Abstract
A key stage during homologous recombination is the processing of the Holliday junction, which determines the outcome of the recombination reaction. To dissect the pathways of Holliday junction processing in a eukaryote, we have targeted an Escherichia coli Holliday junction resolvase to the nuclei of fission yeast recombination-deficient mutants and analysed their phenotypes. The resolvase partially complements the UV and hydroxyurea hypersensitivity and associated aberrant mitoses of an rqh1(-) mutant. Rqh1 is a member of the RecQ subfamily of DNA helicases that control recombination particularly during S-phase. Significantly, overexpression of the resolvase in wild-type cells partly mimics the loss of viability, hyper-recombination and 'cut' phenotype of an rqh1(-) mutant. These results indicate that Holliday junctions form in wild-type cells that are normally removed in a non-recombinogenic way, possibly by Rqh1 catalysing their reverse branch migration. We propose that in the absence of Rqh1, replication fork arrest results in the accumulation of Holliday junctions, which can either impede sister chromatid segregation or lead to the formation of recombinants through Holliday junction resolution.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Cell Nucleus / metabolism
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DNA Helicases / deficiency*
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DNA Helicases / genetics
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DNA, Fungal / genetics
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DNA, Fungal / metabolism
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Endodeoxyribonucleases / genetics
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Endodeoxyribonucleases / physiology*
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Escherichia coli / enzymology*
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Escherichia coli Proteins*
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Eukaryotic Cells / metabolism
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Fungal Proteins / genetics
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Fungal Proteins / physiology*
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Genes, Reporter
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Genes, Synthetic
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Genetic Complementation Test
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Green Fluorescent Proteins
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Holliday Junction Resolvases*
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Luminescent Proteins / genetics
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Models, Genetic
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Nuclear Localization Signals
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Nuclear Proteins / genetics
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Nuclear Proteins / physiology
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Phenotype
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Radiation Tolerance / genetics
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Recombinant Fusion Proteins / physiology
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Recombination, Genetic / physiology*
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S Phase
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Schizosaccharomyces / genetics*
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Schizosaccharomyces / metabolism
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Schizosaccharomyces / radiation effects
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Schizosaccharomyces pombe Proteins*
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Ultraviolet Rays
Substances
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DNA, Fungal
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Escherichia coli Proteins
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Fungal Proteins
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Luminescent Proteins
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Nuclear Localization Signals
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Nuclear Proteins
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Recombinant Fusion Proteins
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Schizosaccharomyces pombe Proteins
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Green Fluorescent Proteins
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Endodeoxyribonucleases
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Holliday Junction Resolvases
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RusA protein, E coli
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DNA Helicases
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Rqh1 protein, S pombe