Date of Completion

4-9-2015

Embargo Period

10-6-2015

Keywords

DNA damage, Mutagenesis, Translesion Synthesis, Abasic Site, 8, 5’-Cyclopurines, O2-Alkylthymidine, DNA Lesions

Major Advisor

Dr. Ashis Basu

Associate Advisor

Dr. Steven Suib

Associate Advisor

Dr. Mark Peczuh

Field of Study

Chemistry

Degree

Doctor of Philosophy

Open Access

Open Access

Abstract

DNA contains the genetic information of all living organisms. Therefore, its integrity and stability are essential for the long-term viability of the organism. However, DNA is a chemical entity, which gets constantly damaged by both endogenous and exogenous DNA damaging agents. These DNA damages may lead to mutations and eventually cause diseases like cancer. Multiple DNA repair mechanisms have evolved in living organisms to repair DNA damages. Even so, not all DNA damages can be repaired before the replication apparatus encounters the DNA damage. As such, cells developed a damage tolerance pathway known as translesion synthesis (TLS) that allows cells to overcome replication blockage and facilitate bypass of the DNA lesions. This process is carried out by TLS polymerases, of which most belong to the Y family of DNA polymerases. These specialized enzymes are capable of bypassing the damaged DNA, but they also are low fidelity enzymes, frequently associated with mutagenesis and carcinogenesis.

In this dissertation, I have investigated the replication bypass of different DNA lesions, including abasic site, 8,5’-cyclopurines and tobacco-specific nitrosamine-derived O2-alkylthimidines in Escherichia coli and in human embryonic kidney (HEK293T) cells. The comparative replicative assays in bacterial and human cells revealed that bypass of these DNA lesions are significantly different in the two systems and that the mammalian polymerases are more efficient in the TLS of strong replication blocking lesions. Based on the cytotoxic and mutagenic properties of these lesions, it is evident that replicative bypass efficiency varies with the complexity of the lesion. Moreover, multiple TLS polymerases are involved in the mutagenesis of different DNA lesions. These studies provide important mechanistic details as to how these lesions are bypassed and reveal their mutagenic properties.

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