title: Activation of the NLRP3 inflammasome by IAV virulence protein PB1-F2 contributes to severe pathophysiology and disease.
abstract: The ability for a host to recognize infection is critical for virus clearance and often begins with induction of inflammation. The PB1-F2 of pathogenic influenza A viruses (IAV) contributes to the pathophysiology of infection, although the mechanism for this is unclear. The NLRP3-inflammasome has been implicated in IAV pathogenesis, but whether IAV virulence proteins can be activators of the complex is unknown. We investigated whether PB1-F2-mediated activation of the NLRP3-inflammasome is a mechanism contributing to overt inflammatory responses to IAV infection. We show PB1-F2 induces secretion of pyrogenic cytokine IL-1β by activating the NLRP3-inflammasome, contributing to inflammation triggered by pathogenic IAV. Compared to infection with wild-type virus, mice infected with reverse engineered PB1-F2-deficient IAV resulted in decreased IL-1β secretion and cellular recruitment to the airways. Moreover, mice exposed to PB1-F2 peptide derived from pathogenic IAV had enhanced IL-1β secretion compared to mice exposed to peptide derived from seasonal IAV. Implicating the NLRP3-inflammasome complex specifically, we show PB1-F2 derived from pathogenic IAV induced IL-1β secretion was Caspase-1-dependent in human PBMCs and NLRP3-dependent in mice. Importantly, we demonstrate PB1-F2 is incorporated into the phagolysosomal compartment, and upon acidification, induces ASC speck formation. We also show that high molecular weight aggregated PB1-F2, rather than soluble PB1-F2, induces IL-1β secretion. Furthermore, NLRP3-deficient mice exposed to PB1-F2 peptide or infected with PB1-F2 expressing IAV were unable to efficiently induce the robust inflammatory response as observed in wild-type mice. In addition to viral pore forming toxins, ion channel proteins and RNA, we demonstrate inducers of NLRP3-inflammasome activation may include disordered viral proteins, as exemplified by PB1-F2, acting as host pathogen 'danger' signals. Elucidating immunostimulatory PB1-F2 mediation of NLRP3-inflammasome activation is a major step forward in our understanding of the aetiology of disease attributable to exuberant inflammatory responses to IAV infection.
authors.affiliation: Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia. jmcauley@unimelb.edu.au
authors.forename: Julie L
authors.initials: JL
authors.lastname: McAuley
authors.forename: Michelle D
authors.initials: MD
authors.lastname: Tate
authors.forename: Charley J
authors.initials: CJ
authors.lastname: MacKenzie-Kludas
authors.forename: Anita
authors.initials: A
authors.lastname: Pinar
authors.forename: Weiguang
authors.initials: W
authors.lastname: Zeng
authors.forename: Andrea
authors.initials: A
authors.lastname: Stutz
authors.forename: Eicke
authors.initials: E
authors.lastname: Latz
authors.forename: Lorena E
authors.initials: LE
authors.lastname: Brown
authors.forename: Ashley
authors.initials: A
authors.lastname: Mansell
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002461:Cell Line, Transformed; D005260:Female; D006801:Humans; D058847:Inflammasomes / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007249:Inflammation / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology / Q000821:virology; D009980:Influenza A virus / Q000235:genetics / Q000276:immunology / Q000378:metabolism* / Q000472:pathogenicity; D007251:Influenza, Human / Q000235:genetics / Q000276:immunology / Q000378:metabolism* / Q000503:physiopathology; D053583:Interleukin-1beta / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D014764:Viral Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D037521:Virulence Factors / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; C508585:IL1B protein, human; D058847:Inflammasomes; D053583:Interleukin-1beta; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C453881:NLRP3 protein, human; C492716:Nlrp3 protein, mouse; C442816:PB1-F2 protein, Influenza A virus; D014764:Viral Proteins; D037521:Virulence Factors