Abstract
The protein kinase Hsk1 is essential for DNA replication in Schizosaccharomyces pombe. It associates with Dfp1/Him1 to form an active complex equivalent to the Cdc7-Dbf4 protein kinase in Saccharomyces cerevisiae. Swi1 and Swi3 are subunits of the replication fork protection complex in S. pombe that is homologous to the Tof1-Csm3 complex in S. cerevisiae. The fork protection complex helps to preserve the integrity of stalled replication forks and is important for activation of the checkpoint protein kinase Cds1 in response to fork arrest. Here we describe physical and genetic interactions involving Swi1 and Hsk1-Dfp1/Him1. Dfp1/Him1 was identified in a yeast two-hybrid screen with Swi1. Hsk1 and Dfp1/Him1 both co-immunoprecipitate with Swi1. Swi1 is required for growth of a temperature-sensitive hsk1 (hsk1ts) mutant at its semi-permissive temperature. Hsk1ts cells accumulate Rad22 (Rad52 homologue) DNA repair foci at the permissive temperature, as previously observed in swi1 cells, indicating that abnormal single-stranded DNA regions form near the replication fork in hsk1ts cells. hsk1ts cells were also unable to properly delay S-phase progression in the presence of a DNA alkylating agent and were partially defective in mating type switching. These data suggest that Hsk1-Dfp1/Him1 and Swi1-Swi3 complexes have interrelated roles in stabilization of arrested replication forks.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Alkylating Agents / pharmacology
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Alleles
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Animals
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Bacterial Proteins / metabolism
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Cell Cycle
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Cell Cycle Proteins / metabolism*
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Cell Cycle Proteins / physiology*
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Chromosomal Proteins, Non-Histone / genetics
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Chromosomal Proteins, Non-Histone / metabolism*
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DNA / metabolism
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DNA Repair
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DNA Replication
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DNA, Single-Stranded / genetics
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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Escherichia coli / metabolism
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Hydroxyurea / pharmacology
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Immunoprecipitation
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Luminescent Proteins / metabolism
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Methyl Methanesulfonate / chemistry
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Mice
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Models, Genetic
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Mutation
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Plasmids / metabolism
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Protein Binding
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Protein Kinases / genetics
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Protein Kinases / metabolism*
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Protein Serine-Threonine Kinases / metabolism*
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Protein Serine-Threonine Kinases / physiology*
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S Phase
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Saccharomyces cerevisiae Proteins / metabolism*
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Saccharomyces cerevisiae Proteins / physiology*
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Schizosaccharomyces / genetics
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Schizosaccharomyces / physiology*
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Schizosaccharomyces pombe Proteins / genetics
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Schizosaccharomyces pombe Proteins / metabolism*
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Schizosaccharomyces pombe Proteins / physiology*
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Temperature
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Transcription Factors / genetics
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Transcription Factors / metabolism*
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Two-Hybrid System Techniques
Substances
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Alkylating Agents
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Bacterial Proteins
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Cell Cycle Proteins
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Chromosomal Proteins, Non-Histone
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DNA, Single-Stranded
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DNA-Binding Proteins
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Dbf4 protein, S cerevisiae
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HIM1 protein, S cerevisiae
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Luminescent Proteins
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Saccharomyces cerevisiae Proteins
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Schizosaccharomyces pombe Proteins
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Swi3 protein, S pombe
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Transcription Factors
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rad52 protein, S pombe
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swi1 protein, S pombe
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yellow fluorescent protein, Bacteria
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DNA
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Methyl Methanesulfonate
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Protein Kinases
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CDC7 protein, S cerevisiae
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Dfp1 protein, S pombe
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HSK1 protein, S pombe
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Protein Serine-Threonine Kinases
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Hydroxyurea