title: Activation of c-jun N-terminal kinase upon influenza A virus (IAV) infection is independent of pathogen-related receptors but dependent on amino acid sequence variations of IAV NS1.
abstract: A hallmark cell response to influenza A virus (IAV) infections is the phosphorylation and activation of c-jun N-terminal kinase (JNK). However, so far it is not fully clear which molecules are involved in the activation of JNK upon IAV infection. Here, we report that the transfection of influenza viral-RNA induces JNK in a retinoic acid-inducible gene I (RIG-I)-dependent manner. However, neither RIG-I-like receptors nor MyD88-dependent Toll-like receptors were found to be involved in the activation of JNK upon IAV infection. Viral JNK activation may be blocked by addition of cycloheximide and heat shock protein inhibitors during infection, suggesting that the expression of an IAV-encoded protein is responsible for JNK activation. Indeed, the overexpression of nonstructural protein 1 (NS1) of certain IAV subtypes activated JNK, whereas those of some other subtypes failed to activate JNK. Site-directed mutagenesis experiments using NS1 of the IAV H7N7, H5N1, and H3N2 subtypes identified the amino acid residue phenylalanine (F) at position 103 to be decisive for JNK activation. Cleavage- and polyadenylation-specific factor 30 (CPSF30), whose binding to NS1 is stabilized by the amino acids F103 and M106, is not involved in JNK activation. Conclusively, subtype-specific sequence variations in the IAV NS1 protein result in subtype-specific differences in JNK signaling upon IAV infection.
  
  
  OBJECTIVE
  Influenza A virus (IAV) infection leads to the activation or modulation of multiple signaling pathways. Here, we demonstrate for the first time that the c-jun N-terminal kinase (JNK), a long-known stress-activated mitogen-activated protein (MAP) kinase, is activated by RIG-I when cells are treated with IAV RNA. However, at the same time, nonstructural protein 1 (NS1) of IAV has an intrinsic JNK-activating property that is dependent on IAV subtype-specific amino acid variations around position 103. Our findings identify two different and independent pathways that result in the activation of JNK in the course of an IAV infection.
authors.affiliation: Institute of Molecular Virology (IMV), Centre of Molecular Biology of Inflammation (ZMBE), Westfälische Wilhelms-Universität Münster, Münster, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Nacken
authors.affiliation: Institute of Molecular Virology (IMV), Centre of Molecular Biology of Inflammation (ZMBE), Westfälische Wilhelms-Universität Münster, Münster, Germany.
authors.forename: Darisuren
authors.initials: D
authors.lastname: Anhlan
authors.affiliation: Institute of Molecular Virology (IMV), Centre of Molecular Biology of Inflammation (ZMBE), Westfälische Wilhelms-Universität Münster, Münster, Germany.
authors.forename: Eike R
authors.initials: ER
authors.lastname: Hrincius
authors.affiliation: Virology Laboratory, National Research Center, Cairo, Egypt.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Mostafa
authors.affiliation: Division of Influenza and other Respiratory Viruses, Robert Koch Institute (RKI), Berlin, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wolff
authors.affiliation: Division of Influenza and other Respiratory Viruses, Robert Koch Institute (RKI), Berlin, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Sadewasser
authors.affiliation: Institute of Medical Virology, Justus Liebig University Giessen, Giessen, Germany.
authors.forename: Stephan
authors.initials: S
authors.lastname: Pleschka
authors.affiliation: Institute of Molecular Virology (IMV), Centre of Molecular Biology of Inflammation (ZMBE), Westfälische Wilhelms-Universität Münster, Münster, Germany Cells in Motion Cluster of Excellence, University of Münster, Münster, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Ehrhardt
authors.affiliation: Institute of Molecular Virology (IMV), Centre of Molecular Biology of Inflammation (ZMBE), Westfälische Wilhelms-Universität Münster, Münster, Germany Cells in Motion Cluster of Excellence, University of Münster, Münster, Germany ludwigs@uni-muenster.de.
authors.forename: Stephan
authors.initials: S
authors.lastname: Ludwig
mesh_terms: D000595:Amino Acid Sequence / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism; D004285:Dogs; D057809:HEK293 Cells; D006801:Humans; D009980:Influenza A virus / Q000235:genetics* / Q000378:metabolism; D007251:Influenza, Human / Q000235:genetics* / Q000378:metabolism / Q000821:virology; D048031:JNK Mitogen-Activated Protein Kinases / Q000235:genetics* / Q000378:metabolism; D061985:Madin Darby Canine Kidney Cells; D016297:Mutagenesis, Site-Directed / Q000379:methods; D053594:Myeloid Differentiation Factor 88 / Q000235:genetics / Q000378:metabolism; D009976:Orthomyxoviridae Infections / Q000235:genetics* / Q000378:metabolism / Q000821:virology; D015398:Signal Transduction / Q000235:genetics; D014212:Tretinoin / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D053594:Myeloid Differentiation Factor 88; D017361:Viral Nonstructural Proteins; D014212:Tretinoin; D048031:JNK Mitogen-Activated Protein Kinases; D053487:DEAD-box RNA Helicases