Tolerance of DNA Mismatches in Dmc1 Recombinase-mediated DNA Strand Exchange

Abstract

Recombination between homologous chromosomes is required for the faithful meiotic segregation of chromosomes and leads to the generation of genetic diversity. The conserved meiosis-specific Dmc1 recombinase catalyzes homologous recombination triggered by DNA double strand breaks through the exchange of parental DNA sequences. Although providing an efficient rate of DNA strand exchange between polymorphic alleles, Dmc1 must also guard against recombination between divergent sequences. How DNA mismatches affect Dmc1-mediated DNA strand exchange is not understood. We have used fluorescence resonance energy transfer to study the mechanism of Dmc1-mediated strand exchange between DNA oligonucleotides with different degrees of heterology. The efficiency of strand exchange is highly sensitive to the location, type, and distribution of mismatches. Mismatches near the 3 end of the initiating DNA strand have a small effect, whereas most mismatches near the 5 end impede strand exchange dramatically. The Hop2-Mnd1 protein complex stimulates Dmc1-catalyzed strand exchange on homologous DNA or containing a single mismatch. We observed that Dmc1 can reject divergent DNA sequences while bypassing a few mismatches in the DNA sequence. Our findings have important implications in understanding meiotic recombination. First, Dmc1 acts as an initial barrier for heterologous recombination, with the mismatch repair system providing a second level of proofreading, to ensure that ectopic sequences are not recombined. Second, Dmc1 stepping over infrequent mismatches is likely critical for allowing recombination between the polymorphic sequences of homologous chromosomes, thus contributing to gene conversion and genetic diversity. Homologous recombination (HR)2 is a major, conserved pathway for the repair of DNA double strand breaks (DSBs). During meiosis, HR establishes physical linkages between homologous chromosomes from maternal and paternal origins as pairs. These linkages produce chiasmata, which are the cytological manifestation of the crossover products of HR. Together with sister chromatid cohesion, these chromosome crossovers ensure the orderly segregation of each chromosome pair in the first meiotic division (1, 2). The cells derived from the meiotic cell division have half the number of chromosomes as their parent, which is an essential feature of sexual reproduction involving the generation of haploid gametes. Errors in meiotic chromosome segregation lead to miscarriages and Down, Klinefelter, Edwards, and Turner syndromes in humans, which are all characterized by aneuploidy stemming from a deficiency in meiotic chromosome segregation (3-5).Meiotic HR is initiated by cleavage of chromosomal DNA at multiple sites by the Spo11 protein to generate DSBs (6). These DSBs are processed by nucleases to generate 3Ј single-stranded DNA (ssDNA) tails (7,8). A recombinase polymerizes on the ssDNA tails to form a helical nucleoprotein filament, termed the presynaptic filament, capable of searching for, interacting with, and invading a homo...

Mek1 Down Regulates Rad51 Activity during Yeast Meiosis by Phosphorylation of Hed1

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Defining the influence of Rad51 and Dmc1 lineage-specific amino acids on genetic recombination

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Stimulated by retinoic acid gene 8 (Stra8) plays important roles in many stages of spermatogenesis

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