The role of p53 in DNA damage-mediated cytotoxicity overrides its ability to regulate nucleotide excision repair in human fibroblasts

The p53 tumour suppressor protein plays a pivotal role in the response of mammalian cells to DNA damage. In addition to its regulatory role in cell cycle progression, p53 regulates apoptosis and can therefore influence cellular survival in response to DNA damage. More recent work has revealed that p53 is also involved in the nucleotide excision repair (NER) of structurally diverse types of DNA damage. The relative influence of p53 on NER and cellular sensitivity to DNA damage was investigated in this study using cells that differ in p53 status. Two cell models were selected: 041 TR fibroblasts in which the expression of p53 is regulated by a tetracycline-inducible promoter, and WI38 primary lung fibroblasts together with their isogenic derivative VA13, in which p53 is abrogated post-translationally by SV40 transformation. Cells were exposed to the clinically and environmentally relevant DNA-damaging agents cisplatin (0-5 microM, 2 h), (+/-)-anti-benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide (0-0.5 microM, 30 min) and UV-C (0-5 J/m2), each of which induce structurally distinct types of DNA damage known to be subject to p53-dependent NER. Sensitivity of the p53-proficient and p53-deficient cells to this DNA damage was then compared at each dose of DNA-damaging agent using the clonogenic survival assay and the colorimetric MTT assay. p53-proficient cells were more sensitive than p53-deficient cells to cisplatin, (+/-)-anti-benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide and UV-C; these differences in cellular sensitivity were more apparent in the 041 TR cells (up to 3.6-, 5.8- and 1.9-fold, respectively) than the WI38/VA13 cells (up to 2.3-, 1.4- and 1.4-fold, respectively). Thus, despite the well-documented persistence of DNA damage in p53-deficient fibroblasts due to impaired NER, loss of p53 results in reduced DNA damage-mediated cytotoxicity.