Role of p53 in radiation induced genomic instability

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We have previouslyreported the induction of mutations at the maternal alleles ofESTR locus and the pink-eyed unstable locus of Fl mice born to irradiated spermatozoa (PNAS 98, 1705, 2001; Radiat. Res. 157, 661, 2002)). These observations indicated the presence of cross-talk between X-irradiated paternal genome andunirradiated maternal genome whichinduces untargeted recombination in zygotic stage embryos and delayed recombination in fetuses. Further study has indicated that thiscross-talk is dependent on the function of p53 which mediates a novel p53 dependent S checkpoint in the zygotic stage mouse embryos (MCB, 22, 2220, 2002).In this study, we have first analyzed the p53 protein domain required for the novel p53 dependent S checkpoint Microinjection analyses of p53 protein with specific mutation demonstrated that this S checkpoint neither required transcription activation domain nor the ATM phosphorylation sites. However, the protein had mutation in the DNA binding domain was unable … More to carry out the S checkpoint function. These suggest that the p53 protein itself may bind to DNA to execute the S checkpoint (Oncogene24, 3229, 2005). In parallel with these results, DNA fiber analyses revealed that p53 dependent S checkpoint functioned in suppressing the progression of replication fork when cells were challenged by X-irradiationOOncogene25, 529, 2006).It can be envisaged that the slowing down of the replication fork progression may facilitate recombination between sisterchromatids. In order to test this possibility, primary mouse embryofibroblasts (MEFs) with the wild type p53 gene and those with the null allele were exposed to X-rays of up to 6 Gy. The frequency of SCE increased dose dependently in the wild type MEFs while that of the p53 null MEFs did not This links the p53 dependent S checkpoint to homologous recombination at least between sister chromatids.Our previous study indicated the increase in the frequency of recombinational mutation at the maternal pink-eyed unstable allele in the retinal pigment epithelial cells of sperm irradiated embryos. Since the retinal pigment epithelial cells start development at day 11 to day 12 of gestation, the frequency of recombination stays elevated for at least 11 to 12 days after fertilization with irradiated sperm. Indeed, the frequency of SCE was higher in primary MEFs of day 12 fetus fertilized by irradiated sperm (in preparation. Altogether, our 3 year mouse study has excavated a novel p53 dependent S checkpoint and its function toupregulate homologous recombination for at least 11 to 12 days in sperm irradiated embryos. In addition, they also demonstrated the DNA damage memory and its function in delayed recombination.In order to study the molecular mechanism, we have also tested Schizosaccharomyces pombe to see if the delayed and untargeted recombination can be induced. S. pombe exhibited upregulated recombination for around 10 cell cycle generations afterX-irradiation. The length of upregulated recombination was cell generation dependent rather than the absolute time dependent as shown by the temperature shift experiments. In addition, this upregulated recombination was not due to bystander factors since the phenomena were not affected by the presence of radical scavengers in the culture media. Concomitantly with the upregulation of the recombination frequency, Rad22 foci were observed for around 10 generations after irradiation. These studies also demonstrated the DNA damage memory and delayed recombination in S. pombe Less