Rad9 modulates the P21WAF1 pathway by direct association with p53

Background: Previous studies suggest that human RAD9 ( hRad9), encoding a DNA damage checkpoint molecule, which is frequently amplified in epithelial tumor cells of breast, lung, head and neck cancer, participates in regulation of the tumor suppressor p53- dependent transactivation of pro- survival P21(WAF1). This study examined the exact mechanism of the hRad9 function, especially through the phosphorylation of the C- terminus, in the transcription regulation of P2(1WAF1). Results: The transfection of phosphorylation- defective hRAD9 mutants of C- terminus resulted in reduction of the p53- dependent P21(WAF1) transactivation; the knockdown of total hRad9 elicited an increased P21(WAF1) mRNA expression. Immunoprecipitation and a ChIP assay showed that hRad9 and p53 formed a complex and both were associated with two p53- consensus DNA- binding sequences in the 5' region of P21(WAF1) gene. The association was reduced in the experiment of phosphorylation- defective hRAD9 mutants. Conclusion: The present study indicates the direct involvement of hRad9 in the p53- dependent P21(WAF1) transcriptional mechanism, presumably via the phosphorylation sites, and alterations of the hRad9 pathway might therefore contribute to the perturbation of checkpoint activation in cancer cells.