Caspase-dependent apoptosis and -independent poly(ADP-ribose) polymerase cleavage induced by transforming growth factor beta1.
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-beta1-induced apoptosis and to examine the involvement of caspases and its relationship with PARP cleavage. TGF-beta1 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-beta1-induced apoptosis; but has no effect on TGF-beta1-induced DNA fragmentation and PARP cleavage. However, BD-fmk, a broad-spectrum caspase inhibitor, completely suppresses TGF-beta1-induced apoptosis, but unexpectedly does not inhibit TGF-beta1-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-beta1-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-beta1-induced caspase-dependent apoptosis is associated with caspase-independent PARP cleavage that requires the TGF-beta1-induced synthesis of new proteins. The results indicate that caspase-3 is not a major caspase involved in TGF-beta1-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.
Yang
Yanan
Y
Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yue-Yang Road, 200031, Shanghai, China.
Zhao
Sheng
S
Song
Jianguo
J
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D002469:Cell Separation; D003513:Cycloheximide / Q000494:pharmacology; D053938:DNA Fragmentation; D003630:Daunorubicin / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D022781:Hepatocytes / Q000473:pathology; D015151:Immunoblotting; D051379:Mice; D008856:Microscopy, Fluorescence; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D051745:Protein Kinase C-delta; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1
D000590:Amino Acid Chloromethyl Ketones; D004791:Enzyme Inhibitors; D011500:Protein Synthesis Inhibitors; C508961:Tgfb1 protein, mouse; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D003513:Cycloheximide; D011065:Poly(ADP-ribose) Polymerases; C497259:Prkcd protein, mouse; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D020169:Caspases; D003630:Daunorubicin