UBC Theses and Dissertations
Roles of the DOG-1 and JRH-1 helicase-like proteins in DNA repair in Caenorhabditis elegans Youds, Jillian L.
Helicases perform vital roles in the cell by unwinding D N A to make it accessible for the essential processes of replication, transcription and repair. In Caenorhabditis elegans, the DOG- 1 helicase-like protein is required for polyG/polyC-tract (G/C-tract) maintenance, as dog-l animals have a mutator phenotype characterized by deletions that initiate in G/C-tracts. DOG-1 may unwind secondary structures that form in polyguanine D N A during lagging strand replication. In order to more completely understand the role of dog-1, genetic interactors were identified, dog-1 functionally interacts with the him-6/BLM helicase. Absence of recombinational repair-implicated proteins in the dog-1 background, including HIM-6/BLM, RAD-51, BRD-1/BARD1 and HIM-9/XPF, as well as the trans-lesion synthesis polymerases polKMD po/7 increased the frequency of animals with G/C-tract deletions, indicating that these pathways are important mechanisms for repair at G/C-tracts in the absence of DOG-1. These data support the hypothesis that persisting D N A secondary structures can cause replication fork stalling, which can be resolved by deletion-free or deletion-prone mechanisms. DOG-1 has highest sequence identity to human BR1P1/FANCJ, which is mutated in patients from the Fanconi Anemia (FA) subgroup J. D N A damage sensitivity experiments indicated that, like chicken F A N C J cells, dog-1 mutants were not significantly sensitive to DNA damage from X-ray or UV-irradiation, but were extremely hypersensitive to the D N A interstrand cross-linking agent UVA-activated trimethylpsoralen. Thus, DOG-1 appears to have a conserved role in cross-link repair and is the C. elegans F A N C J homolog. Characterization of the dog-1/FANCJ-relatsd helicase, Jrh-1, revealed that mutants for this putative helicase are moderately sensitive to cross-linking agents, dog-1 jrh-1 double mutants displayed a synthetic lethal phenotype characterized by excessive recombination intermediates and mitotic catastrophe in the germline. However, absence of JRH-1 did not have any effect on G/C-tract deletions, indicating that JRH-1 does not have a redundant function with DOG-1 at G/C-tracts. Absence of JRH-1 reduced the fitness of eTl and nTl translocation hétérozygotes, but not translocation homozygotes, jrh-1 was synthetically lethal with him-6/BLM and with the endonuclease mus-81, suggesting a possible role for JRH-1 in regulating the balance between different types of repair.
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