DNA-dependent protein kinase (DNA-PK) is composed of a 460-kDa catalytic component (p460) and a DNA-binding component Ku protein. Immunoblot analysis after treatment of Jurkat cells with anti-Fas antibody demonstrated the cleavage of p460 concomitantly with an increase in CPP32/Yama/apopain activity. Recombinant CPP32/Yama/apopain specifically cleaved p460 in the DNA-PK preparation that had been purified from Raji cells into 230- and 160-kDa polypeptides, the latter of which was detected in anti-Fas-treated Jurkat cells. The regulatory component Ku protein was not significantly affected by CPP32/Yama/apopain. DNA-PK activity was decreased with the disappearance of p460 in the incubation of DNA-PK with CPP32/Yama/apopain. These results suggest that the catalytic component of DNA-PK is one of the target proteins for CPP32/Yama/apopain in Fas-mediated apoptosis.