rologie i - European Congress of Virology

5 th European Congress of Virologycycle. We determined the crystal structure of the N terminal domainfrom Influenza virus C M1 and produced low resolution models of fulllength M1 from influenza A and C viruses based on SAXS analyses.Because M1 has to at least partially dissociate from the membraneduring entry, we analyzed potential structural changes induced by theendosomal pH by NMR and tested M1 interaction with model membranesat physiological and acidic pHs. We will present an integratedstructural model of M1 function based on our structural and functionaldata.and is attenuated for propagation in cell culture. Importantly, we alsoshow that the efficient translation of viral mRNAs containing a translationenhancer sequence also contributes to the disassembly of SGs in infectedcells. Further, we show that the nsP3/G3BP interaction also blocksSG induction by other stresses than virus infection such as pateamineA and 2 deoxy d glucose treatment. This is one of few described viralmechanisms for SG disruption and underlines the role of SGs in anti viraldefence.REF 220Influence of the actin bundling protein Fascin on HTLV 1 transmissionChristine GROSS, Martina KALMER, Bernhard FLECKENSTEIN,Andrea K. KRESSInstitute of Clinical and Molecular Virology, Friedrich Alexander UniversitätErlangen Nürnberg, Erlangen, GERMANYHuman T cell lymphotropic virus type 1 (HTLV 1) preferentially infectsCD4+ T cells via cell to cell transmission, which requires both the viralTax protein and the host cell cytoskeleton. We previously reported Taxdependent upregulation of the actin bundling protein Fascin (FSCN 1)in HTLV 1 infected cells. Now, we asked whether Fascin contributes toHTLV 1 transmission. Using confocal microscopy, we found that Tax inducedFascin colocalized with Actin in transfected and chronically infectedT cells. Moreover, both Fascin and Actin accumulated at cell cell contactssuggesting recruitment of Actin by Fascin. Use of shRNAs led to specificand stable repression of Fascin in both Tax transfected and chronicallyinfected T cells without affecting cell vitality. Moreover, the accumulationof both Fascin and Actin at cell cell contacts was reverted in the presenceof Fascin specific shRNAs. In transmission assays, we found that repressionof Fascin in HTLV 1 infected cells led to reduced transactivation ofco cultured reporter T cells. However, a first series of experiments indicatedan increase of the viral core protein gag p19 in the supernatantsof HTLV 1 infected cells when Fascin was repressed, suggesting elevatedvirus release. In contrast, block of both actin and tubulin reduced therelease of gag p19 levels into the supernatants. Taken together, these datasuggest that repression of Fascin re routes HTLV 1 from cell to cell to cellto supernatant transmission.REF 221Sequestration of G3BP by Semliki Forest virus NsP3 inhibits theassembly of stress granules on viral mRNAsMarc PANAS 1 , Margus VARJAK 2 , Aleksei LULLA 2 , Kai Er ENG 1 ,Andres MERITS 2 , Gunilla B. KARLSSON HEDESTAM 1 , Gerald M.MCINERNEY 11 Department of Microbiology, Tumor and Cell Biology, Stockholm,SWEDEN; 2 Tartu University Institute of Technology, Tartu, ESTONIADynamic, mRNA containing stress granules (SGs) form in the cytoplasmof cells under environmental stresses including viral infection. Manyviruses appear to employ mechanisms to disrupt the formation of SGson their mRNAs, suggesting that they represent a cellular defence againstinfection. Here, we report that early in Semliki Forest virus infection, theC terminal domain of the viral non structural protein 3 (nsP3) forms acomplex with Ras GAP SH3 domain binding protein (G3BP) and sequestersit into cytoplasmic foci in a manner that inhibits the formation ofSGs on viral mRNAs. These nsP3+G3BP+ foci contain neither TIA 1/Rnor members of the 43S translation initiation complex indicating that theyare not SGs, but often contain dsRNA suggesting they are viral RNAreplication complexes or similar structures. A viral mutant carrying a Cterminal truncation of nsP3 induces more numerous and persistent SGsREF 222Architecture of respiratory syncytial virus revealed by electron cryotomographySarah BUTCHER 1 , Lassi LILJEROOS 1 , Magdalena KRZYZANIAK 2 ,Ari HELENIUS 21 University of Helsinki, Helsinki, FINLAND; 2 ETH Zurich, Zurich,SWITZERLANDRespiratory syncytial virus is a human pathogen that causes severe infectionof the respiratory tract. Current information about the structure ofthe virus and its interaction with host cells is limited. We carried out anelectron cryotomographic characterization of cell culture grown RSV todetermine the architecture of the virion. The particles ranged from 100 nmto 1000 nm in diameter and were spherical, filamentous, or a mixture ofthe two. The filamentous morphology correlated with the presence of acylindrical matrix protein layer linked to the inner leaflet of the viralenvelope and with local ordering of the glycoprotein spikes identified bysubvolume averaging. Recombinant viruses with only the fusion protein intheir envelope showed that this glycoprotein was predominantly in a formthat resembled the post fusion conformation. The ribonucleocapsids wereleft handed, randomly oriented, and curved inside the virions. In filamentousparticles, they were often adjacent to an intermediate layer of proteinassigned to M2 1. Our results indicate important differences in structurebetween the Paramyxovirinae and Pneumovirinae subfamilies within theParamyxoviridae, and provide new insight into host cell entry and exit ofa serious pathogen.REF 223Structures of Fibre Protein Head Domains from Two New AdenovirusesAbhimanyu Kumar SINGH, Mark VAN RAAIJCentro Nacional de Biotecnologia (CNB CSIC), Madrid, SPAINAdenoviruses are linear, non enveloped, dsDNA viruses which are icosahedralin shape. They are divided into five genera. The fibre protein, one ofthe major structural proteins of adenovirus is the principle determinant ofviral tropism and facilitates primary host cell recognition via its globularhead domain. We have determined crystal structures of fibre proteinhead domains of Turkey Hemorrhagic Enteritis Virus (THEV, TurkeyAdenovirus type 3) from the Siadenovirus genus and Snake Adenovirustype 1 (SnAdV1) from the Atadenovirus genus. THEV exists in virulentas well as in avirulent forms, the difference between them lies in twomutations located in fibre head. Our 2.2 Å structure of the fibre head froman avirulent strain localises these mutations. The secondary structuretopology resembles other adenovirus fibre heads, although a difference isthe presence of an extended beta hairpin, which contacts a neighbouringmonomer. The positively charged surface suggests that the receptor maybe acidic. Curiously, the closest structural homologues are reovirus fibressigmaC and sigma 1. SnAdV1 has one fibre per penton base, but itsreceptor is not known. Using site directed mutagenesis and multi wavelengthanomalous dispersion technique we have determined the structureof its fibre protein head domain at 1.7 Å. The eight beta sheet containingVirologie, Vol 17, supplément 2, septembre 2013S181