Abstract
The cellular DNA-damage response is a signaling network that is vigorously activated by cytotoxic DNA lesions, such as double-strand breaks (DSBs). The DSB response is mobilized by the nuclear protein kinase ATM, which modulates this process by phosphorylating key players in these pathways. A long-standing question in this field is whether DSB formation affects chromatin condensation. Here, we show that DSB formation is followed by ATM-dependent chromatin relaxation. ATM's effector in this pathway is the protein KRAB-associated protein (KAP-1, also known as TIF1beta, KRIP-1 or TRIM28), previously known as a corepressor of gene transcription. In response to DSB induction, KAP-1 is phosphorylated in an ATM-dependent manner on Ser 824. KAP-1 is phosphorylated exclusively at the damage sites, from which phosphorylated KAP-1 spreads rapidly throughout the chromatin. Ablation of the phosphorylation site of KAP-1 leads to loss of DSB-induced chromatin decondensation and renders the cells hypersensitive to DSB-inducing agents. Knocking down KAP-1, or mimicking a constitutive phosphorylation of this protein, leads to constitutive chromatin relaxation. These results suggest that chromatin relaxation is a fundamental pathway in the DNA-damage response and identify its primary mediators.
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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Ataxia Telangiectasia Mutated Proteins
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Blotting, Western
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Cell Cycle Proteins / physiology*
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Cell Line
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Cell Line, Tumor
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Cell Survival / drug effects
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Cell Survival / genetics
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Chromatin / metabolism*
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DNA Damage*
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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DNA-Binding Proteins / physiology*
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Dose-Response Relationship, Drug
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Humans
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Microscopy, Fluorescence
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Mutation / genetics
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Nucleic Acid Synthesis Inhibitors / pharmacology
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Phosphorylation
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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Protein Serine-Threonine Kinases / physiology*
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Repressor Proteins / genetics
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Repressor Proteins / metabolism
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Repressor Proteins / physiology*
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Signal Transduction / physiology*
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Tripartite Motif-Containing Protein 28
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Tumor Suppressor Proteins / genetics
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Tumor Suppressor Proteins / metabolism
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Tumor Suppressor Proteins / physiology*
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Zinostatin / pharmacology
Substances
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Cell Cycle Proteins
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Chromatin
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DNA-Binding Proteins
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Nucleic Acid Synthesis Inhibitors
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Repressor Proteins
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Tumor Suppressor Proteins
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Zinostatin
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TRIM28 protein, human
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Tripartite Motif-Containing Protein 28
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ATM protein, human
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Ataxia Telangiectasia Mutated Proteins
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Protein Serine-Threonine Kinases