Bcl-2 and caspase inhibition cooperate to inhibit tumor necrosis factor-alpha-induced cell death in a Bcl-2 cleavage-independent fashion

The ability of proteins of the Bcl-2 family to either induce or inhibit apo ptosis is dependent on both cell type and the apoptotic stimulus. We have s hown in the murine pro-B cell line FL5.12 that Bcl-2 is incapable of inhibi ting tumor necrosis factor alpha (TNF alpha)-induced cell death and is clea ved during this process. One potential explanation for this observation is that caspase activation directly or indirectly inhibits Bcl-2 function. It has been suggested that caspase cleavage of Bcl-2 is responsible for its in ability to block certain cell deaths. Consistent with Bcl-2 cleavage being a caspase-mediated event, this cleavage is inhibitable by 50 mu M CBZ-Val-A la-Asp-fluoromethylketone (zVAD-fmk). Furthermore, Bcl-2 can cooperate with the caspase inhibitor zVAD-fmk in a dose-dependent manner to block TNF alp ha-induced cell death. Overexpression of Bcl-2 results in a 10-fold decreas e in the amount of zVAD-fmk required to inhibit TNF alpha-induced apoptosis . However, cleavage-defective mutants (D31A and D34A) show no enhanced viab ility relative to wild-type Bcl-2 in response to TNF alpha-induced cell dea th and also show the same cooperativity with zVAD-fmk. These results sugges t that Bcl-2 cleavage is not important for the inhibition of TNF alpha-indu ced cell death but do not preclude an involvement in a post-commitment phas e of apoptosis.