Nuclear Factor 90 Negatively Regulates Influenza Virus Replication by Interacting with Viral Nucleoprotein (cid:1)

Interactions between host factors and the viral replication complex play important roles in host adaptation and regulation of influenza virus replication. A cellular protein, nuclear factor 90 (NF90), was copurified with H5N1 viral nucleoprotein (NP) from human cells in which NP was transiently expressed and identified by matrix-assisted laser desorption ionization–time of flight mass spectrometry analysis. In vitro coimmunoprecipitation of NF90 and NP coexpressed in HEK 293T cells or individually expressed in bacterial and HEK 293T cells, respectively, confirmed a direct interaction between NF90 and NP, independent of other subunits of the ribonucleoprotein complex. This interaction was prevented by a mutation, F412A, in the C-terminal region of the NP, indicating that the C-terminal of NP is required for NF90 binding. RNase V treatment did not prevent coprecipitation of NP and NF90, which demonstrates that the interaction is RNA binding independent. After small interfering RNA knockdown of NF90 expression in A549 and HeLa cells, viral polymerase complex activity and virus replication were significantly increased, suggesting that NF90 negatively affects viral repli- cation. Both NP and NF90 colocalized in the nucleus of virus-infected cells during the early phase of infection, suggesting that the interaction between NF90 and NP is an early event in virus replication. Quantitative reverse transcription-PCR showed that NF90 downregulates both viral genome replication and mRNA transcription in infected cells. These results suggest that NF90 inhibits influenza virus replication during the early phase of infection through direct interaction with viral NP. (NA)

[1]  H. Nair,et al.  Swine-origin influenza A (H1N1) virus infections in a school - New York City, April 2009. , 2009, MMWR. Morbidity and mortality weekly report.

[2]  Y. Guan,et al.  Induction of Proinflammatory Cytokines in Primary Human Macrophages by Influenza A Virus (H5N1) Is Selectively Regulated by IFN Regulatory Factor 3 and p38 MAPK1 , 2009, The Journal of Immunology.

[3]  Jin Hyun Kim,et al.  Viral RNA polymerase complex promotes optimal growth of 1918 virus in the lower respiratory tract of ferrets , 2009, Proceedings of the National Academy of Sciences.

[4]  Y. Guan,et al.  Systemic infection of avian influenza A virus H5N1 subtype in humans , 2009, Human Pathology.

[5]  Wang Guansong,et al.  Update on avian influenza A (H5N1) virus infection in humans , 2009 .

[6]  Y. Guan,et al.  Broad cross-protection against H5N1 avian influenza virus infection by means of monoclonal antibodies that map to conserved viral epitopes. , 2009, The Journal of infectious diseases.

[7]  A. Tomoiu,et al.  Avian Influenza A Virus Polymerase Association with Nucleoprotein, but Not Polymerase Assembly, Is Impaired in Human Cells during the Course of Infection , 2008, Journal of Virology.

[8]  S. Goff,et al.  Host restriction factors blocking retroviral replication. , 2008, Annual review of genetics.

[9]  A. Tomoiu,et al.  Host restriction of avian influenza viruses at the level of the ribonucleoproteins. , 2008, Annual review of microbiology.

[10]  J. Doudna,et al.  An inhibitory activity in human cells restricts the function of an avian-like influenza virus polymerase. , 2008, Cell host & microbe.

[11]  Atsushi Kawaguchi,et al.  Host factors for replication and transcription of the influenza virus genome , 2008, Reviews in medical virology.

[12]  Juan Pablo Albar,et al.  Analysis of the interaction of influenza virus polymerase complex with human cell factors , 2008, Proteomics.

[13]  C. Davis,et al.  Genetic Compatibility and Virulence of Reassortants Derived from Contemporary Avian H5N1 and Human H3N2 Influenza A Viruses , 2008, PLoS pathogens.

[14]  I. Lossos,et al.  NFAR-1 and -2 modulate translation and are required for efficient host defense , 2008, Proceedings of the National Academy of Sciences.

[15]  H. Klenk,et al.  Interaction of Polymerase Subunit PB2 and NP with Importin α1 Is a Determinant of Host Range of Influenza A Virus , 2008, PLoS pathogens.

[16]  Ryosuke Nakano,et al.  An influenza virus replicon system in yeast identified Tat-SF1 as a stimulatory host factor for viral RNA synthesis , 2007, Proceedings of the National Academy of Sciences.

[17]  A. Ostareck-Lederer,et al.  Nuclear factors are involved in hepatitis C virus RNA replication. , 2007, RNA.

[18]  E. Agbottah,et al.  Nuclear Factor 90(NF90) targeted to TAR RNA inhibits transcriptional activation of HIV-1 , 2007, Retrovirology.

[19]  K. Labadie,et al.  Host-range determinants on the PB2 protein of influenza A viruses control the interaction between the viral polymerase and nucleoprotein in human cells. , 2007, Virology.

[20]  Stefan Hippenstiel,et al.  IFNβ induction by influenza A virus is mediated by RIG‐I which is regulated by the viral NS1 protein , 2007, Cellular microbiology.

[21]  Jonas Grossmann,et al.  Identification of cellular interaction partners of the influenza virus ribonucleoprotein complex and polymerase complex using proteomic-based approaches. , 2007, Journal of proteome research.

[22]  J. Taubenberger,et al.  Discovery and Characterization of the 1918 Pandemic Influenza Virus in Historical Context , 2005, Antiviral therapy.

[23]  R. Krug,et al.  The mechanism by which influenza A virus nucleoprotein forms oligomers and binds RNA , 2006, Nature.

[24]  Benjamin Thomas,et al.  Role of Ran Binding Protein 5 in Nuclear Import and Assembly of the Influenza Virus RNA Polymerase Complex , 2006, Journal of Virology.

[25]  Ervin Fodor,et al.  Functional association between viral and cellular transcription during influenza virus infection , 2006, Reviews in medical virology.

[26]  N. Cox,et al.  Lack of transmission of H5N1 avian–human reassortant influenza viruses in a ferret model , 2006, Proceedings of the National Academy of Sciences.

[27]  R. Krug,et al.  Binding of the influenza A virus NS1 protein to PKR mediates the inhibition of its activation by either PACT or double-stranded RNA. , 2006, Virology.

[28]  R. Krug,et al.  The primary function of RNA binding by the influenza A virus NS1 protein in infected cells: Inhibiting the 2'-5' oligo (A) synthetase/RNase L pathway. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[29]  R. Webster,et al.  The polymerase complex genes contribute to the high virulence of the human H5N1 influenza virus isolate A/Vietnam/1203/04 , 2006, The Journal of experimental medicine.

[30]  H. Klenk,et al.  The viral polymerase mediates adaptation of an avian influenza virus to a mammalian host. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[31]  Yi Guan,et al.  Human Infection with an Avian H9N2 Influenza A Virus in Hong Kong in 2003 , 2005, Journal of Clinical Microbiology.

[32]  Ervin Fodor,et al.  Association of the Influenza A Virus RNA-Dependent RNA Polymerase with Cellular RNA Polymerase II , 2005, Journal of Virology.

[33]  Michael B. Mathews,et al.  The double-stranded-RNA-binding motif: interference and much more , 2004, Nature Reviews Molecular Cell Biology.

[34]  Marion Koopmans,et al.  Avian influenza A virus (H7N7) associated with human conjunctivitis and a fatal case of acute respiratory distress syndrome. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[35]  Etsuko Hirayama,et al.  Heat Shock Protein 70 Is Related to Thermal Inhibition of Nuclear Export of the Influenza Virus Ribonucleoprotein Complex , 2004, Journal of Virology.

[36]  R. Webster,et al.  Are We Ready for Pandemic Influenza? , 2003, Science.

[37]  F. Grosse,et al.  Members of the NF90/NFAR protein group are involved in the life cycle of a positive‐strand RNA virus , 2003, The EMBO journal.

[38]  M. Mathews,et al.  Selective regulation of gene expression by nuclear factor 110, a member of the NF90 family of double-stranded RNA-binding proteins. , 2003, Journal of molecular biology.

[39]  G. Barber,et al.  The dsRNA binding protein family: critical roles, diverse cellular functions , 2003, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[40]  R. Krug,et al.  Intracellular warfare between human influenza viruses and human cells: the roles of the viral NS1 protein. , 2003, Virology.

[41]  K. Yano,et al.  Identification of Hsp90 as a Stimulatory Host Factor Involved in Influenza Virus RNA Synthesis* , 2002, The Journal of Biological Chemistry.

[42]  Y. Guan,et al.  Universal primer set for the full-length amplification of all influenza A viruses , 2001, Archives of Virology.

[43]  Maite Huarte,et al.  PA Subunit from Influenza Virus Polymerase Complex Interacts with a Cellular Protein with Homology to a Family of Transcriptional Activators , 2001, Journal of Virology.

[44]  M. Mathews,et al.  Nuclear Factor 90 Is a Substrate and Regulator of the Eukaryotic Initiation Factor 2 Kinase Double-stranded RNA-activated Protein Kinase* , 2001, The Journal of Biological Chemistry.

[45]  A. Mayeda,et al.  Characterization of Two Evolutionarily Conserved, Alternatively Spliced Nuclear Phosphoproteins, NFAR-1 and -2, That Function in mRNA Processing and Interact with the Double-stranded RNA-dependent Protein Kinase, PKR* , 2001, The Journal of Biological Chemistry.

[46]  S. Baird,et al.  Pattern of mutation in the genome of influenza A virus on adaptation to increased virulence in the mouse lung: Identification of functional themes , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[47]  R. Webster,et al.  H9N2 influenza A viruses from poultry in Asia have human virus-like receptor specificity. , 2001, Virology.

[48]  R. Webster,et al.  A DNA transfection system for generation of influenza A virus from eight plasmids. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[49]  R. Webster,et al.  "Ambisense" approach for the generation of influenza A virus: vRNA and mRNA synthesis from one template. , 2000, Virology.

[50]  J. Skehel,et al.  Receptor binding and membrane fusion in virus entry: the influenza hemagglutinin. , 2000, Annual review of biochemistry.

[51]  M. Peiris,et al.  Human infection with influenza H9N2 , 1999, The Lancet.

[52]  P. Digard,et al.  Edinburgh Research Explorer Identification of amino acid residues of influenza virus nucleoprotein essential for RNA binding , 2022 .

[53]  W. Guo,et al.  DRBP76, a Double-stranded RNA-binding Nuclear Protein, Is Phosphorylated by the Interferon-induced Protein Kinase, PKR* , 1999, The Journal of Biological Chemistry.

[54]  E. Medcalf,et al.  Modulation of Nuclear Localization of the Influenza Virus Nucleoprotein through Interaction with Actin Filaments , 1999, Journal of Virology.

[55]  P. L. Boutz,et al.  Influenza Virus Nucleoprotein Interacts with Influenza Virus Polymerase Proteins , 1998, Journal of Virology.

[56]  R. Krug,et al.  Influenza virus NS1 protein interacts with the cellular 30 kDa subunit of CPSF and inhibits 3'end formation of cellular pre-mRNAs. , 1998, Molecular cell.

[57]  M. Peiris,et al.  Clinical features and rapid viral diagnosis of human disease associated with avian influenza A H5N1 virus , 1998, The Lancet.

[58]  R. Webster,et al.  Human influenza A H5N1 virus related to a highly pathogenic avian influenza virus , 1998, The Lancet.

[59]  P. Wang,et al.  The NPI-1/NPI-3 (karyopherin alpha) binding site on the influenza a virus nucleoprotein NP is a nonconventional nuclear localization signal , 1997, Journal of virology.

[60]  R. Lamb,et al.  Orthomyxoviridae: The Viruses and Their Replication. , 1996 .

[61]  A. Smith,et al.  Cloning and expression of cyclosporin A- and FK506-sensitive nuclear factor of activated T-cells: NF45 and NF90. , 1994, The Journal of biological chemistry.

[62]  S. Cusack,et al.  Structure of influenza virus RNP. I. Influenza virus nucleoprotein melts secondary structure in panhandle RNA and exposes the bases to the solvent. , 1994, The EMBO journal.

[63]  B. Murphy,et al.  The avian influenza virus nucleoprotein gene and a specific constellation of avian and human virus polymerase genes each specify attenuation of avian-human influenza A/Pintail/79 reassortant viruses for monkeys , 1987, Journal of virology.

[64]  C. Scholtissek,et al.  The nucleoprotein as a possible major factor in determining host specificity of influenza H3N2 viruses. , 1985, Virology.

[65]  W. J. Bean.  Correlation of influenza A virus nucleoprotein genes with host species. , 1984, Virology.