Abstract
Eukaryotic cells respond to DNA damage and stalled replication forks by activating signaling pathways that promote cell cycle arrest and DNA repair. A systematic screening of the protein kinase small interfering RNA library reveals that Chk1 and ataxia telangiectasia-mutated (ATM) and Rad3-related (ATR) are the main kinases responsible for intra-S-phase checkpoint upon topoisomerase I inhibitor camptothecin-induced DNA damage. It is well known that ATR-Chk1-mediated protein degradation of Cdc25A protein phosphatase is a crucial mechanism conferring this checkpoint activation. Here we describe another mechanism underlying Cdc25A down-regulation in response to DNA damage that occurs at the transcriptional level. We show that activation of tumor suppressor p53 by DNA damage results in inhibition of Cdc25A transcription as a result of activation of transcriptional repressor ATF3 that directly binds to the Cdc25A promoter. In cells deficient in both Chk1 and p53, Cdc25A down-regulation upon camptothecin-induced DNA damage is completely abolished, leading to severe defects in cell cycle checkpoints and remarkable cell death in mitosis. Our findings reveal two independent mechanisms acting in concert in regulation of Cdc25A in DNA damage response. Although Chk1 affects Cdc25A via rapid phosphorylation and protein turnover, inhibition of Cdc25A transcription by p53-ATF3 is required for the maintenance of cell cycle arrest.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Activating Transcription Factor 3 / genetics
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Activating Transcription Factor 3 / metabolism*
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Antineoplastic Agents / pharmacology
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Ataxia Telangiectasia Mutated Proteins
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Camptothecin / pharmacology
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Cell Line, Tumor
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Checkpoint Kinase 1
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DNA Damage / drug effects
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DNA Damage / physiology
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DNA Repair / drug effects
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DNA Repair / physiology
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Down-Regulation / drug effects
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Down-Regulation / physiology
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Humans
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Mitosis / drug effects
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Mitosis / physiology*
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Protein Binding / drug effects
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Protein Binding / physiology
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Protein Kinases / genetics
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Protein Kinases / metabolism*
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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S Phase / drug effects
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S Phase / physiology*
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Signal Transduction / drug effects
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Signal Transduction / physiology
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Transcription, Genetic / drug effects
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Transcription, Genetic / physiology
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Tumor Suppressor Protein p53 / genetics
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Tumor Suppressor Protein p53 / metabolism*
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Tumor Suppressor Proteins / genetics
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Tumor Suppressor Proteins / metabolism
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cdc25 Phosphatases / genetics
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cdc25 Phosphatases / metabolism*
Substances
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ATF3 protein, human
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Activating Transcription Factor 3
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Antineoplastic Agents
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Cell Cycle Proteins
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DNA-Binding Proteins
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TP53 protein, human
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Tumor Suppressor Protein p53
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Tumor Suppressor Proteins
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Protein Kinases
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ATM protein, human
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ATR protein, human
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Ataxia Telangiectasia Mutated Proteins
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CHEK1 protein, human
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Checkpoint Kinase 1
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Protein Serine-Threonine Kinases
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CDC25A protein, human
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cdc25 Phosphatases
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Camptothecin