1. ¹Center for the Development of Molecular Target Drugs, Cancer Research Institute, Kanazawa University, Kanazawa, Ishikawa, Japan
2. ²Department of Pathology, The University of Michigan Medical School, Ann Arbor, MI, USA
3. ³Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada

Correspondence: Dr T Suda, Center for the Development of Molecular Target Drugs, Cancer Research Institute, Kanazawa University, 13-1 Takaramachi, Kanazawa, Ishikawa 920-0934, Japan. E-mail: sudat@kenroku.kanazawa-u.ac.jp

⁴Current address: Department of Pathology, The University of Michigan Medical School, Ann Arbor, Michigan 48109, USA.

Received 10 December 2005; Revised 4 August 2006; Accepted 4 August 2006; Published online 11 September 2006.

Abstract

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.