Date of Award
Doctor of Philosophy (PhD)
Deoxyribonucleic acid (DNA) encodes genetic information essential for cell survival and function. However, it is constantly under assault from endogenous and exogenous damaging agents that not only threaten our own survival but also affect the faithful transmission of genetic information to our offspring. Double-strand breaks (DSBs) are one of the most hazardous forms of DNA damage, which if unrepaired or improperly repaired could lead to plethora of systemic human diseases including cancer. To deal with this problem, cells have evolved with a mechanism called DNA damage response (DDR) to detect, signal, and repair the breaks by inducing multiple cellular events. Resection is one of the key processes of cellular response to DSBs damage and is essential for genome maintenance, cell survival, and tumor suppression. Resection involves selective nucleolytic processing of the 5’ strand DNA at DSB ends to generate 3’ ssDNA overhangs, which in turn control both DNA repair and checkpoint response to the damage. Checkpoints coordinate the damage repair to other cellular processes including cell cycle regulation and gene expression. Despite its critical importance, the biochemical mechanisms and regulation of DSB resection is still not completely understood. Genetic studies in yeasts have suggested two steps mechanisms of resection: initiation by CtIP and MRN (Mre11-Rad50-NBS1) complex and extension by Dna2 and Exo1. We took a multipronged approach to study the resection process and have determined new mechanisms and regulation of both initiation and extension pathways. Here, we report a novel mechanism for the initiation of resection at clean DSBs mediated by Dna2 endonuclease activity. Our results strongly suggest that resection of blocked and free DSB ends is initiated via distinct mechanisms. In addition, we have demonstrated that the extension of resection by Exo1 is regulated both positively and negatively by Poly(ADP-ribosyl)ation, a prominent posttranslational modification at the sites of DNA damage. Our results suggest that Poly(ADP-ribose) not only promote initial damage recruitment of Exo1 but also prevent unscheduled and improper extension of resection. These two separate studies demonstrating new mechanisms for both initiation and extension steps of resection provide some critical new insights into the cellular response to DSBs damage.
Chair and Committee
Peter M. Burgers, Milan G. Chheda, Roberto Galletto, Nima Mosammaparast,
Paudyal, Sharad C., "Mechanisms and Regulation of Resection in DNA Damage Response" (2017). Arts & Sciences Electronic Theses and Dissertations. 1225.