DNA-dependent protein kinase-independent activation of p53 in response to DNA damage

Phosphorylation at serine 15 of the human p53 tumor suppressor protein is i nduced by DNA damage and correlates with accumulation of p53 and its activa tion as a transcription factor. The DNA-dependent protein kinase (DNA-PK) c an phosphorylate serine 15 of human p53 and the homologous serine 18 of mur ine p53 in vitro, Contradictory reports exist about the requirement for DNA -PK in vivo for p53 activation and cell cycle arrest in response to ionizin g radiation. While primary SCID (severe combined immunodeficiency) cells, t hat have defective DNA-PK, show normal p53 activation and cell cycle arrest , a transcriptionally inert form of p53 is induced in the SCID cell line SC GR11. In order to unambiguously define the role of the DNA-PK catalytic sub unit (DNA-PKcs) in p53 activation, we examined p53 phosphorylation in mouse embryonic fibroblasts (MEFs) from DNA-PKcs-null mice. We found a similar p attern of serine 18 phosphorylation and accumulation of p53 in response to irradiation in both control and DNA-PKcs-null MEFs, The induced p53 was cap able of sequence-specific DNA binding even in the absence of DNA-PKcs. Tran sactivation of the cyclin-dependent-kinase inhibitor p21, a downstream targ et of p53, and the G(1) cell cycle checkpoint were also found to be normal in the DNA-PKcs -/- MEFs. Our results demonstrate that DNA-PKcs, unlike the related ATM protein, is not essential for the activation of p53 and G(1) c ell cycle arrest in response to ionizing radiation.