rologie i - European Congress of Virology

5 th European Congress of Virology07. CELLULAR FACTORS CONTROLINGVIRAL INFECTION AND ROLE OF HOSTGENETICSPosters: REF 089 to REF 104REF 089Structural and functional analyses of the innate immune receptortetherinNicolas ASCHMAN 1 , Nolwenn MIGUET 1 , Andreas HINZ 1 , PatriciaRENESTO 1 , Yoshiko USAMI 2 , Heinrich GÖTTLINGER 2 , WinfriedWEISSENHORN 11 Unit of Virus Host Cell Interactions, UMI 3265, Universite Joseph FourierEMBL CNRS, Grenoble, FRANCE; 2 Program in Molecular Medicine,University of Massachusetts Medical School, Worcester, USATetherin is an interferon inducible host factor that inhibits the release ofsome enveloped viruses including HIV 1, from the surface of infectedcells. Many susceptible viruses have evolved proteins that specificallycounteract this activity, such as the accessory protein Vpu in the case ofHIV 1. Tetherin is a type II transmembrane protein with a 170 Å long rodshaped extracellular region, modified with a C terminal glycosyl phosphatidylinositol(GPI) anchor. Crystal structures further reveal a dimericassembly mediated through a disulfide linked coiled coil domain. Althoughthe mechanism of virus particle retention is not fully understood, tetherin’sunique double membrane anchored topology suggests that it insertsinto both cellular and viral membranes during the budding process. Inorder to better characterise the antiviral mechanism of tetherin, we studiedthe role of various determinants such as irregular coiled coil residues,dimer stabilising disulfides and glycosylation by site directed mutagenesis.We propose that the protein’s inherent conformational flexibility isrequired for efficient retention of virions. Furthermore, it has been previouslyobserved that even in the presence of the antagonist Vpu, sometetherin is still incorporated into cell free virions. This led us to speculatethat the antiviral mechanism depends on its cell surface concentration andpossible organisation of tetherin into higher order assemblies at HIV 1budding sites. Molecular details of tetherin structure and function will bepresented.localized on the left arm of the third chromosome, affect the resistance toDCV infection (Magwire et al, 2012). We are now deciphering the roleof this uncharacterized gene in the control of DCV infection. Our loss offunction and gain of function experiments indicate that pastrel encodes amolecule opposing DCV infection, raising the question of the mechanisminvolved. Co localization experiments indicate that Pastrel is localized inlipid droplets. All together, our data suggest a connection between lipiddroplets and restriction of viral infection in Drosophila, as already describedin mammals between the restriction factor Viperin, present on lipiddroplets, and the replication of the human pathogen Hepatitis C Virus(Helbig et al, 2011).REF 092Host dependence of the thermosensibility of measles virus vaccinestrains and critical function of HSP90 in viral replicationLouis Marie BLOYET 1 , Jérémy WELSH 1 , Jessica RABILLOUD 1 ,Branka HORVAT 1 , Denis GERLIER 1 , Boyan GRIGOROV 1CIRI, INSERM U1111, CNRS, Université Lyon 1, ENS Lyon, Lyon,FRANCEAttenuation of measles virus (MeV) that gave rise to a well toleratedvaccine was ultimately reached after adaptation to grow into chickenembryonic fibroblasts (CEF). Moraten and Schwarz vaccines have beenselected for their ability to grow in CEF at 32 ◦ C but not at 37 ◦ C. Thisthermosensibility is not intrinsic to the viruses as they displayed a normalgrowth at 37 ◦ C in human or simian cells. A correlation between the temperaturerestriction of viral growth and a decreased expression of HSP90in CEF lead us to investigate the role of HSP90 in MeV replication cycle.In these cells, HSP90 appears to be a limiting factor since overexpressionof HSP90 in CEF enhanced MeV growth. Furthermore, inhibition ofHSP90 either by siRNA or by addition of a chemical inhibitor decreasedor abrogated the viral RNA synthesis and viral production not only in CEFbut also in other cell types such as Vero cells. Kinetics experiments andreversibility of the inhibition upon drug retrieval pointed to the polymerasecomplex as a likely target of HSP90 activity. Upon blocking HSP90, theamount of MeV L protein detectable by western blot was strongly decreased,while P expression remained unaffected. Treatment of the cells witha proteasome inhibitor did not prevent the decrease induced by the HSP90inhibitor. HSP90 was detected in viral particles suggesting a direct interactionbetween L and HSP90 and ongoing experiments aim at investigatingthis interaction.REF 091Pastrel: a new Drosophila gene restricting infection by the picornalike virus DCVVincent BARBIER, Akira GOTO, Carine MEIGNIN, KarimMAJZOUB, Jean Luc IMLERUPR9022 Institut de Biologie Moléculaire et Cellulaire (I.B.M.C.), Strasbourg,FRANCESince the discovery of the evolutionarily conserved TOLL and IMD pathways,involved in anti fungal and anti bacterial immune responses, thefruit fly Drosophila melanogaster is used as a model to study the molecularmechanisms of innate immunity. To defend against viral pathogens,Drosophila relies on two main facets: the RNA interference (RNAi) pathway,which plays a broad role in the control of viruses, and virus specificinducible responses (Kemp et al, 2013). We also observed that the fly geneticbackground can modulate the resistance to infection by Drosophila CVirus (DCV), a natural pathogen of Drosophila. A genome wide associationstudy recently showed that polymorphisms in the gene named pastrel,REF 093Restriction of rAAV mediated transgene expression in dystrophicmuscles through epigenetic silencing mechanismsJean Baptiste DUPONT 1 , Benoît TOURNAIRE 1 , BéatriceMAROLLEAU 2 , Laetitia VAN WITTENBERGHE 2 , ChristopheGEORGER 2 , Laurence DUBREIL 3 , Emilie LECOMTE 1 , LaurenceJEANSON LEH 2 , Bernard GJATA 2 , Fulvio MAVILIO 2 , Richard O.SNYDER 1,4,5 , Philippe MOULLIER 1,4 , Adrien LÉGER 11 INSERM UMR 1089, Nantes, FRANCE; 2 GENETHON, Evry, FRANCE;3 INRA UMR 703, Nantes, FRANCE; 4 Department of Molecular Geneticsand Microbiology, University of Florida, Gainesville, USA; 5 CERHB,University of Florida, Gainesville, USARepressive epigenetic modifications such as DNA methylation or HistonePost Translational Modifications (HPTMs) can restrict viral genomeexpression and participate in the establishment of latency. During theirlong coevolution with host cells, viruses developed molecular strategiesto counteract epigenetic restriction. However, due to their lack ofviral genes, retro/lentivectors and adenoviral vectors have lost this abilityS144 Virologie, Vol 17, supplément 2, septembre 2013