J Immunol. 2003 Dec 1;171(11):6154-63.

Apoptosis-associated speck-like protein containing a caspase recruitment domain is a regulator of procaspase-1 activation.External

Stehlik, C., Lee, S. H., Dorfleutner, A., Stassinopoulos, A., Sagara, J., Reed, J. C.,
--- - Burnham Institute, La Jolla, CA 92037, USA.
Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC)/target of methylation-induced silencing/PYCARD represents one of only two proteins encoded in the human genome that contains a caspase recruitment domain (CARD) together with a pyrin, AIM, ASC, and death domain-like (PAAD)/PYRIN/DAPIN domain. CARDs regulate caspase family proteases. We show here that ASC binds by its CARD to procaspase-1 and to adapter proteins involved in caspase-1 activation, thereby regulating cytokine pro-IL-1beta activation by this protease in THP-1 monocytes. ASC enhances IL-1beta secretion into the cell culture supernatants, at low concentrations, while suppressing at high concentrations. When expressed in HEK293 cells, ASC interferes with Cardiak/Rip2/Rick-mediated oligomerization of procaspase-1 and suppresses activation this protease, as measured by protease activity assays. Moreover, ASC also recruits procaspase-1 into ASC-formed cytosolic specks, separating it from Cardiak. We also show that expression of the PAAD/PYRIN family proteins pyrin or cryopyrin/PYPAF1/NALP3 individually inhibits IL-1beta secretion but that coexpression of ASC with these proteins results in enhanced IL-1beta secretion. However, expression of ASC uniformly interferes with caspase-1 activation and IL-1beta secretion induced by proinflammatory stimuli such as LPS and TNF, suggesting pathway competition. Moreover, LPS and TNF induce increases in ASC mRNA and protein expression in cells of myeloid/monocytic origin, revealing another level of cross-talk of cytokine-signaling pathways with the ASC-controlled pathway. Thus, our results suggest a complex interplay of the bipartite adapter protein ASC with PAAD/PYRIN family proteins, LPS (Toll family receptors), and TNF in the regulation of procaspase-1 activation, cytokine production, and control of inflammatory responses.
PMID: 14634131External
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
In vitro interaction
  DD1 DD2 Reference
Family DD1 DD2 Method Species Region Expression Species Region Expression
CARD ASC Link ASC GST fusion protein pull-down Not specified Not specified E.coli Not specified Not specified In vitro translation 14634131
CARD ASC Link Caspase1 GST fusion protein pull-down Not specified Not specified E.coli Not specified Not specified In vitro translation 14634131
(Link: click this icon to show interactions only between the two corresponding DDs)
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
In vivo interaction
  Endogenous
expression
Overexpression DD1 DD2 Reference
Family DD1 DD2 Method Species Region Species Region
CARD ASC Link Caspase1 Co-immunoprecipitation HEK293 Not specified Not specified Not specified Not specified 14634131
CARD ASC Link Caspase1 Co-immunoprecipitation THP-1 14634131
CARD ASC Link RIPK2 Co-immunoprecipitation HEK293 Not specified Not specified Not specified Not specified 14634131
(Link: click this icon to show interactions only between the two corresponding DDs)
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
Characterization
  Stoichiometry Affinity Protein-Protein interface Reference
Family DD1 DD2 Method Mutation Complex structure
CARD ASC Link Caspase1 Co-immunoprecipitation L122Q(ASC) 14634131
CARD ASC Link RIPK2 Co-immunoprecipitation L122Q(ASC) 14634131
(Link: click this icon to show interactions only between the two corresponding DDs)
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
Functional Role
Family DD1 DD2 Method Death-related Death-unrelated Reference
CARD ASC Link Caspase1 ELISA IL-1b secretion decrease 14634131
CARD ASC Link RIPK2 Immunoprecipitation Pro-caspase1-RIPK2 complex inhibition 14634131
(Link: click this icon to show interactions only between the two corresponding DDs)