Cell sorting analysis of cell cycle-dependent X-ray sensitivity in end joining-deficient human cells

Non-homologous end joining (NHEJ) plays a major role in the repair of ionizing radiation-induced DNA double-strand breaks (DSBs), especially during the G1-phase of the cell cycle. Using a flow cytometric cell sorter, we fractionated G1- and S/G2-phase cells based on size to assess the DSB-repair act...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 2008-08, Vol.372 (4), p.662-667
Main Authors: Iwabuchi, Kuniyoshi, Hashimoto, Mitsumasa, Matsui, Tadashi, Kurosawa, Aya, Adachi, Noritaka, Date, Takayasu
Format: Article
Language:English
Subjects:
ANIMAL CELLS
CELL CYCLE
Cell Proliferation - drug effects
Cell Separation - methods
Cell sorter
COLONY FORMATION
DNA
DNA Breaks, Double-Stranded
DNA Damage
DNA double-strand breaks
DNA REPAIR
DNA Repair - genetics
DT40
Flow cytometry
Flow Cytometry - methods
G1 Phase - drug effects
G1 Phase - radiation effects
GENETIC RADIATION EFFECTS
HeLa Cells
Histones - analysis
Humans
LABELLING
Nalm-6
Nonhomologous end joining
Radiation Tolerance - genetics
RADIATION, THERMAL, AND OTHER ENVIRONMENTAL POLLUTANT EFFECTS ON LIVING ORGANISMS AND BIOLOGICAL MATERIALS
RADIOSENSITIVITY
Recombination, Genetic - genetics
STRAND BREAKS
X RADIATION
X-ray sensitivity
X-Rays
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Summary:Non-homologous end joining (NHEJ) plays a major role in the repair of ionizing radiation-induced DNA double-strand breaks (DSBs), especially during the G1-phase of the cell cycle. Using a flow cytometric cell sorter, we fractionated G1- and S/G2-phase cells based on size to assess the DSB-repair activity in NHEJ factor-deficient DT40 and Nalm-6 cell lines. Colony formation assays revealed that the X-ray sensitivities of the G1-enriched populations correctly reflected the DSB-repair activities of both the DT40 and Nalm-6 cell lines. Furthermore, as assessed by γ-H2AX foci formation, the sorted cells exhibited less DNA damage than chemically synchronized cells. Given that it does not use fluorescent labeling or chemical agents, this method of cell sorting is simpler and less toxic than other methods, making it applicable to a variety of cell lines, including those that cannot be synchronized by standard chemical treatments.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2008.05.147