Mechanism of ASC-mediated apoptosis: bid-dependent apoptosis in type II cells

Apoptosis-associated speck-like protein containing a CARD (ASC) is an adaptor molecule that mediates apoptotic and inflammatory signals, and implicated in tumor suppression. However, the mechanism of ASC-mediated Apoptosis has not been well elucidated. Here, we investigated the molecular mechanisms of ASC-mediated Apoptosis in several cell lines using a Caspase recruitment domain 12-Nod2 chimeric protein that transduces the signal from muramyl dipeptide into ASC-mediated Apoptosis. Experiments using dominant-negative mutants, small-interfering RNAs and peptide inhibitors for caspases indicated that Caspase-8 was generally required for ASC-mediated Apoptosis, whereas a requirement for caspase-9 depended on the cell type. In addition, caspase-like apoptosis-regulatory protein (CLARP)/Fas-like inhibitor protein, a natural Caspase-8 inhibitor, suppressed ASC-mediated Apoptosis, and Clarp-/- mouse embryonic fibroblasts were highly sensitive to ASC-mediated Apoptosis. Bax-deficient HCT116 cells were resistant to ASC-mediated Apoptosis as reported previously, although we failed to observe colocalization of ASC and Bax in cells. Like Fas-ligand-induced Apoptosis, the ASC-mediated Apoptosis was inhibited by Bcl-2 and/or Bcl-xL in type-II but not type-I cell lines. Bid was cleaved upon ASC activation, and suppression of endogenous Bid expression using small-interfering RNAs in type-II cells reduced the ASC-mediated Apoptosis. These results indicate that ASC, like death receptors, mediates two types of Apoptosis depending on the cell type, in a manner involving Caspase-8.