Analyzing the role of a cellular mechanism in HIV infection and evaluation of novel antiviral compounds [Elektronische Ressource] / vorgelegt von Sylvia Gabriele Münter

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• Gesundheit

Informations

• Publié par: ruprecht-karls-universitat_heidelberg
• Publié le: 01 janvier 2004
• Nombre de lectures: 14
• Langue: English
• Poids de l'ouvrage: 2 Mo

Extrait

INAUGURAL DISSERTATION/ THESE
zur
Erlangung der Doktorwürden
der
Naturwissenschaftlichen-Mathematischen Gesamtfakultät
der
Présentée pour obtenir
Le GRADE de DOCTEUR EN SCIENCES
de
RUPRECHT-KARLS-UNIVERSITÄT HEIDELBERG
und/ et
L’UNIVERSITE PARIS XI ORSAY
vorgelegt von/ par
Diplom-Biochemikerin Sylvia Gabriele Münter
aus/ née à Heidelberg
Tag der Disputation/ Date de la Soutenance: 20.07.2004THEMA/ SUJECT:
Analyzing the role of a cellular mechanism in HIV infection
and
Evaluation of novel antiviral compounds
Prüfungskommission/ Commission d’examen:
Prof. Dr. Hermann Irngartinger
Prof. Michel Jacquet
Prof. Dr. Dr. Jürgen Debus Gutachter/ Rapporteur
Dr. Ulf NehrbassPublication list:
Knoblauch H, Busjahn A, Münter S, Nagy Z, Faulhaber HD, Schuster H, Luft FC.
(1997); Heritability analysis of lipids and three gene loci in twins link the macrophage
scavenger receptor to HDL cholesterol concentrations. Arterioscler Thromb Vasc
Biol. 1997 Oct;17(10):2054-60.
Bajramovic JJ, Münter S, Syan S, Nehrbass U, Brahic M, Gonzalez-Dunia D. (2003);
Borna disease virus glycoprotein is required for viral dissemination in neurons. J
Virol. 2003 Nov;77(22):12222-31.
Roux P, Münter S, Frischknecht F, Herbomel P, Shorte SL. (2004); Focussing light
on infection in four dimensions. Cell Micobiol. 2004 Apr;6(4):333-43. Review.
Münter S, Vazquez-Martinez R, Matunis MJ, David-Watine B, Nehrbass U, Shorte
SL. Nuclear envelope dynamics are driven by high turnover actin polymerization
(submitted).
Presentations at international meetings:
29.08. -04.09.2002 EMBO YIP PhD course Heidelberg: Poster presentation
and oral presentation.
18.06.2003 First Congress of young researchers at the Pasteur
Institute: Oral presentation.
13-17.12.2003 ASCB 2003 Annual Meeting San Francisco:
Poster presentation.Preamble
First of all, I would like to thank Ulf Nehrbass and Jürgen Debus having accompanied
me during my doctoral thesis and for the help and inspiring discussions during the last
four years.
Furthermore, I thank Professor Irngartinger for accepting me as a student under his
supervision in the Department of Chemistry at the University of Heidelberg. Also, I
appreciated a lot his help and advice concerning the chemical syntheses I completed
in his laboratory.
I would like to express my thankfulness to Professor M. Jacquet having accepted to be
part of the evaluation committee of my thesis in the cotutelle program between Paris
and Heidelberg.
Especially, I want to thank Spencer Shorte for the ongoing help and the knowledge I
achieved in fluorescent microscopy; moreover for the excellent supervision related to
important constituents of my thesis and many motivating scientific conversations.
I thank all the members in the BCN lab for their encouragement during my work, the
fantastic working atmosphere (even if sometimes a bit to enthusiastic about the
sound) and the innumerable relaxing moments we spent in the lab … and (just to
name one) in the “Breguet”; Mr. Pubmed and the better half of the “genius couple of
science” for critical reading of this manuscript and all the scientific (besides many
other) conversations; Ivan for the lessons in self-defense; Sophie and Brigitte for
ongoing english-french and french-french translation efforts ; Edith Gouin and
Laurent Blanchoin for outstanding collaboration and help during my thesis; Denise
Guetard for the help and introduction to working secrets in a P3 laboratory; Klaus
Braun for his supervision and help in the very first steps of my work; Rüdiger Pipkorn
for the help with PNA syntheses; the mass spectrometry people at the DKFZ and the
Department of Chemistry at Heidelberg University for the vast amount of samples
they measured for me; Ms. M. Decraene and Ms. Dr. M. Schaade for their support in
administrative questions … and without whom my “convention de cotutelle” would
have never existed.
Also I want to thank all my friends (and at this point especially the Nocke family) for
the pleasurable moments and keeping me up during the last years.
Finally the most special thank to my father, my brother and Felix.Für meinen Vater
und
meine Mutter
(Du hast den Raum mit Sonne geflutet)TABLE OF CONTENTS:
1. Introduction ........................................................................................................................................... 7
1.1 THE HUMAN IMMUNODEFICIENCY VIRUS (HIV)........................................................................ 8
1.1.1 History and origin of the virus .................................................................................................... 8
1.1.2 Status quo of the epidemic........................................................................................................... 9
1.1.3 Retroviruses................................................................................................................................ 10
1.1.4 Genomic organization................................................................................................................ 12
1.1.5 Virion proteins of HIV ............................................................................................................... 14
Gag precursor ...................................................................................................................................................14
MA protein .......................................................................................................................................................14
NC protein...................................15
CA protein16
p6 ......................................................................................................................................................................16
Proteins derived from pol and pro...................................................................................................................17env................................................................................................................................18
Rev....................................................................................................................................................................18
Tat.........................................18
Nef19
Vif and Vpu......................................................................................................................................................19
Viral protein R (Vpr)........................................................................................................................................20
1.1.6 The retroviral life cycle.............................................................................................................. 20
1.2 PEPTIDE NUCLEIC ACIDS (PNAS) ...................................................................................................... 23
1.2.1 Chemical synthesis and properties of PNAs ............................................................................. 25
1.2.2 Antigene and antisense application of PNAs............................................................................ 27
Inhibition of replication.............................27
Inhibition of transcription ................................................................................................................................27
Inhibition of translation..............................28
1.2.3 Cellular uptake of PNA.............................................................................................................. 29
1.2.4 Therapy approach against HIV................................................................................................. 30
1.3 THE ACTIN CYTOSKELETON....................................................................................................... 32
1.3.1 Basic actin biochemistry............................................................................................................ 32
1.3.2. Viral cytoplasmic locomotion................................................................................................... 34
1.3.3 Routing of HIV particles from the MTOC to the nuclear envelope ......................................... 36
1.3.4 Visualization of dynamic actin inside living cells..................................................................... 37
1.4 OBJECTIVES OF THE WORK ........................................................................................................ 39
2. Material and Methods ......................................................................................................................... 40
2.1 PNA and peptide synthesis............................................................................................................ 41
2.2 Experimental systems to test viral production in early and late life cycle ................................. 44
Virus and cell lines...........................................................................................................................................44
Virus infections and assay readout protocols..................................................................................................44
12.3 Cell culture.................................................................................................................................... 45
Cell lines............