title: The mechanism of NLRP3 inflammasome initiation: Trimerization but not dimerization of the NLRP3 pyrin domain induces robust activation of IL-1β.
abstract: NLRP3 inflammasome is a multiprotein platform for the activation of caspase-1. Despite the increasing number of reports linking NLRP3 inflammasome to a variety of diseases, the mechanism behind the NLRP3 activation remains elusive, especially in terms of the early stages which are critical to the NLRP3 inflammasome assembly. In the present study we aimed to determine the minimal oligomerization state required for the NLRP3 inflammasome activation. For this purpose, NLRP3 pyrin domain (NLRP3PYD) was fused to various dimerization and trimerization domains. The constructs were expressed under the inducible promoter in mouse macrophages lacking endogenous NLRP3. Dimerization of the NLRP3PYDeither in parallel or in antiparallel orientation was insufficient for the inflammasome activation. Trimerization of the NLRP3PYDwith the foldon domain, however, induced pyroptosis and robust IL-1β maturation, which was caspase-1 dependent. Interestingly, foldon-induced constitutive activation is resistant to inhibition with NLRP3-specific inhibitor MCC950 and does not lead to ASC speck formation. Although we cannot exclude that wild-type NLRP3 forms higher oligomer species similar to NLRP1 or NLRC4, our results clearly demonstrate that efficient IL-1β response can be achieved by the induced trimerization of the NLRP3PYDdomain.
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana, Slovenia; Graduate School of Biomedicine, University of Ljubljana, 1000 Ljubljana, Slovenia.
authors.forename: Petra
authors.initials: P
authors.lastname: Sušjan
authors.affiliation: Uniform Second-Level Master's Program of Medicine, Medical Faculty, University of Ljubljana, Ljubljana 1000, Slovenia.
authors.forename: Samo
authors.initials: S
authors.lastname: Roškar
authors.affiliation: Department of Synthetic Biology and Immunology, National Institute of Chemistry, Hajdrihova 19, 1000 Ljubljana, Slovenia; EN-FIST Centre of Excellence, Trg Osvobodilne Fronte 13, Ljubljana, Slovenia. Electronic address: iva.hafner@ki.si.
authors.forename: Iva
authors.initials: I
authors.lastname: Hafner-Bratkovič
mesh_terms: D000818:Animals; D020170:Caspase 1 / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D058847:Inflammasomes / Q000276:immunology / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism*; D051379:Mice; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000072417:Protein Domains; D055503:Protein Multimerization; D000071198:Pyrin / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: C587672:IL1B protein, mouse; D058847:Inflammasomes; D053583:Interleukin-1beta; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; D000071198:Pyrin; D011993:Recombinant Fusion Proteins; D020170:Caspase 1
keywords: Inflammasome; NLRP3; Oligomerization; Pyrin