title: Nuclear dynamics of influenza A virus ribonucleoproteins revealed by live-cell imaging studies.
abstract: The negative sense RNA genome of influenza A virus is transcribed and replicated in the nuclei of infected cells by the viral RNA polymerase. Only four viral polypeptides are required but multiple cellular components are potentially involved. We used fluorescence recovery after photobleaching (FRAP) to characterise the dynamics of GFP-tagged viral ribonucleoprotein (RNP) components in living cells. The nucleoprotein (NP) displayed very slow mobility that significantly increased on formation of transcriptionally active RNPs. Conversely, single or dimeric polymerase subunits showed fast nuclear dynamics that decreased upon formation of heterotrimers, suggesting increased interaction of the full polymerase complex with a relatively immobile cellular component(s). Treatment with inhibitors of cellular transcription indicated that in part, this reflected an interaction with cellular RNA polymerase II. Analysis of mutated influenza virus polymerase complexes further suggested that this was through an interaction between PB2 and RNA Pol II separate from PB2 cap-binding activity.
authors.affiliation: Division of Virology, Department of Pathology, University of Cambridge, Cambridge CB2 1QP, UK.
authors.forename: Eva M
authors.initials: EM
authors.lastname: Loucaides
authors.forename: Johann C
authors.initials: JC
authors.lastname: von Kirchbach
authors.forename: Agnes
authors.initials: A
authors.lastname: Foeglein
authors.forename: Jane
authors.initials: J
authors.lastname: Sharps
authors.forename: Ervin
authors.initials: E
authors.lastname: Fodor
authors.forename: Paul
authors.initials: P
authors.lastname: Digard
mesh_terms: D002460:Cell Line; D002467:Cell Nucleus / Q000737:chemistry; D006801:Humans; D009980:Influenza A virus / Q000502:physiology*; D019590:Nucleocapsid Proteins; D011485:Protein Binding; D012319:RNA Polymerase II / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D014758:Viral Core Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism; D014779:Virus Replication
chemical_list: C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; C057027:PB2 protein, Influenzavirus A; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins; D014764:Viral Proteins; D012319:RNA Polymerase II; D012324:RNA-Dependent RNA Polymerase