Abstract
Damage recognition by repair/checkpoint factors is the critical first step of the DNA damage response. DNA double strand breaks (DSBs) activate checkpoint signaling and are repaired by nonhomologous end-joining (NHEJ) and homologous recombination (HR) pathways. However, in vivo kinetics of the individual factor responses and the mechanism of pathway choice are not well understood. We report cell cycle and time course analyses of checkpoint activation by ataxia-telangiectasia mutated and damage site recruitment of the repair factors in response to laser-induced DSBs. We found that MRN acts as a DNA damage marker, continuously localizing at unrepaired damage sites. Damage recognition by NHEJ factors precedes that of HR factors. HR factor recruitment is not influenced by NHEJ factor assembly and occurs throughout interphase. Damage site retention of NHEJ factors is transient, whereas HR factors persist at unrepaired lesions, revealing unique roles of the two pathways in mammalian cells.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Acid Anhydride Hydrolases
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Animals
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Antigens, Nuclear / genetics
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Antigens, Nuclear / metabolism
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Ataxia Telangiectasia Mutated Proteins
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Cell Cycle / physiology*
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Cell Cycle / radiation effects
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Cell Cycle Proteins / metabolism
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Cell Line
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Checkpoint Kinase 2
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Chromosomal Proteins, Non-Histone
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Cohesins
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DNA Damage / physiology*
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DNA Repair / physiology*
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DNA Repair Enzymes / metabolism
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Fungal Proteins / metabolism
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Histones / metabolism
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Humans
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Ku Autoantigen
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Lasers
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MRE11 Homologue Protein
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Mice
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Mice, Knockout
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Nuclear Proteins / metabolism
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Phosphorylation
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Protein Serine-Threonine Kinases / metabolism
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Protein Transport
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Recombination, Genetic
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Signal Transduction / physiology
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Tumor Suppressor Proteins / metabolism
Substances
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Antigens, Nuclear
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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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Fungal Proteins
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H2AX protein, human
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Histones
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MRE11 protein, human
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Mre11a protein, mouse
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NBN protein, human
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Nuclear Proteins
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Tumor Suppressor Proteins
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Checkpoint Kinase 2
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ATM protein, human
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Ataxia Telangiectasia Mutated Proteins
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Atm protein, mouse
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CHEK2 protein, human
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Chek2 protein, mouse
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Protein Serine-Threonine Kinases
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MRE11 Homologue Protein
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Acid Anhydride Hydrolases
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RAD50 protein, human
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Xrcc6 protein, human
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Xrcc6 protein, mouse
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Ku Autoantigen
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DNA Repair Enzymes