The Role Of E3 Ubiquitin Ligase Gp78 And Insig-1 In The Activation Of NLRP3 Inflammasome

NLRP3(NOD-,LRR-and pyrin domain-containing protein 3)inflammasome plays a pivotal role in defending hosts from infection as well as sterile inflammation.The NLRP3 inflammasome must be efficiently activated to cope with the microbial threat and prevent propagation of the infection,but activation must be controlled to prevent hyperinflammation and deleterious consequences for the host organism.Upon activation,the NEK7–NLRP3 interaction increases,then NLRP3 oligomerizes and recruits the ASC protein to nucleate helical ASC filament assembly,resulting in the formation of ASC speck.Assembled ASC recruits pro-caspase-1 and enables the auto-proteolytic activation of pro-caspase-1 into mature caspase-1,then caspase-1 processes inactive pro-IL-1β and pro-IL-18 into mature IL-1β and IL-18.The activation of NLRP3 inflammasome is critically regulated by a deubiquitination mechanism,but little is known about how ubiquitination restrains NLRP3 activity.LPS and ATP/Nigericin induce the activation of NLRP3 inflammasome in peritoneal macrophages.We found that gp78-deficient peritoneal macrophages showed significantly increased caspase-1 maturation and elevated IL-1β and IL-18 secretion compared to control cells.Furthermore,we showed that the membrane-bound E3 ubiquitin ligase gp78 mediates a mixed ubiquitination of NLRP3,which inhibits NLRP3 inflammasome activation by suppressing NLRP3 oligomerization and subcellular translocation.In addition,the ER membrane protein Insulin-induced gene 1(Insig-1)was required for this gp78-NLRP3 interaction and gp78-mediated NLRP3 ubiquitination.gp78 or Insig-1deficiency in myeloid cells led to exacerbated NLRP3 inflammasome-dependent inflammation in vivo,including lipopolysaccharide(LPS)-induced systemic inflammation and alum-induced peritonitis.Taken together,our study elucidates the gp78-mediated NLRP3 ubiquitination as a regulatory mechanism that restrains inflammasome activation and highlights NLRP3 ubiquitination as a potential therapeutic target for inflammatory diseases.