title: HSP90 Chaperoning in Addition to Phosphoprotein Required for Folding but Not for Supporting Enzymatic Activities of Measles and Nipah Virus L Polymerases.
abstract: Nonsegmented negative-stranded RNA viruses, or members of the order Mononegavirales, share a conserved gene order and the use of elaborate transcription and replication machinery made up of at least four molecular partners. These partners have coevolved with the acquisition of the permanent encapsidation of the entire genome by the nucleoprotein (N) and the use of this N-RNA complex as a template for the viral polymerase composed of the phosphoprotein (P) and the large enzymatic protein (L). Not only is P required for polymerase function, but it also stabilizes the L protein through an unknown underlying molecular mechanism. By using NVP-AUY922 and/or 17-dimethylaminoethylamino-17-demethoxygeldanamycin as specific inhibitors of cellular heat shock protein 90 (HSP90), we found that efficient chaperoning of L by HSP90 requires P in the measles, Nipah, and vesicular stomatitis viruses. While the production of P remains unchanged in the presence of HSP90 inhibitors, the production of soluble and functional L requires both P and HSP90 activity. Measles virus P can bind the N terminus of L in the absence of HSP90 activity. Both HSP90 and P are required for the folding of L, as evidenced by a luciferase reporter insert fused within measles virus L. HSP90 acts as a true chaperon; its activity is transient and dispensable for the activity of measles and Nipah virus polymerases of virion origin. That the cellular chaperoning of a viral polymerase into a soluble functional enzyme requires the assistance of another viral protein constitutes a new paradigm that seems to be conserved within the Mononegavirales order.
  
  
  
  Viruses are obligate intracellular parasites that require a cellular environment for their replication. Some viruses particularly depend on the cellular chaperoning apparatus. We report here that for measles virus, successful chaperoning of the viral L polymerase mediated by heat shock protein 90 (HSP90) requires the presence of the viral phosphoprotein (P). Indeed, while P protein binds to the N terminus of L independently of HSP90 activity, both HSP90 and P are required to produce stable, soluble, folded, and functional L proteins. Once formed, the mature P+L complex no longer requires HSP90 to exert its polymerase functions. Such a new paradigm for the maturation of a viral polymerase appears to be conserved in several members of the Mononegavirales order, including the Nipah and vesicular stomatitis viruses.
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.
authors.forename: Louis-Marie
authors.initials: LM
authors.lastname: Bloyet
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.|Laboratoire d'Excellence Ecofect, Lyon, France.
authors.forename: Jérémy
authors.initials: J
authors.lastname: Welsch
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.
authors.forename: François
authors.initials: F
authors.lastname: Enchery
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.|Laboratoire d'Excellence Ecofect, Lyon, France.
authors.forename: Cyrille
authors.initials: C
authors.lastname: Mathieu
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.
authors.forename: Sylvain
authors.initials: S
authors.lastname: de Breyne
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.|Laboratoire d'Excellence Ecofect, Lyon, France.
authors.forename: Branka
authors.identifier: http://orcid.org/0000-0003-0578-7765
authors.initials: B
authors.lastname: Horvat
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France boyan.grigorov@inserm.fr denis.gerlier@inserm.fr.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.|Cancer Research Center of Lyon (CRCL), INSERM, U1052, Université Claude Bernard Lyon 1, Lyon, France.
authors.forename: Boyan
authors.initials: B
authors.lastname: Grigorov
authors.affiliation: CIRI, International Center for Infectiology Research, Université de Lyon, Lyon, France boyan.grigorov@inserm.fr denis.gerlier@inserm.fr.|INSERM, U1111, Lyon, France.|CNRS, UMR5308, Lyon, France.|Université Claude Bernard Lyon 1; Centre International de Recherche en Infectiologie, Lyon, France.|ENS Lyon, Lyon, France.|Laboratoire d'Excellence Ecofect, Lyon, France.
authors.forename: Denis
authors.identifier: http://orcid.org/0000-0001-5539-456X
authors.initials: D
authors.lastname: Gerlier
mesh_terms: D000818:Animals; D002522:Chlorocebus aethiops; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D045464:Henipavirus Infections / Q000378:metabolism* / Q000821:virology; D006801:Humans; D008457:Measles / Q000378:metabolism* / Q000821:virology; D008459:Measles virus / Q000502:physiology; D051379:Mice; D045405:Nipah Virus / Q000502:physiology; D009698:Nucleoproteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D017510:Protein Folding; D018353:Rhabdoviridae Infections / Q000378:metabolism / Q000821:virology; D014709:Vero Cells; D018116:Vesiculovirus / Q000502:physiology; D014764:Viral Proteins / Q000378:metabolism; D014771:Virion / Q000502:physiology; D014779:Virus Replication
chemical_list: D018841:HSP90 Heat-Shock Proteins; D009698:Nucleoproteins; D010750:Phosphoproteins; D014764:Viral Proteins; C435063:L protein, Nipah virus; D012321:DNA-Directed RNA Polymerases; C512399:L polymerase protein, Vesicular stomatitis-Indiana virus