VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

[2] Wennerhold, J. et al (2005): The AraC-type regulator RipA represses aconitase and other ironproteins from Corynebacterium under iron limitation and is itself repressed by DtxR. J Biol Chem,280(49):40500-40508.[3] Krug, A. et al (2005): Identification of AcnR, a TetR-type repressor of the aconitase gene acn inCorynebacterium glutamicum. J Biol Chem, 280(1):585-595.[4] Emer, D. et al (2009): Complex expression control of the Corynebacterium glutamicum aconitasegene: Identification of RamA as a third transcriptional regulator besides AcnR and RipA. JBiotechnol, 140(1-2):92-98.[5] Han, S.O. et al (2008): Effect of carbon source availability and growth phase on expression ofCorynebacterium glutamicum genes involved in the tricarboxylic acid cycle and glyoxylate bypass.Microbiology, 154:3073-3083.SRP008Anisin1, a defensin-like protein in Aspergillus nidulans,senses oxidative stress and balances asexual developmentA. Eigentler* 1 , I. Pócsi 2 , F. Marx-Ladurner 11 Innsbruck Biocenter, Division of Molecular Biology, Innsbruck MedicalUniversity, Innsbruck, Austria2 Faculty of Science and Technology, Department of MicrobialBiotechnology and Cell Biology, University of Debrecen, Debrecen,HungaryIn the genome of A. nidulans a defensin-like protein, Anisin1, was annotatedthat exhibits 51% amino acid identity to the mosquito Aedes (Ae.) aegyptidefensin AaDefA1. Although defensins are widely distributed in nature andtheir function in higher eukaryotes is well characterized, no studies exist sofar on defensins that originate from filamentous ascomycetes. We, therefore,started to characterize the Anisin1 encoding gene in A. nidulans. Expressionstudies in submers cultures indicated that elevated levels of intracellularreactive oxygen species (ROS) triggered the anisin1 expression. We usedspecific mutants of the histidine-to-aspartate signal transduction pathway toshow that anisin1 expression was strongly induced in DsrrA, which suffersfrom a defect to detoxify ROS. In contrast, anisin1 was repressed in A.nidulans strains that efficiently respond to oxidative stress. In A. nidulanswild-type surface cultures, however, the anisin1 transcription correlated withthat of the central regulator for asexual development, brlA, and with catB.This co-regulation was deregulated in DsrrA which might explain thesporulation defect in this mutant. The phenotype analysis of an anisin1deletion mutant revealed an increased oxidative stress sensitivity, a defect inmitospore development and lower conidial counts at 42°C compared to thewild-type. Taken together, our results suggest that in A. nidulans anisin1plays an important role in sensing oxidative stress, in balancingconidiogenesis and in supporting thermotolerance during asexualdevelopment. In analogy to a multiple function of defensins in highereukaryotes, Anisin1 could therefore contribute to the fitness of A. nidulansunder unfavourable growth conditions.SRP009Repair potential in natural drinking water biofilms afterwater treatmentC. Jungfer*, J. Varela Villarreal, K. Brändle, U. Obst, T. SchwartzInstitute of Functional Interphases (IFG), Karlsruhe Institute of Technology(KIT), Eggenstein-Leopoldshafen, GermanyBiofilms are present in drinking water distribution systems despite watertreatment and disinfection at waterworks. They are a possible niche forhygienic relevant bacteria, and therefore a main concern for water industries.Up to now knowledge about survival strategies of bacteria during theirregeneration process in biofilms after disinfection treatments has beenlimited. To get a deeper understanding of this problem, biofilms of groundwater and surface water were investigated in different waterworks.In each waterworks the same pilot scale, built up with different standardpipe materials, was used to simulate a household water distribution system.The water that flowed through the pilot scale was exposed to disinfectionmethods such as UV and chlorine dioxide. Three month old biofilms werecompared using RNA and DNA based methods.When stress markers on RNA level were investigated, UV disinfection wasfound to be responsible for the up-regulation of recA-mediated dark repair innatural biofilms. The highest recA induction in biofilms was associated withcopper, confirming previous investigations from other waterworks. No oronly low recA expression was found in biofilms gained from the waterworksin which drinking water was not disinfected or treated with ClO 2.The total amount of bacteria present in the biofilms did not depend on thedifferent materials or disinfection processes. But DGGE analysis showed asignificant shift in the bacterial population when different materials anddisinfection treatments were used, showing e.g. an interesting speciesselection when grown on copper.SRP010Analysis of antimicrobial peptides, their use for biofilmprotection and the general stress response in fungiB. Gutt* 1 , J. Zoller 1 , A. Rieder 2 , T. Schwartz 2 , C. Bollschweiler 3 , R. Fischer 11 Institute for Applied Biosciences, Karlsruhe Institue of Technology (KIT),Karlsruhe, Germany2 Institute of Functional Interfaces, Karlsruhe Institute of Technology (KIT),Karlsruhe, Germany3 BASF SE, Ludwigshafen, GermanyMicrobial biofilms are ubiquitously found and the chosen life form of mostof the microorganisms. Bacteria, fungi but also algae and protozoa benefitsymbiotically from each other, while they are surrounded by an extracellularmatrix. Some of the biofilms are unwanted, as they cause disease or destroytechnical systems. To prevent biofilm formation on surfaces, we constructedmodified fungal hydrophobins fused to antimicrobial peptides.Hydrophobins are small proteins, which self assemble at any hydrophilichydrophobicinterface into extremely stable amphipathic monolayers.Cationic antimicrobial peptides (AMPs) are only 9 to 50 amino acids in sizeand are proven to be active against bacteria and even against yeasts andfilamentous fungi [1]. They are an alternative to antibiotics and do not affecthuman cells. Different cationic antimicrobial peptides (AMPs) were fused toA. Nidulans hydrophobin DewA and expressed in E. coli.A second aspect is the analysis of the general stress response in A. nidulanswhen these fungi are exposed to those AMPs. To this end, we are currentlyanalyzing different His-Asp phosphorelay signaling systems (also known astwo component systems), which consist of response regulators (rr) andhistidine kinases (hk). A. nidulans has 4 rrs and 15 hks [2], for all of whichwe have corresponding deletion strains, which will be analyzed for theirsensitivity against the cationic antimicrobial peptides.[1] D. Mania et al (2010): Screening for Antifungal Peptides and Their Modes of Action inAspergillus nidulans. Appl. Environ. Microbiol. 76: 7102-7108.[2] D. Hagiwara et al (2007): The SskA and SrrA Response Regulators Are Implicated in OxidativeStress Responses of Hyphae and Asexual Spores in the Phosphorelay Signaling Network ofAspergillus nidulans. Biosci. Biotechnol. Biochem. 71: 1003-1014.SRP011Iron-binding properties of the redox sensor protein HbpSof the three-component system HbpS-SenS-SenR fromStreptomyces reticuliI. Wedderhoff*, H. Schrempf, D. Ortiz de Orué LucanaApplied Genetics of Microorganisms, University of Osnabrueck,Osnabrueck, GermanyThe novel three-component signalling system HbpS-SenS-SenR from thecellulose degrader Streptomyces reticuli has been reported as an example ofa redox sensing pathway in bacteria. This system senses redox stress signalsin form of toxic concentrations of hemin, iron ions or other redox-activecompounds and regulates genes involved in oxidative stress response [1].During these processes, the extracellular oligomer-forming protein HbpSacts in concert with the two-component system SenS-SenR. Moreover,heme-binding and heme-degradation as well as iron-binding properties ofHbpS have been shown to play an important role in the signalling cascade[2,3].Recently, iron-binding motifs (D/E-X-X-E) have been identified in HbpS[3]. In order to get more insights as to their functional role, a set of mutantproteins will be generated and analysed in vitro and in vivo. Furthermore, byusing of the 3D crystal structure of HbpS the localization of these motifswill be addressed. Homologues to HbpS exist in a number of Gram-positiveand Gram-negative bacteria. Based on the structure of HbpS, modelling oftheir structure will be pursued. The obtained results will be discussed inframe of this presentation.[1] Ortiz de Orué Lucana, D. and M.R. Groves (2009): The three-component signalling system HbpS-SenS-SenR as an example of a redox sensing pathway in bacteria. Amino Acids, 37:479-486.[2] Ortiz de Orué Lucana, D. et al (2009): The oligomeric assembly of the novel haem degradingprotein HbpS is essential for interaction with its cognate two-component sensor kinase. J Mol Biol,386: 1108-1122.[3] Ortiz de Orué Lucana, D. et al (2010): Iron-mediated oxidation induces conformational changeswithin the redox-sensing protein HbpS. J Biol Chem, 285: 28086-28096.spektrum | Tagungsband 2011