Pathogens
Mast-cells – Promoters of Hhealth and
Modulators of Disease
ReBirth
Antibodies and Proteinanalysis
PROTRAIN
Exploiting the Potential of Natural
Compounds: Myxobacteria
Genetic and Cellular Mechanisms of
Autoimmune Diseases
DKFZ. The headquarters are in Munich, but an office with
two employees works on the HZI-Campus.
Ascenion Ltd. Co. manages the following areas for the HZI:
• Acquisition and management of intellectual property
• Evaluation of the commercial potential of an invention
before patent fi ling
• Development and employment of strategies for the
exploitation of the HZI patent folio
The HZI-Biotech Campus A new container for the administration
was ready in 2009, and also a new S3-laboratory
is ready for use. The laboratory building D will get a new
facility for office work, which is under construction and
should be ready at the end of 2011.
Biotech Fair on the HZI Campus On September 16, 2010
OMNILAB organized, with the support of HZI and DSMZ,
its 6 th biotech-fair and symposium in the FORUM. It was the
celebration of its 10 th anniversary and the 75 th anniversary
of OMNILAB. About 50 enterprises and several research
institutions of the region presented themselves to about
600 visitors, a new record. The visitors came from the
Braunschweig-Hannover-Magdeburg-Göttingen region. A
special highlight was the BIOTechnikum of the BMBF. The
next fair is foreseen to take place at the HZI facilities on
September 20, 2012.

FACTS AND FIGURES
175
Open Day at HZI On Saturday, May 08, 2010, HZI opened
its doors for everybody to have a look at the activities of the
infection research centre. More than 1,800 visitors from the
region came to get to know more details about infections,
vaccines, biofilms, etc.
A scientific presentation during the Biotech Fair organized by
OMNILAB at the HZI FORUM in September 2010 Photo: OMNILAB
Biotech Network ASEAN-MERCOSUL – Germany The
internet platform of the ASEAN-South American-German
Biotechnetwork (www.asag-biotech.net) was further improved.
Now one can search for most of German institutes that
are working in the biotech area, sensu latu. Furthermore,
several institutes from Latin American countries as well as
from ASEAN countries are listed. Also biotech-industries
and events can be searched for. The platform allows direct
acces to the most important institutions in Germany and
other countries, to organisations that offer fellowships,
calls, and possibilities of networking. The publication
databasis of the Helmholtz Institutions, several patent data
basis, some university libraries can be accessed easily. In
June 2010, HZI together with InWEnt and other partners
organized a big symposium on “Networking and Technology
Transfer”, which took place in Bangkok, Thailand. More
than 150 scientists from Germany, Southeast Asian and
Mercosur countries participated.
Dr. Torsten Lührs explains visitors during the Open Day 2010
how a NMR spectrometer is working. Photo: HZI
Crèche and Kindergarten for Children of HZI Employees
The HZI in cooperation with the Sterntaler Kindergarten in
Braunschweig-Stöckheim offers childcare for those aged one
year and older. In 2010, 19 children were admitted to the
kindergarten, 5 of them in the crèche and the others in the
kindergarten.
HGF-Mentoring Programme 2010/11 In 2010 three
female employees participated in the programme, and it is
expected that another five will participate in 2011.
Audit “berufundfamilie” On November 27, 2010 the HZI
was certificated again for further 3 years for that programme.
Future Day for girls and boys In 2011 nearly 100 pupils
between 12 and 16 from various types of schools visited
the HZI at the Future Day. Fifteen different work groups
participated and about 60 staff members showed the pupils
interesting experiments and/or gave them ideas of the work
at the HZI.
Prof. Dr. Yongyuth Yuthavong, former Minister of Science and
Technology of Thailand and Professor at NSTDA, Thailand,
during his keynote speech on “Aspects of the future development
of biotechnology” at the symposium “Networking and
Technology Transfer” held in Bangkok in June 2010.
(Photo: QSMI, Thai Red Cross).
Girls and boys are very curious to learn more about what HZI
is doing in its research. Photo: HZI

176 FACTS AND FIGURES
Personnel At the end of 2010 the HZI staff comprised 731
persons with full time and part time occupation. Additionally,
251 guests worked in various projects, receiving
their payment from third parties. Along with 188 senior
scientists, almost 300 PhD-students, and 22 engineers were
working at the HZI.
Boards and Assemblies of the HZI The boards and
assemblies of the HZI are the Board of Trustees, the Supervisory
Board, the Scientific Committee and the Managing
Directors.
Board of Trustees The Board of Trustees is formed by the
three trustees of the HZI, the Federal Republic of Germany,
the State of Niedersachsen, and the State of Saarland,
represented by their respective departments, the Federal
Ministry of Education and Research (BMBF), the Finance
Ministry of Niedersachsen, and the Ministry of Economics
of the Saarland.
Supervisory Board The Supervisory Board (SB) oversees
the legality, expedience and economy of the management.
It decides on general research goals, the principal research
policy and financial affairs of the centre. It consists of a
maximum of 11 members.
Scientific Advisory Committee The Scientific Advisory
Committee (SC) consists of external scientific experts. It
advises the Supervisory Board with regard to the R&D programme
as well as the general research strategy of the HZI.
Managing Directors The Managing Directors of the HZI:
Research & Development: Prof. Dr. Dirk Heinz (acting)
Administration: Ulf Richter, MBA
The Scientific and Adminstrative Directors of the HZI,
Prof. Dr. Dirk Heinz (right) and Ulf Richter (left). Photo: HZI
Scientists Council The Scientists Council of the HZI
advises the Management in scientific matters. It consists by
one half of the heads of the departments and by the other
half of elected research group leaders, project leaders or
junior research group leaders. The Managing Directors,
the members of the Steering Committee, one member of
the work council and one member of the administration,
appointed by the Administrative Director, are guests of the
assembly.
Steering Committee The Steering Committee advises the
Managing Directors of the HZI in all important questions
of the Centre. Members are the Managing Directors, the
Programme Speaker, and the Topic Speakers.
Staff Council The Staff Council has certain consultation
and co-determination rights in personnel and social questions.
It consists of 11 members, elected by the HZI staff.
Chairman is John Aubert.
Equal Opportunities Officer is Evelyn Rohn-Stenzel.

FACTS AND FIGURES
177
Members of the Supervisory Board (SB) and the Scientific Advisory Committee (SC), Status: 19.01.2011
Function Name, Title Organisation Locality
Chairman SB Brumme-Bothe, MinDir’in Bärbel BMBF Berlin
Vice-Chairman SB Gevers, MinDirig Dr. Heiko NMWK Hannover
SB Köpke, OR Heinz-Hermann Ministry of Finances, Hannover
Niedersachsen
SB + SC Baum, Prof. Dr. Christopher MHH Hannover
SB Dersch, Prof. Dr. Petra HZI Braunschweig
SB Weiß, Dr. Siegfried HZI Braunschweig
SB + SC Zettlmeissl, Dr. Gerd Intercell AG Wien
SB + SC Müller-Goymann, Prof. Dr. Christel TU Braunschweig
SB + SC Vice-Chairman SC Schendel, Prof. Dr. Dolores HMGU München
SB + SC Kurth, Dr. Bärbel-Maria Robert-Koch-Institut Berlin
SB + SC Daniel, Prof. Dr. Hannelore Wissenschaftszentrum Freising
Weihenstephan
SB + SC Chairman SC Pfeffer, Prof. Dr. med. Klaus Universitätsklinikum Düsseldorf
SC Hacker, Prof. Dr. Jörg Nationale Akademie Halle
der Wissenschaften
SC Rosenthal, Prof. Dr. Walter MDC Berlin
SC Brakhage, Prof. Dr. Axel HKI Jena
SC Apweiler, Dr. Rolf EBI Cambridge
SC Wilmanns, Dr. Matthias EMBL Hamburg
SC Hakenbeck, Prof. Dr. Regine TU Kaiserslautern
SC Hämmerling, Prof. Dr. Günter DKFZ Heidelberg
SC Di Santo, Prof. Dr. James Institut Pasteur Paris

178 FACTS AND FIGURES FACTS AND FIGURES 179
Chart of Organisation, Status February 01, 2011
Scientific Advisory Commitee (SC)
Prof. Dr. med. K. Pfeffer, Chair
Supervisory Board (SB)
MinDir‘in B. Brumme-Bothe, Chair
MinDirig H. Gevers, Nieders. MWK, Vice Chair
Executive Board (DR)
Staff Council (BR)
J. Aubert, Chair
Representative Body for
Disabled Employees
B. Dojka (VPS)
Scientific Director (GFW) Prof. Dr. D. Heinz (acting)
Administrative Director (GFA) U. Richter
Assembly of
Scientists (WV)
Dr. W.-R. Abraham, Chair
Equal Opportunity
Commissioner (GB)
E. Rohn-Stenzel
Dept. of Molecular
Infection Biology (MIBI)
Prof. Dr. Petra Dersch
Dept. of Gene Regulation
and Differentiation (RDIF)
Dr. H. Hauser
Dept. of Molecular
Structural Biology (MOSB)
Prof. Dr. D. Heinz
Dept. of Medical
Microbiology (MMIK)
Prof. Dr. G.S. Chhatwal
Dept. of Infection
Genetics (INFG)
Prof. Dr. K. Schughart
Dept. of Medicinal
Chemistry (MCH)
Prof. Dr. M. Kalesse
Infrastructure
RG Chronic Pseudomonas
Infections (CPI)
Prof. Dr. S. Häußler
RG Molecular
Immuno logy (MOLI)
Dr. S. Weiß
RG Biophysical
Analysis (BA)
Dr. V. Wray
RG Infection
Immunology (INI)
PD Dr. E. Medina
RG Experimental
Animal Unit (TEE)
Dr. H. Riedesel
RG Microbial Drugs
(MWIS)
Prof. Dr. R. Müller
Administration and
Infrastructure (VIN)
U. Richter
Staff Units
U. Richter /
Prof. Dr. D. Heinz
RG Microbial Communication
(KOM)
Prof. Dr. I. Wagner-Döbler
RG Model Systems for
Infection and Immunity
(MSYS) Dr. D. Wirth
RG Recombinant Protein
Expression (RPEX)
Dr. J. van den Heuvel
RG Microbial Interactions
and Processes (MINP)
Prof. Dr. D. Pieper
Dept. of Immune Control
(IMMK)
Prof. Dr. B. Schraven
RG Environmental
Microbiology (UWM)
Prof. Dr. K. Timmis
Human Resources
(PA)
J. Schinkel
Public Relations (ÖA)
M. Braun
RG Genom Analytics
(GMAK)
Dr. R. Geffers
RG Cellular Proteom
Research (CPRO)
Dr. L. Jänsch
Dept. of Vaccinology and
Applied Microbiology (VAC)
Prof. Dr. Dr. C.A. Guzmán
RG System-oriented Immunology
and Inflammation Research
(SIME) Prof. Dr. I. Schmitz
Finance Department
(FA)
D.-M. Reinhardt
Controlling (CO)
- Dr. M. Strätz (DMC) (Deputy)
- NN (BCO)
- Dr. R.-J. Müller (PWC)
JRG Chronic Infections
and Cancer (CHIK)
Prof. Dr. L. Zender
RG Microbial Proteomics
(MPRO)
Prof. Dr. K. Riedel
RG Chemical Microbiology
(CMIK)
Dr. W.-R. Abraham
Dept. of Experimental
Immunology (EXIM)
Prof. Dr. J. Hühn
Purchasing
Department (EM)
B. Balster
Internal Auditing (IR)
R. Lomberg – Anti-corruption-commissioner
JRG Macromolecular
Interactions (MMIA)
Prof. Dr. C. Ritter
JRG Phagosome Biology
(PHAB)
Dr. M. Gutierrez
RG Immunoregulation
(IREG)
PD Dr. D. Bruder
HIPS
Legal Affairs (JUR)
Dr. C. Kügler-
Walkemeyer
Occupational Safety
Specialist (FaSi)
Dr. E. Grund
Scientific Groups
JRG Structure-based
Infection Biology (SBIB)
Dr. T. Lührs
JRG at Centre for Structural
Systems Biology –
DESY, Prof. Dr. I. Kursula
JRG Immune Aging and
Chronic Infections (IMCI)
Dr. Dr. L. Cicin-Sain
JRG Viral Immune
Modulation (VIMM)
Dr. M. Brinkmann
Dept. of Systems
Immuno logy (SIMM)
Prof. Dr. M. Meyer-Hermann
Dept. of Microbial Natural
Products (MINS)
Prof. Dr. R. Müller
JRG Actinobacteria Metabolic
Engineering Group
(AMEG) Dr. A. Luzhetskyy
Patents (PS)
D. Meseke
Technical
Services (TB)
O. Rabe
Scientifi c Information &
Internationalization (WI)
Prof. Dr. R. Jonas
Library (BIB)
A. Plähn
Abbreviations:
Dept Department
RG Research Group
JRG Junior Research Group
PWC Scientific Controlling
BCO Financial Controlling
DMC External Funding Controlling
DSB Data Security
HIPS Helmholtz-Institute for
Pharma ceutical Research Saarland,
Saarbrücken
CIO Chief Information Officer
Dept. of Chemical
Biology (CBIO)
Dr. F. Sasse (acting head)
RG Biologic Systems
Analysis (BISA) Prof. Dr.
U. Bilitewski (acting head)
Dept. of Drug Delivery
(DDEL)
Prof. Dr. C.-M. Lehr
Dept. of Drug Design and
Optimization (DDOP)
Prof. Dr. R. Hartmann
Computer Centre
(RZ)
Dr. N. Bedorf (CIO)
Safety and Environmental
Affairs (SU)
Dr. E. Grund
Organisation/Administration-IT
(ORG)
H. Ohrdorf - DSB

180
IMPRINT
Research Report 2010/11
Published by
Helmholtz Centre for Infection Research GmbH (HZI)
Inhoffenstraße 7
D-38124 Braunschweig, Germany
Telephone +49 (0)531-61 81-0
Telefax +49 (0)531-61 81-2655
info@helmholtz-hzi.de | www.helmholtz-hzi.de
Member of
Helmholtz Association of National Research Centres
Editor-in-Chief:
Prof. Dr. Rainer Jonas (V.i.S.d.P.) | rjo@helmholtz-hzi.de
Editoral Assistance: Monica Kirchner | Dr. Bastian Dornbach | Dr. Clemens Ostrowicz
Text Editing and Realisation: Dr. Jo Schilling | buero@jo-schilling.de
Translations: ADREM Sprachdienstleistungen sowie Ronning Übersetzungsdienst (Twincore)
© 2011 HZI Braunschweig
Photographs
The portraits and further photos were taken by
Bierstedt – pages: 48, 71, 80, 90, 91, 98, 106, 111, 113, 127-129, 172
Dornbach – pages: 30, 35, 39, 118
Gramann – pages: 7, 9, 19, 22, 28, 42, 44, 64, 69, 73, 74, 77, 84, 97, 102, 103, 108, 110, 119,
132, 138, 175, 176
Hans – pages: 79, 86, 89
Hübner – pages: 114, 175
Krämer – pages: 15, 30, 34, 40, 74, 76, 85, 88, 94-96, 99, 104, 112, 115, 117, 126
Sondermann – page: 5
HZI collection – pages: 2, 52, 59, 60, 67, 70, 72, 78, 81, 87, 93, 105, 107, 112, 122, 124, 130, 131
HIPS collection – pages: U2, U4, 2, 36, 39, 40, 125
Twincore collection – page: U1, U2, 2
The following portraits were made available by the authors – pages: 28 (Ciesek), 28 (Steinmann),
40 (Schäfer), 75 (Dersch), 92 (Hühn), 105 (Zender), 116 (Meyer-Hermann), 120 (Riedel), 123 (Kursula)
For all other photographs, the authors and/or institutions are mentioned below the legends.
Layout and design:
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ISSN 1865-9713

Celle
Gifhorn
Lüneburg
B 4
Hannover
Dortmund
BS-Hafen
B 214
BS-Flughafen
Kreuz
BS-Nord
A 2
Berlin
A 391
A 392
B 4
Kreuz Ölper
B 248
B 1
A 391
Braunschweig
Peine
Hildesheim
Hauptbahnhof
B 1
Königslutter
Helmstedt
B 1
Dreieck
BS-Südwest
Kreuz
BS-Süd
A 39
Wolfsburg
A 39
Abfahrt
BS-Mascherode/
Stöckheim
Salzgitter
Göttingen
Kassel
B 248
A 395
Salzgitter-Bad
Wolfenbüttel
Bad Harzburg
Site plan of the Helmholtz Centre for Infection Research

ISSN 1865-9713
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