Caspase-dependent apoptosis and -independent poly(ADP-ribose) polymerase cleavage induced by transforming growth factor β1

Abstract

Apoptosis is an important cell suicide program which involves the caspases activation and is implicated in physiological and pathological processes. Poly(ADP-ribose) polymerase (PARP) cleavage is often associated with apoptosis and has been served as one hallmark of apoptosis and caspase activation. In this study, we aimed to determine TGF-β1-induced apoptosis and to examine the involvement of caspases and its relationship with PARP cleavage. TGF-β1 induces strong apoptosis of AML-12 cells which can be detected by DNA fragmentation, FACS, and morphological assays. Z-VAD-fmk, a selective caspase inhibitor, partially inhibits the TGF-β1-induced apoptosis; but has no effect on TGF-β1-induced DNA fragmentation and PARP cleavage. However, BD-fmk, a broad-spectrum caspase inhibitor, completely suppresses TGF-β1-induced apoptosis, but unexpectedly does not inhibit TGF-β1-induced PARP cleavage. Furthermore, Z-VAD-fmk treatment is able to completely inhibit the daunorubicin-induced apoptosis in A-431 cells, but only slightly blocks the daunorubicin-induced PARP cleavage, whereas BD-fmk can inhibit both daunorubicin-induced apoptosis and PARP cleavage completely. In addition, we observed that both TGF-β1-induced apoptosis and PARP degradation in AML-12 cells can be completely blocked by inhibiting the protein synthesis with cycloheximide. These results demonstrate for the first time that TGF-β1-induced caspase-dependent apoptosis is associated with caspase-independent PARP cleavage that requires the TGF-β1-induced synthesis of new proteins. The results indicate that caspase-3 is not a major caspase involved in TGF-β1-induced apoptosis in AML-12 cells, and is not required for apoptosis-associated DNA fragmentation. The results also suggest that PARP cleavage may occur as an independent event that can be disassociated with cell apoptosis.