ZMBH J.Bericht 2000 - Zentrum für Molekulare Biologie der ...

with low levels of the enzyme were highly susceptible
to oxidative stress.
A variety of drugs is excreted from cancer cells in the
form of glutathione-drug conjugates by ABC transporters
of the MRP (multidrug resistance protein)
family. In some Leishmania species, arsenite and antimonial
drugs can be sequestered as conjugates with
trypanothione. The only drugs available against latestage
trypanosomiasis in east Africa are arsenical
derivatives, but resistance to these drugs is emerging.
To find out if MRP-like trypanothione-arsenical conjugate
pumps might play a role in resistance, we have
cloned and sequenced trypanosome homologues of
two Leishmania MRP-like genes. Cells over-expressing
the transporters will be characterised with respect
to drug sensitivity and other characteristics.
Metabolic compartmentation in Toxoplasma
Martina Ding, Frank Voncken
D. Soldati (ZMBH) and I became intrigued by the
complete absence of reports of peroxisomes in malaria
parasites and related organisms such as Toxoplasma. A
partial sequence for catalase (a peroxisomal marker)
was identified in the Toxoplasma sequence database.
The predicted protein has all concensus amino acids
for enzyme activity, and a C-terminal peroxisomal
targeting signal. To our surprise, immunofluorescent
labelling of Toxoplasma with antibodies to two different
anti-peptide antibodies gave predominantly cytosolic
labelling, and the addition of the targeting signal
to the green flourescent protein also resulted in cytosolic
flourescence. Cell fractionations gave similar
results. Thus so far we have no evidence that Toxoplasma
contains peroxisomes.
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Collaborations
Dr. Luise Krauth-Siegel (Biochemie-Zentrum, Heidelberg)
– trypanothione reductase.
Dr. Dietmar Steverding (Parasitologie, Heidelberg) –
transferrin receptor.
Dr. Martina Bremser and Prof. F. Wieland (Biochemie-Zentrum,
Heidelberg) - coatomer.
Dr. Dominique Soldati (ZMBH) – Toxoplasma peroxisomes
Drs. Jürgen Benting and Kai Simons, EMBL (rafts)
Dr. Paul Michels (Institute for Cellular Pathology,
Brussels, Belgium) - glycosomes and glycolysis.
Prof. Kenneth Stuart (Seattle Biomedical Research
Institute, Washington, Seattle) and Prof. Ullrich
Goeringer (Universität Darmstadt) – inducible knockouts
of genes involved in RNA editing.
Dr. Roland Kaminsky and Dr. Pascal Maser (Tropeninstitut,
Basel) - drug resistance.
External Funding
During the period reported our research was supported
by grants from the Deutsche Forschungsgemeinschaft
(SFB 352 “Molekulare Mechanismen intrazellulärer
Transportprozesse”, SFB 544 “Kontrolle tropischer
Infektionskrankheiten”, Graduiertenkolleg “Kontrolle
der Genexpression in pathogenen Mikroorganismen”
and Projekt-’Sachbeihilfen’) from the Human Frontier
Science Programme Organization, the EMBO, the
DAAD, the BMBF (Forschungsschwerpunkt Tropenmedizin)
and from the Fonds der Chemischen Industrie.
PUBLICATIONS
Original articles
Ansorge, I., Steverding, D., Melville, S., Hartmann,
C., and Clayton, C. (1999). Transcription of ‘inactive’
expression sites in African trypanosomes leads
to expression of multiple transferrin receptor RNAs
in bloodstream forms. Mol. Biochem. Parasitol. 101,
81-94.
Benting, J., Rietveld, A., Ansorge, I., and Simons, K.
(1999). Acyl and alkyl chain length of GPI anchors is
critical for raft association in vitro. FEBS Lett. 462,
47-50.
Blattner, J., Helfert, S., Michels, P., and Clayton,
C. E. (1998). Compartmentation of phosphoglycerate
kinase in Trypanosoma brucei plays a critical role
in parasite energy metabolism. Proc. Natl. Acad. Sci.
USA 95, 11596-11600.
Clayton, C.E., Ha, S., Rusche, L., Hartmann, C.,
Beverley, S. M. (2000). Tests of heterologous promoters
and intergenic regions in Leishmania major. Mol.
Biochem. Parasit. 105, 163-167.
Ding, M., Clayton, C.E. and Soldati, D. (2000). Toxoplasma
gondii catalase: are there peroxisomes in Toxoplasma?
. J. Cell Sci. 105, (in press).
Drozdz, M., and Clayton, C. E. (1999). Confirmation
of a regulatory 3‘-untranslated region from Trypanosoma
brucei. RNA 5, 1632-1644.
Hartmann, C., Hotz, H.-R., McAndrew, M., and Clayton,
C. (1998). Effect of multiple downstream splice
sites on polyadenylation in Trypanosoma brucei. Mol.
Biochem. Parasitol. 93, 149-152.
Hotz, H.-R., Biebinger, S., Flaspohler, J., and Clayton,
C. E. (1998). PARP gene expression: regulation at
many levels. Mol. Biochem. Parasitol. 91, 131-143.
Krieger, S., Schwarz, W., Ariyanagam, M.R., Fairlamb,
A., Krauth-Siegel, L., and Clayton, C. E. (2000).
Trypanosomes lacking trypanothione reductase are
avirulent and show increased sensitivity to oxidative
stress. Mol. Microbiol. 35, 542-552.
Lorenz, P., Meier, A., Erdmann, R., Baumgart, E., and
Clayton, C. (1998). Elongation and clustering of glycosomes
in Trypanosoma brucei overexpressing the
glycosomal Pex11p. EMBO J. 17, 3542-3555.
McAndrew, M., Graham, S. and Clayton, C.E. (1998).
Testing promoter activity in the trypanosome genome:
isolation of a metacyclic-type VSG promoter, and
unexpected insights into RNA polymerase II transcription.
Exp. Parasitol. 90, 65-76.
Reviews
Hotz, H.-R., Biebinger, S., Flaspohler, J., and Clayton,C.E.
(1998). PARP Gene expression: regulation at
many levels. Francqui Symposium Proceedings and
Mol. Biochem. Parasitol. 91, 131-143.
Clayton, C. E., et al. (1998). Genetic nomenclature for
Trypanosoma and Leishmania. Mol. Biochem. Parasitol.
97, 221-224.
Clayton, C. E. (1999). Genetic manipulation of Kinetoplastida.
Parasitol. Today 15, 372-378.
Roditi, I., and Clayton, C. E. (1999). An unambiguous
nomenclature for the major surface glycoprotein
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