title: A Delicate Balance Between Repair and Replication Factors Regulates Recombination Between Divergent DNA Sequences in Saccharomyces cerevisiae.
abstract: Single-strand annealing (SSA) is an important homologous recombination mechanism that repairs DNA double strand breaks (DSBs) occurring between closely spaced repeat sequences. During SSA, the DSB is acted upon by exonucleases to reveal complementary sequences that anneal and are then repaired through tail clipping, DNA synthesis, and ligation steps. In baker's yeast, the Msh DNA mismatch recognition complex and the Sgs1 helicase act to suppress SSA between divergent sequences by binding to mismatches present in heteroduplex DNA intermediates and triggering a DNA unwinding mechanism known as heteroduplex rejection. Using baker's yeast as a model, we have identified new factors and regulatory steps in heteroduplex rejection during SSA. First we showed that Top3-Rmi1, a topoisomerase complex that interacts with Sgs1, is required for heteroduplex rejection. Second, we found that the replication processivity clamp proliferating cell nuclear antigen (PCNA) is dispensable for heteroduplex rejection, but is important for repairing mismatches formed during SSA. Third, we showed that modest overexpression of Msh6 results in a significant increase in heteroduplex rejection; this increase is due to a compromise in Msh2-Msh3 function required for the clipping of 3' tails. Thus 3' tail clipping during SSA is a critical regulatory step in the repair vs. rejection decision; rejection is favored before the 3' tails are clipped. Unexpectedly, Msh6 overexpression, through interactions with PCNA, disrupted heteroduplex rejection between divergent sequences in another recombination substrate. These observations illustrate the delicate balance that exists between repair and replication factors to optimize genome stability.
authors.affiliation: Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853.
authors.forename: Ujani
authors.initials: U
authors.lastname: Chakraborty
authors.affiliation: Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853.
authors.forename: Carolyn M
authors.initials: CM
authors.lastname: George
authors.affiliation: Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853.
authors.forename: Amy M
authors.initials: AM
authors.lastname: Lyndaker
authors.affiliation: Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853 eea3@cornell.edu.
authors.forename: Eric
authors.initials: E
authors.lastname: Alani
mesh_terms: D053843:DNA Mismatch Repair; D004260:DNA Repair; D004261:DNA Replication; D004271:DNA, Fungal; D004268:DNA-Binding Proteins / Q000378:metabolism; D015870:Gene Expression; D005838:Genotype; D059765:Homologous Recombination; D008954:Models, Biological; D009154:Mutation; D011485:Protein Binding; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*
chemical_list: D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins
keywords: DNA mismatch repair; Msh6 overexpression; PCNA; Rmi1; Sgs1; Top3; heteroduplex rejection