The Roles of chromatin modifying enzymes in DNA double-strand break repair
- Kelly Elizabeth McCann.
- June 2010.
- Physical description
- online resource (xviii, 242 pages) : illustrations (some color)
- McCann, K. E. (Kelly Elizabeth).
- Brown, J. M. (J. Martin) thesis advisor (primary).
- Cimprich, Karlene. thesis advisor.
- Ford, James M. (James Matthew). thesis advisor.
- Wysocka, Joanna, Ph. D. thesis advisor.
- Stanford University. Program in Cancer Biology. degree grantor.
- Stanford University. Committee on Graduate Studies.
- Includes bibliographical references (p. 227-242).
- Efficient DNA double-strand break (DSB) repair is essential for maintaining the stability of the genome. A single long-lived DSB can cause cell lethality (Bennett et al., 1996). Humans with defects in DSB repair are sensitive to DNA damaging agents and are predisposed to developing cancers, as exemplified by such diseases as Nijmegan breakage syndrome, Ataxia telangiectasia, and breast cancer susceptibility in women with mutations in the BRCA1 and BRCA2 genes (Duker, 2002). Repair of DNA damage requires activation of cell cycle checkpoint controls, recruitment of repair proteins to DNA lesions, and transcriptional activation of relevant genes. As shown by our data, deletion of particular histone modification genes produces sensitivity to ionizing radiation in Saccharomyces cerevisiae, suggesting a role for chromatin modification enzymes in the repair process as well. Because damage recognition and repair of lesions are both influenced by chromatin structure, we began by studying the role of histone acetylation in the DNA damage response. Acetylation of the N-terminal tails of histone H4 opens the chromatin to allow repair enzymes to access broken DNA. Through a screen of the yeast deletion pool, we found that deletion of BRE1 and DOT1 genes causes sensitivity to ionizing radiation. Ubiquitination of H2B on lysine 123 (H2B-K123) by ubiquitin ligase Bre1 is necessary for methylation of H3 on lysine 79 (H3-K79) by Dot1. Through the histone modifications they catalyze, these proteins are involved in many aspects of the DNA repair response, as outlined in Sections C, D, and E. Our studies focused on homologous recombination repair defects, genome-wide expression patterns in BRE1 and DOT1 deletion mutants, an analysis of the data regarding proteins purported to bind to methylated H3-K79, and optimization of a protein purification strategy to find Dot1 binding partners. Our yeast deletion pool screen also predicted a role for N-terminal acetyltransferase complex NatB in DNA double-strand break repair. In Section F, we build a case for acetylation of DNA end-binding protein Mre11.
- Publication date
- Submitted to the Department of Cancer Biology and the Committee on Graduate Studies of Stanford University.
- Thesis (Ph.D.)--Stanford University, 2010.