Date of Award

Spring 2015

Project Type


Program or Major


Degree Name

Doctor of Philosophy

First Advisor

Kevin M Culligan

Second Advisor

Vaughn S Cooper

Third Advisor

Estelle M Hrabak


Challenging human health issues include treatments for genetic diseases and providing improved agricultural crop output to feed the growing world population. The project described here, which focuses on how cells respond to chromosomal (genomic) damage, has significant implications in each example. In humans, accumulation of DNA damage induced mutations can result in genetic diseases such as cancer, and in plants can similarly result in genome instability, reducing productivity. Organisms from human to plants have conserved mechanisms to counteract DNA damage. However, detailed genetic and biochemical information on plant DNA repair systems is still limited. The goal of this dissertation was to characterize the role of the Replication Protein A1 (RPA1) genes in DNA repair and related cellular processes such as DNA replication and meiotic recombination in Arabidopsis thaliana.

Besides being a good model system in plant molecular genetics research, A. thaliana provides a unique advantage for the study of DNA metabolism. Unlike most animal systems, loss-of-function mutations in known DNA damage response genes are generally not lethal in A. thaliana. This makes genetic characterization of its pathways more straight-forward and economical. In addition, since plants have similar DNA repair genes found from yeast to humans, it is possible that novel mechanisms and pathways discovered in plants will also be found in animals, potentially leading to insights of how mutations accumulate, and the resulting effects on disease and cancer progression in humans.

RPA is a heterotrimeric, single-stranded DNA (ssDNA) binding protein complex that is required for multiple processes in eukaryotic DNA metabolism, including replication, repair and recombination. RPA homologues have been identified in all eukaryotic organisms examined and are composed of subunits RPA1, RPA2 and RPA3, approximately 70, 34, and 14 kDa in size, respectively. Animals and yeast have only a single RPA1 subunit and use posttranslational modification to switch its function from DNA replication to repair. In contrast, plants typically encode multiple paralogs of RPA1 subunits and likely use different regulatory mechanisms.

A. thaliana encodes five RPA1 (RPA1A to RPA1E) subunits, and the studies presented in this dissertation demonstrate that these paralogs form three distinct functional groups composed of RPA1A (group A), RPA1B, RPA1D (group B) and RPA1C, RPA1E (group C) as determined by phylogenetic and functional genetic analysis.

DNA damage hypersensitivity, gene expression, fertility/meiotic analysis, and DNA replication assays suggest that the three groups play both unique and overlapping roles in DNA metabolism. While the RPA1C and RPA1A groups are primarily responsible for DNA repair and meiotic recombination, respectively, the RPA1B group promotes DNA replication. In addition, amino acid and nucleotide sequence analysis reveals that these groups have unique domains, motifs, cis-elements, gene expression profiles, and conservation that are consistent with the proposed functions.