title: A loss of function analysis of host factors influencing Vaccinia virus replication by RNA interference.
abstract: Vaccinia virus (VACV) is a large, cytoplasmic, double-stranded DNA virus that requires complex interactions with host proteins in order to replicate. To explore these interactions a functional high throughput small interfering RNA (siRNA) screen targeting 6719 druggable cellular genes was undertaken to identify host factors (HF) influencing the replication and spread of an eGFP-tagged VACV. The experimental design incorporated a low multiplicity of infection, thereby enhancing detection of cellular proteins involved in cell-to-cell spread of VACV. The screen revealed 153 pro- and 149 anti-viral HFs that strongly influenced VACV replication. These HFs were investigated further by comparisons with transcriptional profiling data sets and HFs identified in RNAi screens of other viruses. In addition, functional and pathway analysis of the entire screen was carried out to highlight cellular mechanisms involved in VACV replication. This revealed, as anticipated, that many pro-viral HFs are involved in translation of mRNA and, unexpectedly, suggested that a range of proteins involved in cellular transcriptional processes and several DNA repair pathways possess anti-viral activity. Multiple components of the AMPK complex were found to act as pro-viral HFs, while several septins, a group of highly conserved GTP binding proteins with a role in sequestering intracellular bacteria, were identified as strong anti-viral VACV HFs. This screen has identified novel and previously unexplored roles for cellular factors in poxvirus replication. This advancement in our understanding of the VACV life cycle provides a reliable knowledge base for the improvement of poxvirus-based vaccine vectors and development of anti-viral theraputics.
authors.affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Philippa M
authors.initials: PM
authors.lastname: Beard
authors.affiliation: Division of Pathway Medicine, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Samantha J
authors.initials: SJ
authors.lastname: Griffiths
authors.affiliation: Institute for Informatics, Ludwig-Maximilians-Universität München, München, Germany.
authors.forename: Orland
authors.initials: O
authors.lastname: Gonzalez
authors.affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Ismar R
authors.initials: IR
authors.lastname: Haga
authors.affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Tali
authors.initials: T
authors.lastname: Pechenick Jowers
authors.affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Danielle K
authors.initials: DK
authors.lastname: Reynolds
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Kings Buildings, Edinburgh, Midlothian, United Kingdom.
authors.forename: Jan
authors.initials: J
authors.lastname: Wildenhain
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Kings Buildings, Edinburgh, Midlothian, United Kingdom.
authors.forename: Hille
authors.initials: H
authors.lastname: Tekotte
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Kings Buildings, Edinburgh, Midlothian, United Kingdom.
authors.forename: Manfred
authors.initials: M
authors.lastname: Auer
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Kings Buildings, Edinburgh, Midlothian, United Kingdom; Institute for Research in Immunology and Cancer, Universite de Montreal, Montreal, Quebec, Canada.
authors.forename: Mike
authors.initials: M
authors.lastname: Tyers
authors.affiliation: Division of Pathway Medicine, University of Edinburgh, Edinburgh, Midlothian, United Kingdom; SynthSys, Centre for Integrative Systems at Edinburgh, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Peter
authors.initials: P
authors.lastname: Ghazal
authors.affiliation: Institute for Informatics, Ludwig-Maximilians-Universität München, München, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Zimmer
authors.affiliation: Division of Pathway Medicine, University of Edinburgh, Edinburgh, Midlothian, United Kingdom.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Haas
mesh_terms: D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D057166:High-Throughput Screening Assays; D054884:Host-Pathogen Interactions / Q000235:genetics*; D006801:Humans; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D015203:Reproducibility of Results; D015398:Signal Transduction; D014158:Transcription, Genetic; D014615:Vaccinia / Q000235:genetics* / Q000378:metabolism / Q000821:virology*; D014616:Vaccinia virus / Q000502:physiology*; D014779:Virus Replication
chemical_list: D034741:RNA, Small Interfering