Tumor Suppressor p53 Can Participate in Transcriptional Induction of the GADD45 Promoter in the Absence of Direct DNA Binding

The GADD45 gene is a growth arrest-associated gene that is induced by certain DNA-damaging agents and other stresses, such as starvation, in all mammalian cells. In addition to a strong p53-binding element in an intronic sequence, we have recently found that p53, while not required or sufficient alone, may contribute to the stress responsiveness of the promoter. Much of the responsiveness was localized to a GC-rich motif in the proximal promoter which contains multiple Egr1 sites and a larger WT1 site; this 20-bp WT1 motif is identical to the WT1-binding site in the PDGF-A gene. In extracts from a human breast carcinoma cell line expressing p53 and WT1, which is known to associate with p53 in vivo, evidence was obtained that these proteins are in a complex that binds this 20-bp element. A combination of p53 and WT1 expression vectors strongly induced a GADD45-reporter construct, while mutation of the WT1-Egr1 site in the promoter prevented this induction. Abrogation of p53 function by a dominant-negative vector or abrogation of WT1 function by an antisense vector markedly reduced the induction of this promoter. Since p53 does not bind directly to the promoter, these results indicate that p53 can contribute to the positive regulation of a promoter by protein-protein interactions.