Activation of ATM and phosphorylation of p53 by heat shock.

p53 protein is phosphorylated in response to various stresses. Here we examined phosphorylation of p53 protein in normal human diploid cells after heat shock at 43 degrees C for 2 h. We found that heat shock stimulates phosphorylation of p53 at Ser15 but not at Ser20, while X-irradiation at 4 Gy and 10 J/m(2) of UV induces phosphorylation of p53 at Ser15 and less significantly at Ser20. Increased phosphorylation of Ser15 was also observed in heat shocked GM638, the SV40-transformed human fibroblast cell line. Although X-ray irradiation induced phosphorylation of Ser6, 9, 20, and 37 in GM638 cells, heat shock did not affect the phosphorylation level of these serines. We observed little or no phosphorylation of p53 at Ser15 in two primary ataxia telangiectasia fibroblast cells, that are defective in ATM. Using an in vitro kinase assay, we confirmed that immunoprecipitated ATM from both heat-shocked and X-irradiated normal human diploid cells can phosphorylate p53 at Ser15 to a similar extent. These results indicate that heat shock induces phosphorylation of p53, especially at Ser15, and its phosphorylation is mediated by ATM kinase.