Loss of Caspase-9 Reveals Its Essential Role for Caspase-2 Activation and Mitochondrial Membrane Depolarization

Abstract

Caspase-9 plays an important role in apoptosis induced by genotoxic stress. Irradiation and anticancer drugs trigger mitochondrial outer membrane permeabilization, resulting in cytochrome c release and caspase-9 activation. Two highly contentious issues, however, remain: It is unclear whether the loss of the mitochondrial membrane potential ΔΨMcontributes to cytochrome c release and whether caspases are involved. Moreover, an unresolved question is whether caspase-2 functions as an initiator in genotoxic stress-induced apoptosis. In the present study, we have identified a mutant Jurkat T-cell line that is deficient in caspase-9 and resistant to apoptosis. Anticancer drugs, however, could activate proapoptotic Bcl-2 proteins and cytochrome c release, similarly as in caspase-9–proficient cells. Interestingly, despite these alterations, the cells retained ΔΨM. Furthermore, processing and enzyme activity of caspase-2 were not observed in the absence of caspase-9. Reconstitution of caspase-9 expression restored not only apoptosis but also the loss of ΔΨMand caspase-2 activity. Thus, we provide genetic evidence that caspase-9 is indispensable for drug-induced apoptosis in cancer cells. Moreover, loss of ΔΨMcan be functionally separated from cytochrome c release. Caspase-9 is not only required for ΔΨMloss but also for caspase-2 activation, suggesting that these two events are downstream of the apoptosome.