Constitutive expression of the cyclin-dependent kinase inhibitor p21 is transcriptionally regulated by the tumor suppressor protein p53

The tumor suppressor protein p53 has been implicated in the response of cells to DNA damage. Studies to date have demonstrated a role for p53 in the transcriptional activation of target genes in the cellular response to DNA damage that results in either growth arrest or apoptosis, In contrast, here is demonstrated a role for p53 in regulating the basal level of expression of the cyclin-dependent kinase inhibitor p21 in the absence of treatment with DNA-damasng agents. Wild-type p53-expressing MCF10F cells had detectable levels of p21 mRNA and protein, whereas the p53-negative Saos-2 cells did not. Saos-2 cells were infected with recombinant retrovirus to establish a proliferating pool of cells with a comparable constitutive level of expression of wild-type p53 protein to that seen in untreated MCF10F cells. Restoration of wild-type but not mutant p53 expression recovered a basal level of expression of p21 in these cells. Constitutive expression of luciferase reporter constructs containing the p21 promoter was inhibited by co-transfection with the human MDM2 protein or a dominant-negative p53 protein and was dependent on the presence of p53 response elements in the reporter constructs. Furthermore, p53 in nuclear extracts of untreated cells was capable of binding to DNA in a sequence-specific manner. These results implicate a role for p53 in regulating constitutive levels of expression of p21 and demonstrate that the p53 protein is capable of sequence-specific DNA binding and transcriptional activation in untreated, proliferating cells.