Contribution of caspase-3 differs by p53 status in apoptosis induced by X-irradiation

We investigated the effect of p53 status on involvement of caspase-3 activa tion in cell death induced by X-irradiation, using rat embryonic fibroblast s (REFs) transduced with a temperature-sensitive mutant (mt) p53 gene. Cell s with wild-type (wt) p53 showed greater resistance to X-irradiation than c ells with mt p53. In cells with wt p53, X-irradiation-induced apoptosis was not inhibited by the caspase-3 inhibitor acetyl-L-aspartyl-L-methionyl-L-g lutaminyl-L-aspartyl-aldehyde (Ac-DMQD-CHO) and caspase-3 activity was not elevated following X-irradiation, although induction of p53 and p21/WAF-1 p rotein was observed. In contrast, irradiated cells with mt p53 showed 89% i nhibition of cell death with Ac-DMQD-CHO and 98% inhibition with the antiox idant N-acetyl-L-cysteine (NAC), In cells with mt p53, caspase-3 activity w as increased approximately 5 times beyond baseline activity at 24 h after i rradiation. This increase was almost completely inhibited by NAG. However, inhibition of caspase-3 by Ac-DMQD-CHO failed to decrease production of rea ctive oxygen species by cells with mt p53. Differential involvement of casp ase-3 is a reason for differences in sensitivity to X-irradiation in cells with different p53 status. Caspase-3 activation appears to occur downstream from generation of reactive oxygen species occurring independently of wt p 53 during X-irradiation-induced cell death.