Abstract
The Dbf4/Drf1-dependent S-phase-promoting kinase Cdc7 (Ddk) is thought to be an essential target inactivated by the S-phase checkpoint machinery that inhibits DNA replication. However, we show here that the complex formation, chromatin association, and kinase activity of Ddk are not inhibited during the DNA-damage-induced S-phase checkpoint response in Xenopus egg extracts and mammalian cells. Instead, we find that Ddk plays an active role in regulating S-phase checkpoint signaling. Addition of purified Ddk to Xenopus egg extracts or overexpression of Dbf4 in HeLa cells downregulates ATR-Chk1 checkpoint signaling and overrides the inhibition of DNA replication and cell-cycle progression induced by DNA-damaging agents. These results indicate that Ddk functions as an upstream regulator to monitor S-phase checkpoint signaling. We propose that Ddk modulates the S-phase checkpoint control by attenuating checkpoint signaling and triggering DNA replication reinitiation during the S-phase checkpoint recovery.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Adaptor Proteins, Signal Transducing / metabolism*
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Animals
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Cell Cycle Proteins / metabolism*
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Cell Extracts
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Checkpoint Kinase 1
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Chromatin / drug effects
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Chromatin / metabolism
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Chromosomal Proteins, Non-Histone / metabolism*
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DNA Damage*
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Down-Regulation / drug effects
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Etoposide / pharmacology
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Formins
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HeLa Cells
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Humans
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Oocytes / cytology
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Oocytes / drug effects
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Oocytes / enzymology
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Phosphorylation / drug effects
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Protein Kinases / metabolism
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Protein Serine-Threonine Kinases / metabolism*
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S Phase / drug effects
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Signal Transduction / drug effects
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Xenopus
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Xenopus Proteins / metabolism*
Substances
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Adaptor Proteins, Signal Transducing
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Cell Cycle Proteins
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Cell Extracts
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Chromatin
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Chromosomal Proteins, Non-Histone
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DBF4 protein, human
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DBF4B protein, Xenopus
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DIAPH1 protein, human
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Dbf4 protein, Xenopus
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Formins
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Xenopus Proteins
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Etoposide
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Protein Kinases
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CDC7 protein, Xenopus
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CDC7 protein, human
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CHEK1 protein, human
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Checkpoint Kinase 1
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Chek1 protein, Xenopus
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Protein Serine-Threonine Kinases