Presentation Title

Examination of WT AND Mutant p53 on DNA Binding in vivo

Start Date

November 2016

End Date

November 2016

Location

HUB 302-60

Type of Presentation

Poster

Abstract

The tumor suppressor and transcription factor p53 is a regulatory protein that participates in the prevention of cancer and other cell processes such as apoptosis, DNA repair, and cell cycle inhibition. However, when mutated p53 is highly expressed in various types of cancer and can no longer perform its function. In this study, the six hotspot mutations in the DNA binding domain were examined for DNA binding in vivo. Wild type and mutant p53 were transfected into mammalian cells and Chromatin Immunoprecipitation Assay (ChIP) was performed to determine the ability of p53 to bind to mdm2 P2 promoter in vivo. In this assay, protein-DNA complexes were formed and immunoprecipitated using p53 specific antibodies. The purified protein-DNA complexes were then quantified by a real time PCR. Our results showed that some p53 mutants were able to bind to DNA (R248Q and R175C) while other mutants could not bind as well. It is possible that mutants are aggregating rather than binding to the DNA. To test this hypothesis, an OC antibody will be used to test for aggregation potential of the wild type and mutant p53.

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Nov 12th, 1:00 PM Nov 12th, 2:00 PM

Examination of WT AND Mutant p53 on DNA Binding in vivo

HUB 302-60

The tumor suppressor and transcription factor p53 is a regulatory protein that participates in the prevention of cancer and other cell processes such as apoptosis, DNA repair, and cell cycle inhibition. However, when mutated p53 is highly expressed in various types of cancer and can no longer perform its function. In this study, the six hotspot mutations in the DNA binding domain were examined for DNA binding in vivo. Wild type and mutant p53 were transfected into mammalian cells and Chromatin Immunoprecipitation Assay (ChIP) was performed to determine the ability of p53 to bind to mdm2 P2 promoter in vivo. In this assay, protein-DNA complexes were formed and immunoprecipitated using p53 specific antibodies. The purified protein-DNA complexes were then quantified by a real time PCR. Our results showed that some p53 mutants were able to bind to DNA (R248Q and R175C) while other mutants could not bind as well. It is possible that mutants are aggregating rather than binding to the DNA. To test this hypothesis, an OC antibody will be used to test for aggregation potential of the wild type and mutant p53.