Abstract
Origins of DNA replication are licensed in G1 by recruiting the minichromosome maintenance (MCM) proteins to form a prereplicative complex (pre-RC). Prior to initiation of DNA synthesis from each origin, a preinitiation complex (pre-IC) containing Cdc45 and other proteins is formed. We report that Cdc7-Dbf4 protein kinase (DDK) promotes assembly of a stable Cdc45-MCM complex exclusively on chromatin in S phase. In this complex, Mcm4 is hyperphosphorylated. Studies in vitro using purified DDK and Mcm4 demonstrate that hyperphosphorylation occurs at the Mcm4 N terminus. However, the DDK substrate specificity is conferred by an adjacent DDK-docking domain (DDD), sufficient for facilitating efficient phosphorylation of artificial phosphoacceptors in cis. Genetic evidence suggests that phosphorylation of Mcm4 by DDK is important for timely S phase progression and for cell viability upon overproduction of Cdc45. We suggest that DDK docks on and phosphorylates MCM proteins at licensed origins to promote proper assembly of pre-IC.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Motifs
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Apoenzymes / metabolism
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Binding Sites
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Cell Cycle Proteins / chemistry
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Cell Cycle Proteins / metabolism*
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Cell Cycle Proteins / physiology
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Chromatin / metabolism
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Conserved Sequence
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DNA Replication
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DNA-Binding Proteins / chemistry
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DNA-Binding Proteins / metabolism
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DNA-Binding Proteins / physiology
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Minichromosome Maintenance Complex Component 4
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Models, Biological
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Molecular Sequence Data
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Nuclear Proteins / chemistry
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Nuclear Proteins / metabolism
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Nuclear Proteins / physiology
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Phosphorylation
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Protein Serine-Threonine Kinases / chemistry
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Protein Serine-Threonine Kinases / metabolism*
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Protein Serine-Threonine Kinases / physiology
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Protein Structure, Tertiary
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S Phase / physiology*
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Saccharomyces cerevisiae / cytology
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins / chemistry
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Saccharomyces cerevisiae Proteins / metabolism*
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Saccharomyces cerevisiae Proteins / physiology
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Sequence Alignment
Substances
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Apoenzymes
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CDC45 protein, S cerevisiae
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Cell Cycle Proteins
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Chromatin
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DNA-Binding Proteins
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Dbf4 protein, S cerevisiae
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Nuclear Proteins
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Saccharomyces cerevisiae Proteins
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CDC7 protein, S cerevisiae
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Protein Serine-Threonine Kinases
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MCM4 protein, S cerevisiae
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Minichromosome Maintenance Complex Component 4