Analysis of the impact of a p53 mutation in a homogeneous genetic background

Abstract

In more than 50% of cancers, p53, a tumor suppressor gene involved cell cycle arrest and apoptosis, has been seen to be heavily mutated making it an important gene to study. There are several studies on p53 and its role in cancer, but they ignore the impact of genetic background. Past studies have shown that genetic background can have a significant effect on the phenotypic consequences of cancer driver mutations, however, all these studies are carried out in a heterogeneous environment. The goal of my study was to utilize the CRISPR Cas 9 system to create a loss of function mutation in the p53 gene in a well characterized human cell line (HEYA8F8) and to evaluate the impact of this mutation on cell growth and apoptotic function in identical genetic backgrounds. The resulting mutation was a deletion in codons 33-36 of exon 4 which decreased the length of the protein from 393 to 389 amino acids. Using the cell lines with the specified deletion, growth rates over 96 hours were compared, which resulted in higher cell counts for the mutant in comparison to the wildtype. Assay for drug sensitivity using cisplatin, the standard of care for many cancers, showed that mutant cell lines had decreased apoptotic function (higher cell viability) in comparison to the wild type. The overall results demonstrated that mutations in p53 increase cell viability when treated with chemotherapy and an increase in cell proliferation. We believe that the cell lines with the loss of function mutations in p53 generated will provide an ideal experimental set up to study how the genetic background can evolve to enhance cancer in future studies.