THE APOPTOTIC AND TRANSCRIPTIONAL TRANSACTIVATION ACTIVITIES OF P53 CAN BE DISSOCIATED

Previous studies have shown that the apoptotic response of cells follo wing DNA damage requires p53 expression. Wild-type p53 protein levels increase in response to DNA damage and its growth-suppressive action i s thought to be mediated by transcriptional activation of the p21/WAF1 /CIP1 gene, the product of which is a potent inhibitor of cyclin-depen dent kinases. The mechanism by which elevated p53 levels lead to apopt osis is not known, but is believed to involve transcriptional activati on of apoptotic genes, such as BAX. We have studied transformed human cells that constitutively express high levels of the R273H mutant p53, which has been reported to lack transcriptional activation activity. We used the inability to induce the p21/Waf1/Cip1 protein as a marker to verify the lack of transcriptional activation activity. Cells expre ssing the R273H mutant of p53 do not show an increase in p21/Waf1/Cip1 following irradiation with ionizing or UVB radiation. Surprisingly, t hese cells are very susceptible to induction of apoptosis by UVB radia tion, as seen by the formation of a nucleosomal ladder and the proteol ytic cleavage of poly(ADP-ribose) polymerase. This suggests that the R 273 mutant p53 can function normally in apoptosis but not in transcrip tional activation following DNA damage. Furthermore, an inhibitor of R NA polymerase II is a potent inducer of apoptosis in these cells, demo nstrating that transcription is not required for apoptosis and suggest ing that stalled RNA polymerase II complexes can initiate apoptosis. I nterestingly, proteolytic cleavage of p53 occurs during apoptosis in t hese cells, generating a 45-kDa fragment and liberating the DNA repair helicase binding domain of p53. We propose that the peptide liberated from the carboxy terminus of p53 may contribute to its apoptotic acti vity, possibly through interaction with the XPB and XPD DNA helicases.