p53 activates transcription by binding as a tetramer to a consensus sequence (the p53 HH site) within the promoter of target genes. Although transcriptional repression by p53 may be equally important, less is known about the mechanism by which p53 recognizes promoters of repressed genes. In the genes studied to date, the lack of a consensus p53 binding site within the repressed promoter has led investigators to propose that p53 works indirectly, either by inhibiting a bound activator or by repressing the basal machinery. We have re-addressed this question with respect to the MDR1 promoter, beginning with the novel hypothesis that the "switch" of p53 from an "activator" to a "repressor" may involve interaction with a non-consensus DNA binding site; in fact, binding itself may require or induce an altered conformation of the p53 tetramer, resulting in the unmasking of repressor domains. Based on this hypothesis, we have identified a novel arrangement of p53 half-sites within the MDR1 promoter which are capable of interacting with p53 as efficiently as the consensus HH site. Importantly, mutation of this site (the HT site) abrogates both p53 binding and p53- mediated repression of the MDR1 promoter. Interestingly, the p53 family members are not able to repress through the HT site and, in fact activate MDR1 transcription though an independent region. We propose to continue to investigate p53-mediated repression through the HT site in the following Aims: 1) We will analyze the interaction of the HT-p53 complex versus the HH-p53 complex with transcriptional co-factors, both in vitro and in vivo: 2) We will determine the domains of p53 involved in repression through the HT element and evaluate the effect of post-translational modifications on modulating this repression; 3) We will continue to characterize the HT element, both in the MDR1 promoter and in other p53-repressed promoters in which we have identified HT sites; and 4) We will pursue our novel observation of a diverse effect of p53 and its family members on MDR1 transcription by analyzing p73/p53 chimeric proteins.
|Effective start/end date||9/1/01 → 8/31/06|
- National Cancer Institute: $231,300.00
- National Cancer Institute: $244,913.00
- National Cancer Institute: $226,800.00
- National Cancer Institute: $264,600.00
- Cancer Research
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