Phosphorylation sites in the amino-terminal region of mouse p53

Abstract

Phosphorylation is an attractive mechanism for regulating the functions of p53. The p34cdc2 kinase, which is involved in regulation of the cell cycle, phosphorylates serine-315 of human p53 in vitro. Casein kinase II phosphorylates serine-389 of mouse p53 in vitro. The amino-terminal region of mouse p53 contains a cluster of potential serine phosphorylation sites. Those sites have been proposed to be sites for phosphorylation by a double-stranded DNA-dependent kinase (DNA-PK) from HeLa cells and can be dephosphorylated by protein phosphatase 2A. To identify in vivo phosphorylation sites in the amino-terminal region of mose p53, we mutated potential phosphorylation sites and analyzed the mutant proteins by tryptic phosphopeptide mapping. We identified serine-7, -9, -18, and -37 as in vivo phosphorylation sites. We further showed that mouse p53 expressed in bacteria is phosphorylated by DNA-PK on aminoterminal serine residues in vitro.