Thesis
Investigating the role of Timeless in acetaldehyde-mediated DNA damage response and repair
Master of Science (M.S.), Drexel University
Jun 2017
DOI:
https://doi.org/10.17918/etd-7531
Abstract
Esophageal squamous cell carcinoma (ESCC) is highly associated with chronic alcohol consumption. Acetaldehyde, the intermediate metabolite in alcohol metabolism, is considered to be a carcinogenic molecule involved in ESCC development. Adducts, DNA-protein crosslinks, and DNA interstrand crosslinks are generated via acetaldehyde interacting with deoxyguanosine residues of DNA. These lesions physically impede the replication machinery, resulting in stalled replication forks. Timeless, a subunit of the fork protection complex, functions at the DNA replication fork to provide stability and prevent fork collapse, which can occur due to prolonged fork stalling. Timeless is also responsible for initiating cell cycle checkpoint activation in response to replication stresses, such as dNTP deprivation by hydroxyurea. Recently, Timeless has been implicated in DNA double-strand break repair by promoting homologous recombination. Yet, other studies have shown an increased frequency of sister chromatid recombination in cells depleted of Timeless. Whether Timeless promotes cell cycle arrest and DNA damage recovery in response to acetaldehyde-mediated DNA damage is largely unknown. Here, we report that Timeless depletion reduces proliferation of cells that are exposed to acetaldehyde. Chk1 and p53 protein levels are reduced in cells depleted of Timeless, resulting in a loss of ability to efficiently arrest the cell cycle in response to acetaldehyde. Ubiquitinated FancD2 persists in response to increasing concentrations of acetaldehyde, suggestive of Fanconi Anemia pathway activation and ICL repair. In addition, there is a significant increase in 53BP1 and BRCA1 foci formation in cells with acetaldehyde-induced DNA damage in the absence of Timeless. Finally, Timeless depletion resulted in an increase in homologous recombination capacity, while the rate of non-homologous end joining is reduced, suggesting a role of Timeless as an anti-recombinase. Taken together, our data show that Timeless functions to provide cellular tolerance to acetaldehyde-mediated DNA damage, as well as to promote DNA damage recovery, specifically through non-homologous end joining.
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Details
- Title
- Investigating the role of Timeless in acetaldehyde-mediated DNA damage response and repair
- Creators
- Amber Theriault - DU
- Contributors
- Eishi Noguchi (Advisor) - Drexel University (1970-)
- Awarding Institution
- Drexel University
- Degree Awarded
- Master of Science (M.S.)
- Publisher
- Drexel University; Philadelphia, Pennsylvania
- Number of pages
- ix, 70 pages
- Resource Type
- Thesis
- Language
- English
- Academic Unit
- Biochemistry and Molecular Biology; College of Medicine; Drexel University
- Other Identifier
- 7531; 991014632696104721