P53 Interacts with the DNA Mismatch Repair System to Modulate the Cytotoxicity and Mutagenicity of Hydrogen Peroxide

This study focused on the question of how the DNA mismatch repair (MMR) system and p53 interact to maintain genomic integrity in the presence of the mutagenic stress induced by hydrogen peroxide (H(2)O(2)). The cytotoxic and mutagenic effects of H(2)O(2) were compared in four colon carcinoma sublines: HCT116, HCT116/E6, HCT116+ch3, and HCT116+ch3/E6, representing MMR(-)/p53(+), MMR(-)/p53(-), MMR(+)/p53(+), and MMR(+)/p53(-) phenotypes, respectively. Loss of p53 in MMR-proficient cells did not significantly alter cellular sensitivity to H(2)O(2), but disruption of p53 in MMR-deficient cells resulted in substantial resistance to H(2)O(2) (IC(50) values of 203.8 and 66.2 microM for MMR(-)/p53(-) and MMR(-)/p53(+) cells, respectively). The effect of loss of p53 and MMR function on sensitivity to the mutagenic effect of H(2)O(2) paralleled the effects on cytotoxic sensitivity. In MMR-deficient cells, loss of p53 resulted in a 3.5- and 2.2-fold increase in the generation of 6-thiogunaine and ouabain-resistant clones, respectively. Loss of MMR in combination with loss of p53 synergistically increased the frequency of frameshift mutations in the CA repeat tracts of the out-of-frame shuttle vector pZCA29 and further promoted instability of microsatellite sequences under H(2)O(2) stress. Flow cytometric analysis showed that H(2)O(2) treatment produced a G(l) and G(2)/M phase arrest in MMR(+)/p53(+) cells. Loss of MMR did not alter the ability of H(2)O(2) to activate either checkpoint; loss of p53 in either the MMR-proficient or deficient cells resulted in impairment of the G(l) arrest and a more pronounced G(2)/M arrest. H(2)O(2) caused a greater and more longed increase in p53 protein levels in MMR-proficient than in the MMR-deficient cells. The results demonstrate that the effect of disabling p53 function is modulated by the proficiency of the MMR system (and vice versa) and that there is an overlap between the functions of p53 and the MMR system with respect to the activation of apoptosis and mutagenesis after an oxidative stress.