Abstract
The regulated loading of the Mcm2-7 DNA helicase (comprising six related subunits, Mcm2 to Mcm7) into pre-replicative complexes at multiple replication origins ensures precise once per cell cycle replication in eukaryotic cells. The origin recognition complex (ORC), Cdc6 and Cdt1 load Mcm2-7 into a double hexamer bound around duplex DNA in an ATP-dependent reaction, but the molecular mechanism of this origin 'licensing' is still poorly understood. Here we show that both Mcm2-7 hexamers in Saccharomyces cerevisiae are recruited to origins by an essential, conserved carboxy-terminal domain of Mcm3 that interacts with and stimulates the ATPase activity of ORC-Cdc6. ATP hydrolysis can promote Mcm2-7 loading, but can also promote Mcm2-7 release if components are missing or if ORC has been inactivated by cyclin-dependent kinase phosphorylation. Our work provides new insights into how origins are licensed and reveals a novel ATPase-dependent mechanism contributing to precise once per cell cycle replication.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphatases / metabolism*
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Adenosine Triphosphate / metabolism
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Amino Acid Sequence
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Cell Cycle Proteins / metabolism
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Chromosomal Proteins, Non-Histone / metabolism
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DNA Replication / genetics*
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DNA-Binding Proteins / metabolism
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Enzyme Activation
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Hydrolysis
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Minichromosome Maintenance Complex Component 3
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Minichromosome Maintenance Complex Component 7
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Nuclear Proteins / metabolism
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Protein Binding
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Replication Origin / genetics*
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Saccharomyces cerevisiae / cytology
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Saccharomyces cerevisiae / enzymology
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins / metabolism
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Sequence Alignment
Substances
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Cell Cycle Proteins
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Chromosomal Proteins, Non-Histone
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DNA-Binding Proteins
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Nuclear Proteins
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Saccharomyces cerevisiae Proteins
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TAH11 protein, S cerevisiae
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Adenosine Triphosphate
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Adenosine Triphosphatases
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MCM2 protein, S cerevisiae
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MCM3 protein, S cerevisiae
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MCM7 protein, S cerevisiae
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Minichromosome Maintenance Complex Component 3
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Minichromosome Maintenance Complex Component 7