The role of Msh2 DNA mismatch pair and P27(kip1) cell cycle regulation on mutagenesis and carcinogenesis




Zhang, Shulin

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Transgenic rodents harbouring the E. coli lacI gene greatly facilitate the study of mutations in vivo where the effects of age, diet, lifestyle, sex, tissue and species specificity can be assessed. In addition, it also permits the investigation of mutations in a specifically altered genetic background. In this thesis, I used the lacI transgenic rodents to study the effect of strains and species difference on spontaneous mutation in the liver, as well as the influence of the DNA repair gene Msh2 and the cell cycle regulation gene p27 on mutagenesis and carcinogenesis. By studying spontaneous mutations in different strains and species of rodents which has different transgene insertion sites and constructs, we demonstrate that despite such differences, the spontaneous mutation frequency and spectra are similar. The major parts of the thesis demonstrate the impact of a deficiency in the Msh2 and p27 gene on spontaneous and chemically induced mutations. The mutator phenotype of thymic lymphoma arising in an Msh2 deficient background was also studied. A deficiency in the Msh2 gene caused an significant increase in mutation frequency in three parts of the colon with a distinct mutational spectrum characterized by an increase of G:C>A:T transitions. However, we did not detect the differences in mutation frequency and spectrum among the three parts of the colon. The mutagenesis of a colonic mutagen and carcinogen 2-amino-1 -methyl-6-phenolimidazo[4,5-b]pyridine (PhIP) was investigated. Msh2 deficiency was found to increase PhIP induced colon mutagenesis in a synergistic manner. Msh2+/- mice displayed a significantly increased frequency of -1 frameshifts in the spontaneous and PhIP treatment group indicating that Msh2 germ line mutation carriers are also at an increased risk of developing cancers. Msh2 thymic lymphomas exhibit a large increase in mutation frequency and an altered mutational spectrum featured by an increase of base substitutions occurring at A:T basepairs, -1 frameshifts and complex mutations. The influence of a deficiency in the p27 cell cycle control gene on mutagenesis is addressed in the next section of the thesis. We created a novel double transgenic mouse strain bearing a different functional status of p27 gene as well as the lacI transgene. P27 deficient mice exhibit similar levels of spontaneous mutation and a similar mutational spectrum as p27 wild type and heterozygous mice. However, after N-nitroso-N-ethylurea (ENU) treatment, hypermutability was detected in p27-/- mice. Interestingly, p27 heterozygous mice displayed an intermediate sensitivity upon ENU treatment indicating an haplo-insufficiency of the p27 gene in protecting against chemically induced mutagenesis. All three genotypes of p27 mice displayed a similar mutational specificity after ENU treatment characterized by the mutations occurring at A:T base pairs. These results show that both Msh2 and p27 homozygous deficient mice are more susceptible to chemically induced mutation than wild type mice. In contrast to the finding of Msh2 mice, p27 functional status does not affect the mutational spectrum recovered in lacI transgene. This illustrates the different mechanisms of DNA mismatch repair and cell cycle regulation in maintaining genomic integrity. The haplo-insufficiency of some genes in safeguarding genomic stability highlights the importance of tumor screening in carrier populations.



Mutagenesis, Cancer, Genetic aspects