Abstract
The histone H3 variant CENP-A is required for epigenetic specification of centromere identity through a loading mechanism independent of DNA sequence. Using multiphoton absorption and DNA cleavage at unique sites by I-SceI endonuclease, we demonstrate that CENP-A is rapidly recruited to double-strand breaks in DNA, along with three components (CENP-N, CENP-T, and CENP-U) associated with CENP-A at centromeres. The centromere-targeting domain of CENP-A is both necessary and sufficient for recruitment to double-strand breaks. CENP-A accumulation at DNA breaks is enhanced by active non-homologous end-joining but does not require DNA-PKcs or Ligase IV, and is independent of H2AX. Thus, induction of a double-strand break is sufficient to recruit CENP-A in human and mouse cells. Finally, since cell survival after radiation-induced DNA damage correlates with CENP-A expression level, we propose that CENP-A may have a function in DNA repair.
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.
MeSH terms
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Animals
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Autoantigens / chemistry
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Autoantigens / genetics
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Autoantigens / metabolism*
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Biological Transport, Active
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Cell Line
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Centromere / metabolism
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Centromere Protein A
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Chromosomal Proteins, Non-Histone / chemistry
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Chromosomal Proteins, Non-Histone / genetics
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Chromosomal Proteins, Non-Histone / metabolism*
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DNA Breaks, Double-Stranded*
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DNA Damage / physiology
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DNA Repair / physiology
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Deoxyribonucleases, Type II Site-Specific / metabolism
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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Histones / metabolism
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Humans
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Kinetics
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Mice
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Models, Biological
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Protein Structure, Tertiary
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
Substances
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Autoantigens
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CENPA protein, human
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Cenpa protein, mouse
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Centromere Protein A
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Chromosomal Proteins, Non-Histone
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H2AX protein, human
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Histones
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Recombinant Fusion Proteins
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Green Fluorescent Proteins
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Deoxyribonucleases, Type II Site-Specific