Cryopyrin and pyrin activate caspase-1, but not NF-κB, via ASC oligomerization

Abstract

Mutations in cryopyrin and pyrin proteins are responsible for several autoinflammatory disorders in humans, suggesting that these proteins play important roles in regulating inflammation. Using a HEK293 cell-based reconstitution system that stably expresses ASC and procaspase-1 we demonstrated that neither cryopyrin nor pyrin or their corresponding disease-associated mutants could significantly activate NF-jB in this system. However, both cryopyrin and two disease-associated cryopyrin mutants induced ASC oligomerization and ASC-dependent caspase-1 activation, with the disease-associated mutants being more potent than the wild-type (WT) cryopyrin, because of increased selfoligomerization. Contrary to the proposed anti-inflammatory activity of WT pyrin, our results demonstrated that pyrin, like cryopyrin, can also assemble an inflammasome complex with ASC and procaspase-1 leading to ASC oligomerization, caspase-1 activation and interleukin-1b processing. Thus, we propose that pyrin could function as a proinflammatory molecule.

Identification and functional consequences of a recurrent NLRP12 missense mutation in periodic fever syndromes

“…Indeed, our results show that the p.Arg352Cys missense mutation is associated with a gain of function of caspase 1 processing, while the previously reported nonsense and splice site mutations lead to a loss of function of NF-B inhibition (1). Similar observations were made for missense and nonsense mutations identified in NLRP3 (6,7,12).…”

“…Since missense mutations identified in NLRP3 activate the proteolytic cleavage of procaspase 1 into caspase 1 (6,7), we studied the role of wild-type and mutant NLRP12 proteins in caspase 1 processing. Unlike cell systems in which ASC and procaspase 1 are transiently expressed, we used a previously validated system of HEK 293T cells stably expressing these proteins at physiologic levels (293-C1-ASC) (6,7). Transfection of these cells with pNLRP12-WT or pNLRP12-Arg352Cys induced the processing of procaspase 1 (Figure 2).…”

“…Previous studies have shown that NLRP3 NBS mutants display increased abilities to induce speck formation in the presence of ASC (6,7). To test the effect of p.Arg352Cys on NLRP12-ASC interaction and speck formation, we transiently transfected 293-C1-ASC_GFP cells either with an empty vector or with plasmids encoding normal or mutant NLRP12 proteins.…”

The estrogen-responsive B box protein: a novel enhancer of interleukin-1β secretion

“…This secretion-promoting effect of EBBP for caspase-1 was much stronger after cotransfection of pro-and mature IL-1b expression plasmids (lanes 13-16, 21-24, supernatant). Highest secretion was observed after coexpression of the cytokine with either EBBP or individual EBBP domains (lanes [14][15][16][22][23][24]. Surprisingly, forced expression of EBBP and pro-and to a lesser extent mature IL-1b resulted in degradation of procaspase-1 (lanes 14, 15, 22, 23, lysate and supernatant).…”

“…21 It is not known how the disease-causing mutations, which locate mainly to the carboxy-terminal RFP (SPRY; B30.2) domain, influence the amino-terminal pyrin domain. 22,23 Furthermore, recent results suggest that Pyrin activates caspase-1 via Asc oligomerization, 24 suggesting that it is an inflammasome activator. Apart from its pyrin domain, Pyrin is a typical member of the TRIM (tripartite motif) family.…”