Date on Master's Thesis/Doctoral Dissertation

5-2007

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Pharmacology and Toxicology

Committee Chair

McGregor, W. Glenn

Author's Keywords

Carcinogen; DNA polymerases; REV1; Translesion; DNA adducts; Carcinogenesis; Polymerase eta; Polymerase iota

Subject

DNA polymerases; Cancer--Genetic aspects

Abstract

Recent advances in understanding the molecular mechanisms of mutagenesis indicate that most mutations are dependent on the activity of translesion synthesis DNA polymerases. The impact of reducing the level of these polymerases on mutagenesis and carcinogenesis in mouse models is poorly defined. Using knock out strategies we were able to remove polymerase eta (pol ç), and polymerase iota (pol é) and pol eta/pol iota from the mouse and lower REV1 in the mouse lung. This dissertation reports the changes in UV-induced carcinogenesis and mutagenesis that were observed. Mutagenesis data in pol iota deficient cells clearly indicate pol iota as a mutagenic TLS polymerase in UV lesion bypass. Pol iota removal effectively lowered the mutational frequency in both pol eta null and wild-type backgrounds. Unexpectedly, after Hprt mutant screening, pol eta and pol iota deficient cells were found to participate in UV lesion bypass in a strand-specific manner. This suggests that not only does bypass of photoproducts occur but occurs in an asymmetrical fashion, with preference of polymerases for leading or lagging strand. To examine the hypothesis that reducing the mutagenic load will reduce the incidence of cancer, we examined how pol iota status contributes to carcinogenesis. Despite the fact that pol iota was mutagenic in bypass of UV induced lesions, its removal accelerated cancer formation in the pol eta null background. The mechanisms behind this tumor suppressor function remain elusive, but indicate pol iota may have additional cellular roles in conjunction with its polymerase activity. Examining the hypothesis that reducing the mutagenic load will reduce the incidence of cancer, we developed strategies to reduce REV1 in mouse models of lung carcinogenesis. Endogenous REV1 mRNA in the lung was effectively lowered with the use of gene delivery of REV1 targeting ribozyme. This reduction was found to effectively decrease the multiplicity of B[a]P-induced lung tumors. This reduction did not affect the size or types of tumors induced suggesting inhibition of cancer formation occurred at the initiation step. Collectively, these data yield insight into the molecular mechanisms of mutagenesis that induce cancer formation.

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