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Mismatch repair proteins regulate heteroduplex formation during mitotic recombination in yeast
1Graduate Program in Genetics and Molecular Biology and Department of Biology, Emory University, Atlanta, Georgia 30322, USA.
Abstract:
Mismatch repair (MMR) proteins actively inhibit recombination between diverged sequences in both prokaryotes and eukaryotes. Although the molecular basis of the antirecombination activity exerted by MMR proteins is unclear, it presumably involves the recognition of mismatches present in heteroduplex recombination intermediates. This recognition could be exerted during the initial stage of strand exchange, during the extension of heteroduplex DNA, or during the resolution of recombination intermediates. We previously used an assay system based on 350-bp inverted-repeat substrates to demonstrate that MMR proteins strongly inhibit mitotic recombination between diverged sequences in Saccharomyces cerevisiae. The assay system detects only those events that reverse the orientation of the region between the recombination substrates, which can occur as a result of either intrachromatid crossover or sister chromatid conversion. In the present study we sequenced the products of mitotic recombination between 94%-identical substrates in order to map gene conversion tracts in wild-type versus MMR-defective yeast strains. The sequence data indicate that (i) most recombination occurs via sister chromatid conversion and (ii) gene conversion tracts in an MMR-defective strain are significantly longer than those in an isogenic wild-type strain. The shortening of conversion tracts observed in a wild-type strain relative to an MMR-defective strain suggests that at least part of the antirecombination activity of MMR proteins derives from the blockage of heteroduplex extension in the presence of mismatches.
Insights
Mismatch repair (MMR) proteins inhibit recombination between diverged DNA sequences. In yeast, MMR deficiency leads to longer gene conversion tracts, suggesting MMR proteins block heteroduplex extension.
Area of Science:
- Molecular Biology
- Genetics
- DNA Repair
Background:
- Mismatch repair (MMR) proteins are crucial for maintaining genomic stability by correcting errors during DNA replication and recombination.
- The precise mechanism by which MMR proteins inhibit recombination between diverged sequences remains incompletely understood.
- MMR's antirecombination activity is hypothesized to involve recognition of mismatches within recombination intermediates.
Purpose of the Study:
- To investigate the role of MMR proteins in regulating gene conversion tract length during mitotic recombination.
- To elucidate the molecular basis of MMR's antirecombination activity by comparing recombination products in wild-type and MMR-defective yeast strains.
Main Methods:
- Utilized an established assay system with 350-bp inverted-repeat substrates in Saccharomyces cerevisiae.
- Sequenced products of mitotic recombination between 94%-identical substrates in both wild-type and MMR-defective yeast strains.
- Mapped gene conversion tracts to quantify differences in length and characterize recombination events.
Main Results:
- Demonstrated that most mitotic recombination events between diverged sequences occur via sister chromatid conversion.
- Observed significantly longer gene conversion tracts in MMR-defective yeast strains compared to isogenic wild-type strains.
- The shortening of conversion tracts in wild-type strains indicates MMR's role in limiting heteroduplex extension.
Conclusions:
- MMR proteins actively suppress recombination between diverged DNA sequences.
- The antirecombination activity of MMR proteins is, in part, mediated by blocking heteroduplex DNA extension when mismatches are present.
- Findings provide mechanistic insight into how MMR proteins contribute to genome stability by limiting aberrant recombination events.