title: Influenza a virus polymerase is an integral component of the CPSF30-NS1A protein complex in infected cells.
abstract: The NS1A protein of influenza A virus binds the cellular CPSF30 protein, thereby inhibiting the 3'-end processing of all cellular pre-mRNAs, including beta interferon pre-mRNA. X-ray crystallography identified the CPSF30-binding pocket on the influenza virus A/Udorn/72 (Ud) NS1A protein and the critical role of two hydrophobic NS1A amino acids outside the pocket, F103 and M106, in stabilizing the CPSF30-NS1A complex. Although the NS1A protein of the 1997 H5N1 influenza A/Hong Kong/483/97 (HK97) virus contains L (not F) at position 103 and I (not M) at position 106, it binds CPSF30 in vivo to a significant extent because cognate (HK97) internal proteins stabilize the CPSF30-NS1A complex in infected cells. Here we show that the cognate HK97 polymerase complex, containing the viral polymerase proteins (PB1, PB2, and PA) and the nucleocapsid protein (NP), is responsible for this stabilization. The noncognate Ud polymerase complex cannot carry out this stabilization, but it can stabilize CPSF30 binding to a mutated (F103L M106I) cognate Ud NS1A protein. These results suggested that the viral polymerase complex is an integral component of the CPSF30-NS1A protein complex in infected cells even when the cognate NS1A protein contains F103 and M106, and we show that this is indeed the case. Finally, we show that cognate PA protein and NP, but not cognate PB1 and PB2 proteins, are required for stabilizing the CPSF30-NS1A complex, indicating that the NS1A protein interacts primarily with its cognate PA protein and NP in a complex that includes the cellular CPSF30 protein.
authors.affiliation: Institute for Cellular and Molecular Biology, University of Texas at Austin, 78712, USA.
authors.forename: Rei-Lin
authors.initials: RL
authors.lastname: Kuo
authors.forename: Robert M
authors.initials: RM
authors.lastname: Krug
mesh_terms: D002460:Cell Line; D039223:Cleavage And Polyadenylation Specificity Factor / Q000378:metabolism*; D006801:Humans; D053124:Influenza A Virus, H5N1 Subtype / Q000502:physiology*; D019590:Nucleocapsid Proteins; D011485:Protein Binding; D016601:RNA-Binding Proteins / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D014758:Viral Core Proteins / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism
chemical_list: D039223:Cleavage And Polyadenylation Specificity Factor; C085820:INS1 protein, influenza virus; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; C061290:PA protein, influenza viruses; C057027:PB2 protein, Influenzavirus A; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; C077346:influenza virus polymerase basic protein 1; D012324:RNA-Dependent RNA Polymerase