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Transcriptional repression by p53 through direct binding to a novel DNA element
R A Johnson1, T A Ince, K W Scotto
1Program in Molecular Pharmacology and Therapeutics, Memorial Sloan-Kettering Cancer Center, New York, New York 1002, USA.
Abstract:
The tumor suppressor protein p53 has been well documented as a transcriptional activator involved in the regulation of a number of critical genes involved in the cell cycle, response to DNA damage, and apoptosis. Activation by p53 requires the interaction of the protein with a consensus binding site consisting of two half-sites, each comprising two copies of the sequence PuPuPuC(A/T) arranged head-to-head and separated by 0-13 base pairs. In addition to activation, p53 has been shown to be a potent repressor of transcription. However, the basis for p53-mediated repression is not well understood and has been proposed to occur indirectly through interactions with other promoter-bound transcription factors. In the present study, we show that p53 can repress transcription directly by binding to a novel head-to-tail (HT) site within the MDR1 promoter. A mutation that disrupted p53 binding to the MDR1 HT site blocked p53-mediated repression of the MDR1 promoter in transfection assays. Replacement of the HT site with a head-to-head (HH) site converted the activity of p53 from repression to activation, indicating that simple recruitment of p53 to the promoter is not sufficient for repression and that the orientation of the binding element determines the fate of p53-regulated promoters.
Insights
The tumor suppressor protein p53 directly represses transcription by binding to a novel head-to-tail site on the MDR1 promoter. Binding site orientation dictates whether p53 activates or represses gene expression.
Area of Science:
- Molecular Biology
- Genetics
- Cancer Research
Background:
- The tumor suppressor protein p53 is a key regulator of cell cycle, DNA damage response, and apoptosis.
- p53 typically functions as a transcriptional activator, binding to specific head-to-head sites.
- The mechanisms underlying p53-mediated transcriptional repression are not fully understood.
Purpose of the Study:
- To investigate the direct mechanisms of p53-mediated transcriptional repression.
- To identify novel p53 binding sites involved in repression.
- To determine the role of binding site orientation in p53 function.
Main Methods:
- Analysis of p53 binding to the MDR1 promoter.
- Site-directed mutagenesis of p53 binding sites.
- Transfection assays to assess transcriptional activity.
Main Results:
- p53 directly represses transcription via a novel head-to-tail (HT) binding site in the MDR1 promoter.
- Disruption of p53 binding to the HT site abolished repression.
- Conversion of the HT site to a head-to-head (HH) site switched p53 function from repression to activation.
Conclusions:
- p53 can directly repress transcription through specific HT binding sites.
- The orientation of the p53 binding element is critical in determining its transcriptional outcome (activation vs. repression).
- These findings provide new insights into the complex regulatory roles of p53 in gene expression.