The understanding of basic DNA repair mechanisms is crucial, as altered regulation of these systems are observed in cancerous cells and also allow for antibiotic resistance in prokaryotes. In E. coli, two major mechanisms of transcription-coupled nucleotide excision repair (TCR) are responsible for removing bulky lesions in DNA that transcribing RNA polymerase encounters- the Mfd-mediated ?forward translocation? model and the UvrD-mediated ?backtracking? model. In this project I will use a genome-wide assay called XR-Seq to further understand E. coli TCR across the genome by determining locations of the genome repaired by each pathway. Further, I will perform NET-Seq in -/+ UV conditions to correlate transcription of both the sense and antisense nascent transcripts to TCR. Together, these studies will provide important information concerning the two known TCR pathways in E. coli by helping us understand how they may work together or separately across the genome and how their absence impacts genome integrity. Ultimately, this work will contribute to further defining the mechanisms of important repair processes that occur in prokaryotes and provide a guideline for future genome-wide studies of other important DNA repair processes.
Altered or dis-regulation of DNA repair systems leads to the tolerance of mutations that can promote the growth of cancer cells or allow bacteria to develop resistance to stresses such as antibiotics. This project will further our knowledge of DNA repair mechanism, specifically mechanism coupled to transcription that exist across all domains of life. Therefore, the detailed datasets and analysis produced for my project will help design work to understand repair in higher organisms.
