POSITIVE AND NEGATIVE REGULATORY ELEMENTS IN THE MURINE P53 PROMOTER

In order to understand the basis for regulated as well as de-regulated expression of the p53 tumor suppressor gene, we have focused on chara cterizing the transcriptional regulation of the p53 gene. Here we pres ent evidence for the existence of two additional upstream regulatory e lements in the murine p53 promoter. One of these sites maps to a regio n between -296 to -270 and the second one between -255 to -226 relativ e to the major transcription initiation site. These two sites are refe rred to as binding sites for PBF I and II, respectively. Nucleotide ba ses that have been found to be critical for the binding of nuclear fac tors to these sites are 5'-AGA-3' (-282 to -280) in binding site I and 5'-ACAG-3' (-246 to -243) in binding site II. Mutational analyses in conjunction with transient transfection assays indicated that the fact or that binds to the region between -245 to -242 (PBF II) plays a posi tive regulatory role p53 promoter activity. This was demonstrated by t he observation that promoter mutations that abolished binding to this site, showed a decreased level of activity as compared to the wild typ e promoter. In analogous experiments, mutational anlayses and transien t transfection assays indicated that the factor that binds to the regi on between -282 to -280 (PBF I) plays a negative regulatory role in p5 3 promoter activity. This was demonstrated by the observation that pro moter mutations that abolished binding to this site, showed an increas ed level of activity as compared to the wild type promoter.