Highly regenerative formerly tissues this kind of as blood will have to possess effective DNA harm responses (DDR) that stability long-term regeneration with safety from leukemogenesis. Hematopoietic stem cells (HSCs) sustain life-long blood production, nonetheless their response to E3 Ligase DNA damage stays largely unexplored. We report that human HSCs exhibit delayed DNA double-strand break rejoining, persistent gamma H2AX foci, and enhanced p53- and ASPP1-dependent apoptosis right after gamma-radiation when compared to progenitors. p53 inactivation or Bcl-2 overexpression decreased radiation-induced apoptosis and preserved in vivo repopulating HSC function. Despite very similar protection from irradiation-induced apoptosis, only Bcl-2-overexpressing HSCs showed higher self-renewal capability, establishing that intact p53 positively regulates self-renewal independently from apoptosis. The reduced self-renewal of HSCs with inactivated p53 was related with increased spontaneous gamma H2AX foci in secondary transplants of HSCs. Our data reveal distinct physiological roles of p53 that collectively assure optimal HSC function: apoptosis regulation and prevention of gamma H2AX foci accumulation on HSC self-renewal.