Cleavage of Bax enhances its cell death function

Members of the Bcl-2 family of proteins are key regulators of apoptosis. So me of these proteins undergo posttranslational modification, such as phosph orylation or proteolysis, that serves to alter their function. Caspases are known to cleave Bid, a proapoptotic family member, as well as Bcl-2 and Bc l-X-L, two prosurvival family members, which activate their cytotoxic activ ity resulting in the release of cytochrome c from mitochondria. Previously we showed that Bax was cleaved by calpain rather than by caspases from full -length 21 kDa to generate a cleavage fragment of 18 kDa. Since cleavage of Bid serves to activate its cytotoxic activity, we wanted to determine if t he p18 form of Bax exhibited increased cytotoxicity compared to pal Bax. Us ing a transient transfection system in human embryonic kidney 293T cells we show that the p18 form of Bax displays a more potent ability to induce cel l death. The pancaspase inhibitor Z-VAD-fmk completely blocked apoptosis in duced by p21 Bax but only partially inhibited apoptosis induced by p18 Bax. Cyclosporin A, an inhibitor of the mitochondrial permeability transition ( PT) pore, had no effect on Bax-mediated apoptosis of 293T cells suggesting that apoptosis was independent of the PT. Thus cleavage of pal Bax during a poptosis to the p18 form may serve to increase the intrinsic cytotoxic prop erties of this proapoptotic molecule and enhance its cell death function at the mitochondria. (C) 2000 Academic Press.