Inhibitor of apoptosis protein hILP undergoes caspase-mediated cleavage during T lymphocyte apoptosis

Several endogenous or viral inhibitors of apoptosis, including Bcl-2, Bcl-x (L),, FLIP, p35, and CrmA, have been shown to be cleaved by caspases during apoptosis, In this study, we demonstrate that the endogenous inhibitor of apoptosis, hILP/XIAP, is also cleaved in apoptotic T lymphocytes, generatin g at least one prominent fragment of 29 kDa, This p29 cleaved fragment was detected in Jurkat cells induced to apoptose by anti-Fas antibody, staurosp orin, or VP-16. The cleavage of hILP appears to be caspase mediated because the production of the p29 protein was inhibited by the pan-caspase peptide inhibitor, Z-VAD.FMK. In Jurkat cells engineered to overexpress CrmA, clea vage of hILP in response to anti-Fas antibody or staurosporin was inhibited , whereas overexpression of Bcl-2 abrogated the cleavage in response to VP- 16, Cleavage of hILP was also observed in cell-free reactions using in vitr o translated hILP and recombinant caspase-3 or -7. Moreover, we found that the p29 hILP fragment retained the ability to bind caspase-3 and -7, as sho wn previously For full-length or BIR-2 hILP, The p29 cleavage product was a lso detected during T-cell receptor-mediated apoptosis in peripheral blood lymphocytes from normal donors. Furthermore, tumor-associated T lymphocytes purified from ascites of patients with ovarian cancer expressed fragmented hILP, which was not detected in control T cells purified from peripheral b lood of normal donors. Our results suggest that the cleavage of hILP repres ents an important event in apoptosis of T lymphocytes in both normal and pa thological in vivo settings.