MUTATION OF PHOSPHOSERINE-389 AFFECTS P53 FUNCTION IN-VIVO

To study the importance of phosphorylation for p53 transactivation fun ction, we generated mutations at each of its known phosphorylated seri ne amino acids. Mutations of murine p53 serine residues individually t o either alanine or glutamic acid at positions 7, 9, 12, 18, 37, 312, and 389 resulted in equivalent levels of transcriptional activation in standard transient transfection experiments. However, when p53 transc riptional activity was measured in cells that attain G(1) arrest upon contact inhibition, wild-type p53 was inactive, and only alteration at serine 389 to glutamic acid resulted in a functional p53 protein. Thi s Ser --> Glu mutant also has an increased ability to bind DNA. Elimin ation of the phosphorylation site by substitution of an alanine amino acid resulted in loss of transcriptional activity. We also demonstrate d that specific phosphorylation of p53 at serine 389 is induced by cyc lin E overexpression in high-density cells. Our data establish for the first time that phosphorylation of p53 at serine 389 is important in activating its function in vivo.