Date of Award

Winter 2019

Project Type

Dissertation

Program or Major

Biochemistry

Degree Name

Doctor of Philosophy

First Advisor

Kevin Culligan

Second Advisor

Jessica Bolker

Third Advisor

mary Katherine Lockwood

Abstract

The DNA of every organism is under continuous attack from both environmental agents, errors in DNA replication, and reactive oxygen species produced from cellular metabolism. The preservation of genome stability and integrity is essential to counteract threats to genome integrity. Eukaryotic organisms have evolved a complex set of mechanisms to sense DNA damage and initiate repair of the damage or death of the cell, collectively called the DNA damage response (DDR).

One of the main causes of detrimental DNA damage is due to microbial pathogens, both in mammals and in plants. In humans Escherichia coli, Helicobacter pylori and Pseudomonas aeruginosa are Gram-negative bacteria that cause disease and are a leading risk factor in cancer. Little is known about the mechanism pathogens use to cause DNA damage and the consequent activation of repair pathways to combat the pathogen.

Arabidopsis thaliana is a good model system for the study of DNA repair pathways, since, unlike most animal systems, loss-of-function mutations in the DNA damage response pathway are non-lethal. The genome of Arabidopsis encodes many orthologs of mammalian repair proteins making it a good model organism for gene-gene interactions. A comparative analysis of genes implicated in DDR in both plants and animals, using gene ontology and bioinformatic tools, revealed that fundamental mechanisms underlying the maintenance of genome integrity, as well as the associated genes, are conserved between animals and plants. This research suggests that plants can be used as a first step screening tool of DNA repair targeted therapies and their role in cancer treatment (Nikitaki et al., 2017).

RPA is a heterotrimeric single-stranded binding protein complex that is required for eukaryotic cell replication, repair and recombination. RPA homologs have been identified in all eukaryotic organism and are ubiquitous in cells. Plants encode multiple paralogs of RPA1 with each performing different functions in DNA metabolism. Arabidopsis encodes five RPA1 subunits (RPA1A-RPAE). RPA1C and RPA1E function in DNA repair as determined by phylogenetic and genetic analysis.

This dissertation is an investigation into the role of plant Replication Protein A 1 (RPA1) genes in the DNA damage response of Arabidopsis in response to infection and pathogenesis of the microbial pathogen Pseudomonas syringae. The Arabidopsis-Pseudomonas syringae interaction is a well-studied pathosystem that has triggered research on the mechanisms underlying plant recognition of pathogens, defense responses and pathogen virulence.

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