CLEAVAGE AND INACTIVATION OF DNA-DEPENDENT PROTEIN-KINASE CATALYTIC SUBUNIT DURING APOPTOSIS IN XENOPUS EGG EXTRACTS

DNA-dependent protein kinase (DNA-PK) consists of a 460 kDa subunit th at contains the catalytic domain (DNA-PKcs) complexed with two polypep tides of 70 kDa and 80 kDa (Ku70 and Ku80) which comprise the Ku autoa ntigen, DNA-PKcs requires association with DNA via Ku for catalytic ac tivation and is implicated in double strand break repair, V(D)J recomb ination and transcription, We have utilised a cell-free system of conc entrated Xenopus laevis egg extracts to investigate the regulation and possible functions of DNA-PK, Recently, we have shown that this syste m can reproduce events of apoptosis, including activation of an apopto tic protease that cleaves poly(ADP-ribose) polymerase, Here, we report that DNA-PK is rapidly inactivated with the onset of apoptosis in sys tem, Loss of activity is concomitant with cleavage of the catalytic su bunit, whereas the Ku subunits are stable. Cleavage and inactivation o f DNA-PKcs is prevented by prior addition of the anti-apoptotic protei n Bcl-2 or inhibition of an apoptotic protease that has characteristic s of the CPP-32/Ced-3 family of cysteine proteases that cleave poly(AD P-ribose) polymerase. These results suggest that cleavage and inactiva tion of DNA-PKcs prevents this factor from functioning in DNA repair, recombination or transcriptional regulation during apoptosis.