title: Annealing of Complementary DNA Sequences During Double-Strand Break Repair inDrosophilaIs Mediated by the Ortholog of SMARCAL1.
abstract: DNA double-strand breaks (DSBs) pose a serious threat to genomic integrity. If unrepaired, they can lead to chromosome fragmentation and cell death. If repaired incorrectly, they can cause mutations and chromosome rearrangements. DSBs are repaired using end-joining or homology-directed repair strategies, with the predominant form of homology-directed repair being synthesis-dependent strand annealing (SDSA). SDSA is the first defense against genomic rearrangements and information loss during DSB repair, making it a vital component of cell health and an attractive target for chemotherapeutic development. SDSA has also been proposed to be the primary mechanism for integration of large insertions during genome editing with CRISPR/Cas9. Despite the central role for SDSA in genome stability, little is known about the defining step: annealing. We hypothesized that annealing during SDSA is performed by the annealing helicase SMARCAL1, which can anneal RPA-coated single DNA strands during replication-associated DNA damage repair. We used unique genetic tools inDrosophila melanogasterto test whether the fly ortholog of SMARCAL1, Marcal1, mediates annealing during SDSA. Repair that requires annealing is significantly reduced inMarcal1null mutants in both synthesis-dependent and synthesis-independent (single-strand annealing) assays. Elimination of the ATP-binding activity of Marcal1 also reduced annealing-dependent repair, suggesting that the annealing activity requires translocation along DNA. Unlike the null mutant, however, the ATP-binding defect mutant showed reduced end joining, shedding light on the interaction between SDSA and end-joining pathways.
authors.affiliation: Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, North Carolina 27599.
authors.forename: Julie Korda
authors.initials: JK
authors.lastname: Holsclaw
authors.affiliation: Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, North Carolina 27599 sekelsky@unc.edu.|Department of Biology, University of North Carolina, Chapel Hill, North Carolina 27599.|Integrative Program in Biological and Genome Sciences, University of North Carolina, Chapel Hill, North Carolina 27599.
authors.forename: Jeff
authors.identifier: 0000-0002-4424-677X
authors.initials: J
authors.lastname: Sekelsky
mesh_terms: D000255:Adenosine Triphosphate / Q000235:genetics; D000818:Animals; D064113:CRISPR-Cas Systems; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage / Q000235:genetics; D059766:DNA End-Joining Repair / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics; D000072669:Gene Editing; D042822:Genomic Instability / Q000235:genetics*; D000071796:SMARCB1 Protein / Q000235:genetics*
chemical_list: D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D000071796:SMARCB1 Protein; D000255:Adenosine Triphosphate
keywords: DSB repair; Drosophila; SDSA; SMARCAL1; homologous recombination