Investigating the roles of Chl1 helicase in DNA damage tolerance pathways
Reyes, Teresa Anne Clarisse
Reyes, Teresa Anne Clarisse
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Publication Date
2023-01-09
Type
Thesis
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Abstract
Faithful DNA replication is fundamental to ensure proper genomic stability. Eukaryotic cells have evolved sophisticated networks to deal with DNA lesions encountered during chromosome replication. The DNA damage response covers several processes important for genome stability, including detection of DNA lesions, checkpoint activation, cell cycle control and tolerance/repair of the DNA damage. DNA damage tolerance (DDT) pathways involve mutagenesis and recombination-mediated DNA damage bypass mechanisms specialized in the tolerance of lesions encountered during DNA replication. Several studies provided evidence that replication in the presence of DNA damage involves transient formation of recombination-like structures consisting of sister chromatid junctions (SCJs) that resemble double Holiday Junctions (dHJs) and are resolved by the conserved RecQ helicase Sgs1/BLM together with Top3 and Rmi1. The mechanism by which dHJs are being initially formed remains less understood. Chl1 is a conserved 5’ to 3’ DNA helicase, whose orthologues in humans are mutated in the developmental disorder Warsaw breakage syndrome (DDX11) and Fanconi anemia (FANCJ), respectively, both of which are characterized at the cellular level by DNA homologous recombination repair defects. Because Chl1 and its homologs play critical roles in maintaining genome integrity, we addressed potential functions for Chl1 in DDT using budding yeast cells as a model system. In addition to knockout mutants, we used different alleles of Chl1 that limit its expression to either S or G2/M phase of the cell cycle, abrogate specifically its recruitment to the replication fork or its helicase activity. We identified that Chl1 helicase is required for cell survival in the presence of DNA alkylating agents and promotes replication associated recombination. Its action is not restricted to S phase and genetic conditions involving PCNA polyubiquitylation, suggesting a more general role for Chl1 in facilitating downstream steps of recombination. Interestingly, we obtained genetic evidences for Chl1 acting in compensation with Srs2 helicase.
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NUI Galway