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Title: Investigating the role of ERCC6L2 in the DNA damage response
Author: Carnie, Christopher J.
ISNI:       0000 0004 7654 2041
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2018
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ERCC6L2 is a SNF2 ATPase found mutated in a small number of patients with severe bone marrow failure (BMF). Studies identifying ERCC6L2 mutations have suggested a role for this uncharacterised protein in the DNA damage response (DDR), but its function has remained unclear. Beyond a conserved catalytic core coupling ATP hydrolysis to a DNA translocase activity, ERCC6L2 contains a Tudor domain and a number of patches of strongly conserved residues, but no other characterised domains are annotated. This project aimed to ascertain whether or not ERCC6L2 is involved in specific arms of the DDR and to investigate the contributions of putative functional domains to such a role. This doctoral thesis confirms a role for ERCC6L2 in the response to DNA damage. Experiments using overexpressed ERCC6L2 indicate that ERCC6L2 is recruited to sites of DNA damage in a manner dependent on several different regions of the protein, none of which are characterised domains. Notably, a range of clinical ERCC6L2 mutants fail to localise to DNA damage sites, perhaps explaining a DDR defect in ERCC6L2-mutant patients. Clonogenic survival assays using ERCC6L2-/- cell lines are sensitive to DNA damage-inducing agents known to cause DNA double strand breaks (DSBs) and DNA replication perturbations. Notably, ERCC6L2's role in the response to these drugs is dependent on its ATPase activity, which is likely lost in several BMF-causing point mutations. Interestingly, ERCC6L2-/- cells also display a profound defect in DNA replication upon genotoxic treatment. Importantly, this defect is rescued by overexpression of wild-type ERCC6L2 but not by overexpression of a catalytic-dead mutant. Overall, this work demonstrates a role for ERCC6L2 in the DNA damage response, which is dependent on its catalytic activity, as well as identifying several important domains and features of ERCC6L2 which impact upon its function. Although the mechanistic basis of ERCC6L2 function in the context of DSB repair and/or DNA replication is still unclear, these findings might provide valuable insights into genome stability and the basis of ERCC6L2 deficiency disorders.
Supervisor: Ahel, Dragana Sponsor: EPA Research Fund
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available