INCREASED SEQUENCE-SPECIFIC P53 DNA-BINDING ACTIVITY AFTER DNA-DAMAGEIS ATTENUATED BY PHORBOL ESTERS

Damage to cellular DNA greatly increases the levels of the tumor-suppr essor gene p53 and induces cell cycle arrest in G1. A critical functio n of wild-type p53 is its ability to bind to specific DNA sequences. T he effect of DNA damage on the sequence-specific DNA-binding propertie s of cellular p53 was investigated using DNA gel mobility-shift assays with nuclear extracts from NIH-3T3 cells. DNA damage (initiated by ra diation) induced a rapid, cycloheximide-sensitive increase in the leve ls of nuclear p53-DNA binding activity and an increase in the half-lif e of the p53 protein. Increased p53-DNA binding activity could be dete cted at low (0.2 Gy), non-lethal doses of radiation. The tumor promote r 12-O-tetradecanoyl phorbol 13-acetate (TPA) attenuated the DNA damag e-induced increase in p53-DNA binding activity by decreasing the half- life of the p53 protein. The tumor promoter properties of TPA may ther efore be mediated by interfering with the cellular p53 response to DNA damage. The increased levels of p53 bound to specific DNA sequences f ollowing DNA damage may induce cell cycle arrest. p53-mediated growth arrest could occur by inhibition of DNA replication and/or alterations in transcription of cell cycle genes.