| Cell cycle-dependent expression of phosphorylated histone H2AX: reduced expression in unirradiated but not X-irradiated G1-phase cells. | |
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MedLine Citation:
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PMID: 12751958 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Exposure of cells to ionizing radiation causes phosphorylation of histone H2AX at sites flanking DNA double-strand breaks. Detection of phosphorylated H2AX (gammaH2AX) by antibody binding has been used as a method to identify double-strand breaks. Although generally performed by observing microscopic foci within cells, flow cytometry offers the advantage of measuring changes in gammaH2AX intensity in relation to cell cycle position. The importance of cell cycle position on the levels of endogenous and radiation-induced gammaH2AX was examined in cell lines that varied in DNA content, cell cycle distribution, and kinase activity. Bivariate analysis of gammaH2AX expression relative to DNA content and synchronization by centrifugal elutriation were used to measure cell cycle-specific expression of gammaH2AX. With the exception of xrs5 cells, gammaH2AX level was approximately 3 times lower in unirradiated G(1)-phase cells than S- and G(2)-phase cells, and the slope of the G(1)-phase dose-response curve was 2.8 times larger than the slope for S-phase cells. Cell cycle differences were confirmed using immunoblotting, indicating that reduced antibody accessibility in intact cells was not responsible for the reduced antibody binding in G(1)-phase cells. Early apoptotic cells could be easily identified on flow histograms as a population with 5-10-fold higher levels of gammaH2AX, although high expression was not maintained in apoptotic cells by 24 h. We conclude that expression of gammaH2AX is associated with DNA replication in unirradiated cells and that this reduces the sensitivity for detecting radiation-induced double-strand breaks in S- and G(2)-phase cells. |
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Authors:
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Susan H MacPhail; Judit P Banáth; Ying Yu; Eric Chu; Peggy L Olive |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Radiation research Volume: 159 ISSN: 0033-7587 ISO Abbreviation: Radiat. Res. Publication Date: 2003 Jun |
Date Detail:
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Created Date: 2003-05-19 Completed Date: 2003-07-02 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 0401245 Medline TA: Radiat Res Country: United States |
Other Details:
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Languages: eng Pagination: 759-67 Citation Subset: IM; S |
Affiliation:
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Medical Biophysics Department, British Columbia Cancer Research Centre, British Columbia, Vancouver, British Columbia, Canada. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Apoptosis Cell Cycle* DNA / analysis DNA Damage* Flow Cytometry G1 Phase Histones / analysis*, immunology Humans Lymphocytes / drug effects, radiation effects Phosphorylation S Phase Tumor Cells, Cultured X-Rays |
| Chemical | |
Reg. No./Substance:
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0/H2AFX protein, human; 0/Histones; 9007-49-2/DNA |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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