A Double-Strand Break Repair Defect in ATM-Deficient Cells Contributes to Radiosensitivity

Kühne, Martin; Riballo, Enriqueta; Rief, Nicole; Rothkamm, Kai; Jeggo, Penny A.; Löbrich, Markus

doi:10.1158/0008-5472.can-03-2384

Cited by 324 publications

(258 citation statements)

References 59 publications

(51 reference statements)

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“…Pulsed-field gel electrophoresis (PFGE) was carried out as described before [18]. Cells were harvested and about 1×10 5 cells were embedded in 1% low-melting agarose.…”

Section: Pulsed-field Gel Electrophoresis and Colony Formation Assaysmentioning

confidence: 99%

Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair

Fang

Wang

et al. 2007

Cell Res

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“…Pulsed-field gel electrophoresis (PFGE) was carried out as described before [18]. Cells were harvested and about 1×10 5 cells were embedded in 1% low-melting agarose.…”

Section: Pulsed-field Gel Electrophoresis and Colony Formation Assaysmentioning

confidence: 99%

Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair

Fang

Wang

et al. 2007

Cell Res

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“…We found no difference in DNA repair capacity using any of these tests. These assays were chosen because they are rapid and simple, and they have successfully been employed to screen for radiation-sensitive individuals and individuals with cancer-prone syndromes (Rothfuss et al, 1998;Djuzenova et al, 1999;Kuhne et al, 2004). As a panel, the three tests measure both the immediate effect of radiation on DNA breakage, and DNA repair capacity up to 24 h post-irradiation.…”

Section: Discussionmentioning

confidence: 99%

“…g-H2AX nuclear foci are counted using immunofluorescence. Detection of foci is used to evaluate the induction of radiation-induced double-stranded DNA breaks in lymphocytes (Petersen et al, 2001;Fernandez-Capetillo et al, 2003;Pilch et al, 2003;Rothkamm and Lobrich, 2003;Kuhne et al, 2004). The alkaline comet assay measures both acute DNA damage and rate of repair (Olive, 2002) and was measured immediately post-irradiation and 1 h thereafter.…”

Section: Discussionmentioning

confidence: 99%

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

et al. 2007

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The BRCA1 gene product helps to maintain genomic integrity through its participation in the cellular response to DNA damage: specifically, the repair of double-stranded DNA breaks. An impaired cellular response to DNA damage is a plausible mechanism whereby BRCA1 mutation carriers are at increased risk of breast cancer. Hence, an individual's capacity to repair DNA may serve as a useful biomarker of breast cancer risk. The overall aim of the current study was to identify a biomarker of DNA repair capacity that could distinguish between BRCA1 mutation carriers and non-carriers. DNA repair capacity was assessed using three validated assays: the single-cell alkaline gel electrophoresis (comet) assay, the micronucleus test, and the enumeration of g-H2AX nuclear foci. DNA repair capacity of peripheral blood lymphocytes from 25 cancer-free female heterozygous BRCA1 mutation carriers and 25 noncarrier controls was assessed at baseline and following cell exposure to g -irradiation (2 Gy). We found no significant differences in the mean tail moment, in the number of micronuclei or in the number of g-H2AX nuclear foci between the carriers and non-carriers at baseline, and following g-irradiation. These data suggest that these assays are not likely to be useful in the identification of women at a high risk for breast cancer. British Journal of Cancer (2007) The inheritance of a deleterious mutation in the breast cancer susceptibility gene, BRCA1, has been associated with a lifetime risk of breast cancer of between 45 and 87% (Ford et al, 1998;Antoniou et al, 2003). Genetic, reproductive, and environmental factors have all been suggested to influence breast cancer risk in BRCA1 mutation carriers (reviewed in Narod and Offit (2005)). The use of breast cancer as an endpoint to evaluate the protective role of potential modifying factors is not always feasible. Thus, there is a need to ascertain biomarkers of cancer susceptibility in these women. In turn, the identification of a valid biomarker of breast cancer risk would allow us to identify mutation carriers and help improve our ability to target, and to evaluate the effect of both dietary and lifestyle alterations, as well as, medical diagnostic and therapeutic interventions on breast cancer risk.The BRCA1 protein plays a vital role in maintaining genomic stability because of its key role in the repair of double-stranded DNA breaks by homologous recombination (reviewed in Welcsh et al (2000)). If double-stranded DNA breaks are left unrepaired, or are repaired inaccurately, aberrant chromosome breaks, deletions, and translocations may accumulate. Thus, an impaired cellular response to DNA damage appears to be a plausible mechanism whereby BRCA1 mutation carriers are at an increased risk of breast cancer (Scott, 2004). Hence, the evaluation of an individual's capacity to repair DNA may serve as a biomarker of breast cancer risk in carriers of these mutations.Two previous studies have shown higher levels of chromosomal aberrations and chromosomal breaks among women with a BRCA...

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“…Recently, γH2AX foci analysis has become a widely used tool to quantify DSBs on a single cell level after IR (2). γH2AX foci numbers of 30-35 per Gy per G1 cell have been reported (3)(4)(5)(6)(7)(8) suggesting that there is a 1:1 correlation between γH2AX foci and DSBs after IR of nonreplicating cells. However, the precise DSB induction yields following irradiation can depend on a number of factors (see, eg, (9)) and, we show here, are critically dependent upon the surface upon which the cells are grown and irradiated.…”

Section: Dna Double-strand Breaks (Dsbs) Represent Important Lesions mentioning

confidence: 99%

X-irradiation of cells on glass slides has a dose doubling impact

et al. 2007

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Immunofluorescence detection of γH2AX foci is a widely used tool to quantify the induction and repair of DNA double-strand breaks induced by ionising radiation. We observed that Xirradiation of mammalian cells exposed on glass slides induced 2-fold higher foci numbers compared to irradiation with γ-rays. Here, we show that the excess γH2AX foci after Xirradiation are produced from secondary radiation particles generated from the irradiation of glass slides. Both 120 kV X-rays and 137 Cs γ-rays induce ~20 γH2AX foci per Gy in cells growing on thin (~2 µm) plastic foils immersed in water. The same yield is obtained following γ-irradiation of cells growing on glass slides. However, 120 kV X-rays produce ~40 γH2AX foci per Gy in cells growing on glass, two-fold greater than obtained using cells irradiated on plastic surfaces. The same increase in γH2AX foci number is obtained if the plastic foil on which the cells are grown is irradiated on a glass slide. Thus, the physical proximity to the glass material and not morphological differences of cells growing on different surfaces accounts for the excess γH2AX foci. The increase in foci number depends on the energy and is considerably smaller for 25 kV relative to 120 kV X-rays, a finding which can be explained by known physical properties of radiation. The kinetics for the loss of foci, which is taken to represent the rate of DSB repair, as well as the Artemis dependent repair fraction, was similar following X-or γ-irradiation, demonstrating that DSBs induced by this range of treatments are repaired in an identical manner.3

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A Double-Strand Break Repair Defect in ATM-Deficient Cells Contributes to Radiosensitivity

Cited by 324 publications

References 59 publications

Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair

Cell polarity protein Par3 complexes with DNA-PK via Ku70 and regulates DNA double-strand break repair

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

X-irradiation of cells on glass slides has a dose doubling impact

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