DNA damage response pathways in DNA double strand break repair and hepatic metabolism
Moore, Elizabeth Susanne
The maintenance of genomic integrity and cellular homeostasis relies upon the appropriate selection and coordination of signaling and effector pathways in response to damage or perturbations. The DNA damage response (DDR) is an intricate network of surveillance, signal transduction, checkpoint activation, and repair protein recruitment and function. The DDR is capable of exquisite modulation to appropriately respond to a wide array of damage types, extents, and contexts that depend on cell type, cell cycle stage, cellular metabolic state, and extracellular cues. One of the most deleterious DNA lesions is a double stranded break (DSB). Multiple repair pathways have evolved to repair DSBs that differ in repair factors, mechanism, kinetics, and fidelity, and pathway choice is incompletely understood. Lower accuracy non-homologous end joining (NHEJ) and high fidelity homologous recombination (HR) are the two major DSB repair mechanisms. We investigated the role of the HUS1 component of the RAD9A-HUS1-RAD1 (9-1-1) DNA damage response clamp in DSB repair. Combined absence of the HR component Rad54 and Hus1 deficiency increased genome instability and genotoxin sensitivity, suggesting both Hus1 and Rad54 contribute to HR. In contrast, combined loss of the NHEJ component Prkdc and Hus1 deficiency resulted in no significant increase in genomic instability and a partial rescue in genotoxin sensitivity in vivo. This suggests that the 9-1-1 clamp may have a role in suppressing inappropriate NHEJ in certain contexts and/or that alternative repair pathways are disinhibited in the absence of NHEJ and deficiency of Hus1. The data suggest the 9-1-1 clamp plays a role in DSB repair pathway selection. Cell metabolism and genome integrity are intersecting and coordinated, and many canonical DDR proteins also regulate metabolism. This work describes a novel connection between a component of the Fanconi Anemia (FA) DNA repair pathway and hepatic metabolism. Altered energy states have been described in FA deficient cells, and human patients with a defect in the FA pathway are predisposed to metabolic disease through an unknown etiology. Upon challenge with a high fat, high cholesterol diet, FA-deficient Fancd2-/- male mice developed hepatic pathology accompanied by altered expression of cholesterol and bile acid metabolism genes and displayed numerous differential abundances of hepatic lipid species, in the absence of significant elevation in DNA damage or DDR activation. This indicates a role for the FA pathway in hepatic metabolic homeostasis. This work advances our understanding of the molecular function and dynamics of the 9-1-1 DDR repair complex and the mechanisms through which cells respond to DSBs and replication stress. Finessed understanding of the interplay between DDR factors can help us better identify derangements that result in deleterious phenotypes as well as identify DDR pathway components upon which cancer cells have increased reliance, so they may be targeted to increase therapeutic success. This work also advances the known roles of the Fanconi Anemia pathway beyond canonical DNA repair, which may lead to improved understanding of the etiology of FA phenotypes and therapeutic strategies. Elucidating the many connections between the DDR and cellular metabolic homeostasis has broad implications for understanding cancer cell biology.
Genetics; Molecular biology; Biology
Weiss, Robert S.
Schimenti, John C.; Wakshlag, Joseph J.; Cummings, Bethany P.
Biomedical and Biological Sciences
Ph.D., Biomedical and Biological Sciences
Doctor of Philosophy
dissertation or thesis