Activation of caspases in p53-induced transactivation-independent apoptosis

Though p53-induced apoptosis plays an important role in tumor suppression, the mechanism(s) by which p53 induces apoptosis is still unclear. To elucid ate the p53-induced apoptotic pathway, we examined the role of p53 transact ivation activity and caspase in J138V5C cells carrying a human temperature- sensitive (ts) p53 mutant (138Ala-->Val). The results showed that p53-induc ed apoptosis was not blocked by cycloheximide, which effectively prevented the expression of p53 target genes, indicating that transactivation was not essential for p53-induced apoptosis in this system. Western blot analysis showed that PARP, CPP32 and ICH-I precursors were cleaved during apoptosis, The CPP32-preferential tetrapeptide inhibitor Ac-DEVD-CHO blocked the clea vage of ICH-1 and PARP precursors, suggesting that CPP32 or some other DEVD -sensitive caspase(s) is the upstream activator of ICH-1. We also examined the role of the Fas pathway by using Fas and Fas ligand-neutralizing antibo dies. Both antibodies failed to block p53-induced apoptosis, suggesting tha t the Fas pathway was not essential for p53-induced apoptosis in this syste m. Taken together, our results indicate that p53-induced, transactivation-i ndependent apoptosis in Jurkat cells involves sequential activation of CPP3 2 or some other DEVD-sensitive caspase(s) and ICH-1, via a Fas-independent pathway.