RT Journal Article SR Electronic T1 Bid and Calpains Cooperate to Trigger Oxaliplatin-Induced Apoptosis of Cervical Carcinoma HeLa Cells JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 998 OP 1010 DO 10.1124/mol.109.058156 VO 76 IS 5 A1 Sergio Anguissola A1 Barbara Köhler A1 Robert O'Byrne A1 Heiko Düssmann A1 Mary D. Cannon A1 Frank E. Murray A1 Caoimhin G. Concannon A1 Markus Rehm A1 Donat Kögel A1 Jochen H. M. Prehn YR 2009 UL http://molpharm.aspetjournals.org/content/76/5/998.abstract AB The Bcl-2 homology 3-only protein Bid is an important mediator of death receptor-induced apoptosis. Recent reports and this study suggest that Bid may also mediate genotoxic drug-induced apoptosis of various human cancer cells. Here, we characterized the role of Bid and the mechanism of Bid activation during oxaliplatin-induced apoptosis of HeLa cervical cancer cells. Small hairpin RNA-mediated silencing of Bid protected HeLa cells against both death receptor- and oxaliplatin-induced apoptosis. Expression of a Bid mutant in which caspase-8 cleavage site was mutated (D59A) reactivated oxaliplatin-induced apoptosis in Bid-deficient cells but failed to reactivate death receptor-induced apoptosis, suggesting that caspase-8-mediated Bid cleavage did not contribute to oxaliplatin-induced apoptosis. Overexpression of bcl-2 or treatment with the pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-dl-Asp-fluoromethylketone abolished caspase-2, -8, -9, and -3 activation as well as Bid cleavage in response to oxaliplatin, suggesting that Bid cleavage occurred downstream of mitochondrial permeabilization and was predominantly mediated by caspases. We also detected an early activation of calpains in response to oxaliplatin. Calpain inhibition reduced Bid cleavage, mitochondrial depolarization, and activation of caspase-9, -3, -2, and -8 in response to oxaliplatin. Further experiments, however, suggested that Bid cleavage by calpains was not a prerequisite for oxaliplatin-induced apoptosis: single-cell imaging experiments using a yellow fluorescent protein-Bid-cyan fluorescent protein probe demonstrated translocation of full-length Bid to mitochondria that was insensitive to calpain or caspase inhibition. Moreover, calpain inhibition showed a potent protective effect in Bid-silenced cells. In conclusion, our data suggest that calpains and Bid act in a cooperative, but mutually independent, manner to mediate oxaliplatin-induced apoptosis of HeLa cells. © 2009 The American Society for Pharmacology and Experimental Therapeutics