The tumor suppressor p53 is a master regulatory gene that directly controls differential expression of target genes in a sequence-specific manner from promoter response elements (RE). Mutations in p53 that alter transactivation functions, such as strength of activation of various genes and/or spectrum of genes regulated, may lead to changes in the p53 transcriptional network (1), which could result in phenotypic changes in cellular stress responses. We are determining the functional fingerprints of mutant p53s associated with various cancers, including breast cancer, Li Fraumeni syndrome (LFS) and adrenal cortical carcinoma (ACC) in terms of p53 transactivation at various REs in human promoters. To assess the ability of mutant p53 to interact with REs in vivo, we utilized a yeast model that allows for rapid analysis of p53 function within chromatin under conditions of variable expression (1). The previously described system that examines transactivation capacities of wild-type and mutant p53 towards many REs under isogenic conditions has been expanded to include a quantitative reporter. Surprisingly, the amount of wild-type p53 expression required to initially induce transactivation was comparable for all REs regardless of transactivation at saturating amounts of p53, indicating that differences in transactivation capacity in vivo are not simply determined by on-rates. These results challenge current notions that the ability of wild-type p53 to transactivate from various REs is simply due to differences in RE binding affinities. Among 24 non-hotspot p53 mutants from sporadic, BRCA1/2- and LFS-associated breast cancers, 9 showed altered function–not just complete loss–towards at least one of the REs. The functional mutants examined were in the DNA binding (i.e., H214R) or tetramerization (i.e., R337C) domain. Often the changes were detected only at low p53 levels. The induction patterns in the quantitative assay were consistent with the functional mutations simply modulating transactivation levels. If these observations translate to human cells, then levels of p53 could determine changes in the spectrum of genes transactivated and the extent of individual gene transactivation. Interestingly, the ACC mutation R337H exhibited a very different pattern of transactivation in that more p53 was required for the initial detection of transactivation. This may reflect differences in the ability of the mutant proteins to form tetramers. We conclude that there are many subtle p53 mutations whose effects may only be detected at low expression levels. Consistent with our previous reports, these p53 mutations can result in changes in the spectrum of REs transactivated. The quantitative assay is useful to address the impact of subtle differences in REs on transactivation as well as the effects of functional mutations on transactivation parameters. 1. PNAS 17 (2003) 9934.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]