Thesis (M. Sc.)--York University, 2003. Graduate Programme in Biology. / Typescript. Includes bibliographical references (leaves 81-88). Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://wwwlib.umi.com/cr/yorku/fullcit?pMQ82948.

Thesis (Ph. D.)--University of Hong Kong, 1998. / Includes bibliographical references (leaf 151-168).

Thesis (M. Phil.)--University of Hong Kong, 1994. / Includes bibliographical references (leaves 137-156).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1970. / Vita. Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Thesis (Ph.D.)--Virginia Commonwealth University, 2008. / Prepared for: Dept. of Microbiology & Immunology. Title from thesis submission form. Includes bibliographical references.

Thesis (Ph. D.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed Oct. 16, 2007). Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Texas at Austin, 2007. / Vita. Includes bibliographical references.

The initiation of the cancer process is associated with mutations. Analysis of environmental exposure to chemical or physical agents causing these genetic alterations is of great importance in order to develop strategies for avoiding or reducing cancer risk in humans. The causality between mutagenesis and carcinogenesis also prompts the concept that the modifying effect on mutagenesis by a compound would be predictive of the cancer preventive potential of that compound. The Big Blue© transgenic assay, using the E. coli lacI gene as the mutational target provides an opportunity to evaluate mutagenesis and its modulation m vivo. This model system was used to study the tissue-specific effect of the potential chemopreventive agent conjugated linoleic acid (CLA), on the mutagenicity of the suspected human carcinogen, 2-amino-1 -methyl-6- phenylimidazo[4,5-b]pyridine (PhIP). PhIP and CLA were selected for study since both compounds are consumed by humans on a daily basis, and are suspected to be related to the human risk of colon, breast, and prostate cancers. The mutagenicity of PhIP in Big Blue© rats was shown to be tissue-, sex-, and dose-dependent. PhIP was found to be a potent mutagen in the colon, followed by the cecum, prostate, and kidney. Compared with the background mutational spectra, the PhlP-induced spectra were characterized by an elevated proportion o f-1 frameshifts, consisting mainly of deletions of single G:C base pair. However, the induced spectra varied among tissues. A sex-dependent induction of mutation by PhIP was observed in the kidney such that the PhlP-induced mutation frequency was twice as high in male rats as in female rats; the biological significance of this difference is not clear. In contrast, although PhIP has been shown to induce colon tumors preferentially in male rats, and only rarely in female rats, no difference in mutational response was detected between the colons of male and female rats treated with PhIP. Experiments were performed to examine the in vivo effect of CLA on mutagenesis. Similar to what is seen for the mutagenicity of PhIP, the modification by CLA depends on tissue, sex, and dose of administration. CLA showed a modest protection against PhlP-induced mutagenesis in the distal part of the colon, in the prostate, and in the kidney of female rats. However, significant changes in the overall PhlP-induced mutation spectrum were seen only in the prostate. The antimutagenic effect of CLA may be directly responsible for its cancer prevention «^lability, since PhlP-induced aberrant crypt foci in the colon of male rats were completely inhibited by CLA However, CLA was not totally innocuous. When supplemented at 0.5%, CLA acted as a comutagen of PhIP, increasing the PhlP-induced MF in the cecum, although this effect was not observed when CLA was supplemented at 1%. The differences in effect may be related to the antioxidant or pro-oxidant activities of CLA isomers under experimental conditions. Due to the artificial nature of the lambda/LIZ lacI transgene and the possible absence of DNA repair in this transgene, the suitability of the Big Blue© transgenic assay as a mutational test system has been questioned. We examined the repair of UV- and benzo(α)pyrene diol epoxide-induced DNA damage in this non-transcribed lambda construct of the Big Blue© rat-2 transgenic cell line and demonstrated that DNA damage is indeed repaired in this transgenic construct. Lastly, since CLA altered the mutational spectra in the prostate in a way consistent with an effect of mismatch repair, the possibility of an effect of CLA on mismatch repair was explored in bacteria. Although CLA was found to increase mutant frequency in a mismatch repair proficient E. coli strain, but not in deficient strains, the mechanism by which CLA operates remains unclear. Altogether, the data demonstrate the mutagenicity of PhIP and its modulation by CLA as a function of tissue, sex, and dose of administration, and support the application of the Big Blue© transgenic assay as a screening tool for mutagens and chemopreventive agent / Graduate

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. / Graduate

Recovery of DNA synthesis in UV-irradiated E. coli. E. coli undergoing the SOS response to DNA damage (e.g. UV-irradiation) displays transient inhibition of DNA synthesis. The mechanism of the inhibition and of the recovery of DNA synthesis following UV-irradition was analysed in several mutants defective in repair or in the regulation of the RecA-LexA dependent SOS response. The results indicated that inhibition is not an inducible function and is probably due to the direct effect of lesions in the template blocking replisome movement. In contrast, recovery of DNA synthesis after UV required protein synthesis and was clearly shown to be an SOS function under RecA control. In addition, analyses involving a recA200 (temperature sensitive RecA) and recA constitutive mutants demonstrated that RecA protein was directly required for recovery in addition to its regulatory role. These experiments however also suggested that an inducible Irr (inducible replisome reactivation) factor under RecA control was also required for post UV-recovery. In vitro mutagenesis. In this part of my study attempts were also made to establish an vitro system for untargeted mutagenesis based upon a plasmid replication system. This involved establishing optimum conditions for extracting plasmid DNA (replicated vitro) and a highly efficient transformation system, in order to analyse directly mutagenic events.