Following genotoxic stress, p53 either rescues a damaged cell or promotes its elimination. The parameters determining a specific outcome of the p53 response are largely unknown. In mouse fibroblasts treated with different irradiation schemes, we monitored transcriptional and non-transcriptional p53 activities and identified determinants that initiate an anti- or a pro-apoptotic p53 response within the context of p53-independent stress signaling. The primary, transcription-mediated p53 response in these cells is anti-apoptotic, while induction of p53-dependent apoptosis requires an additional, transcription-independent p53 activity, provided by high intracellular levels of activated p53. High intracellular levels of p53 were selectively generated after apoptosis-inducing high-dose UV-irradiation, and correlated with a strongly delayed upregulation of Mdm2. Following high-dose UV-irradiation, p53 accumulated in the cytoplasm and led to activation of the pro-apoptotic protein Bax. As p53-dependent Bax-activation is transcription-independent, we postulated that certain transcription-deficient mutant p53 proteins might also exert this activity. Indeed we found an endogenous, transcription-inactive mutant p53 that upon genotoxic stress induced Bax-activation in vivo. Our results demonstrate the impact and in vivo relevance of non-transcriptional mechanisms for wild-type and mutant p53-mediated apoptosis.Oncogene advance online publication, 10 October 2005; doi:10.1038/sj.onc.1209126.