Impaired Repair of Ionizing Radiation-Induced DNA Damage in Cockayne Syndrome Cells

Cramers, Patricia; Verhoeven, Esther E.A.; Filon, A.R.; Rockx, Davy; Santos, Susy J.; Leer, Anneke A. van der; Kleinjans, Jos; Zeeland, A.A. van; Mullenders, Leon H.F.

doi:10.1667/rr1972.1

Cited by 17 publications

(11 citation statements)

References 55 publications

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“…This is in line with an increasing amount of literature indicating that CS (but not XP cells) display increased vulnerability to inducers of oxidative DNA damage, and that CSB and CSA play a role in repair of oxidative DNA damage independent of the NER machinery Cramers et al, 2011;D'Errico et al, 2007;Melis et al, 2012;Nardo et al, 2009;Spivak and Hanawalt, 2006). CSB has been proposed to play a role in several oxidative DNA damage repair pathways, including transcription-coupled base excision repair, genome-wide base excision repair, and mitochondrial base excision repair, but additional repair unrelated mechanisms such as transcriptional bypass could play a role as well (Charlet-Berguerand et al, 2006;Cramers et al, 2011;Menoni et al, 2012;Nouspikel, 2008;Scheibye-Knudsen et al, 2013;Spivak and Hanawalt, 2006;Stevnsner et al, 2008). The increased vulnerability to ionizing radiation of Csb m/m versus Xpa À/À cells does not apply to embryonic stem cells, as Csb m/m and Xpa À/À embryonic stem cells display the same survival after exposure to gamma radiation.…”

Section: Sensitivity Of Csb M/m Cells To Uv and Inducers Of Oxidativesupporting

confidence: 88%

Cockayne syndrome pathogenesis: Lessons from mouse models

Jaarsma

Pluijm

Horst

et al. 2013

Mechanisms of Ageing and Development

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Section: Sensitivity Of Csb M/m Cells To Uv and Inducers Of Oxidativesupporting

confidence: 88%

Cockayne syndrome pathogenesis: Lessons from mouse models

Jaarsma

Pluijm

Horst

et al. 2013

Mechanisms of Ageing and Development

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“…The sensitivity of CSB-defective cells to oxidative DNA damage (10,11) indicates that CSB deficiency is phenotypically distinct from classic NER deficiencies and indicates that CSB function is not limited to monoadduct lesions. Therefore, transcription blockage by DSBs may also elicit CSB function to recruit repair factors most suitable for strand break repair at actively transcribed regions.…”

Section: Csb Function Is Necessary For the Recruitment Of Recombinatimentioning

confidence: 99%

“…As noted, in addition to UV sensitivity, CS patients also manifest severe neurodegeneration (8,9), suggesting the importance of CS proteins in maintaining genome stability against a broad spectrum of DNA damage. For example, CSB-defective cells are also sensitive to ionizing radiation (IR) (10,11), which is phenotypically distinctive from classic NER deficiencies and indicates that CSB function is not limited to UV-derived photo lesions. In addition, CSB-deficient mice exhibit a subset of symptoms found in patients, primarily UVC sensitivity and susceptibility to skin cancer (12).…”

mentioning

confidence: 99%

DNA damage during the G0/G1 phase triggers RNA-templated, Cockayne syndrome B-dependent homologous recombination

Wei

Nakajima

Böhm

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

132

138

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Damage repair mechanisms at transcriptionally active sites during the G0/G1 phase are largely unknown. To elucidate these mechanisms, we introduced genome site-specific oxidative DNA damage and determined the role of transcription in repair factor assembly. We find that KU and NBS1 are recruited to damage sites independent of transcription. However, assembly of RPA1, RAD51C, RAD51, and RAD52 at such sites is strictly governed by active transcription and requires both wild-type Cockayne syndrome protein B (CSB) function and the presence of RNA in the G0/G1 phase. We show that the ATPase activity of CSB is indispensable for loading and binding of the recombination factors. CSB counters radiation-induced DNA damage in both cells and zebrafish models. Taken together, our results have uncovered a novel, RNA-based recombination mechanism by which CSB protects genome stability from strand breaks at transcriptionally active sites and may provide insight into the clinical manifestations of Cockayne syndrome.NA double strand breaks (DSBs) are a most severe type of DNA damage caused by endogenous metabolic processes and exogenous exposure to radiation and chemicals. Unrepaired DSBs induce genomic instability, carcinogenesis, and premature aging. In mammalian cells, DSBs are repaired by either the nonhomologous end joining (NHEJ) or the homologous recombination (HR) pathway. Although it is a common understanding that HR primarily takes place in response to strand breaks in the S-G2 phases of the cell cycle where the undamaged sister chromatids are present as donor templates, recent studies have suggested that homologous pairing also occurs during the G0/G1 phase and is associated with transcription (1), although the mechanisms remain to be elucidated. At active transcription sites, RNA polymerase II (RNA POLII) can bypass base modifications such as 8-oxo guanine but not single strand breaks (SSBs) and DSBs (2-5), indicating that unrepaired strand breaks at transcriptionally active (TA) sites can be especially deleterious and may lead to secondary damage.The Cockayne syndrome B (CSB) gene is defective in approximately two-thirds of patients with Cockayne syndrome (CS), an autosomal recessive disease with diverse clinical signs including severe growth failure, progressive neurodegeneration, and hypersensitivity to sunlight. CSB has an established role in transcription-coupled nucleotide excision repair (TC-NER) of photo lesions. When RNA POLII is stalled at bulky lesions, CSB is loaded to facilitate NER of the transcribed strand (6, 7). As noted, in addition to UV sensitivity, CS patients also manifest severe neurodegeneration (8, 9), suggesting the importance of CS proteins in maintaining genome stability against a broad spectrum of DNA damage. For example, CSB-defective cells are also sensitive to ionizing radiation (IR) (10, 11), which is phenotypically distinctive from classic NER deficiencies and indicates that CSB function is not limited to UV-derived photo lesions. In addition, CSB-deficient mice exhibit a subset of sy...

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“…If this is true, this raises some interesting questions since TCR is not normally associated with the repair of DNA damage induced by ionising radiation. However, it is worth noting that recent evidence has demonstrated that CSA and CSB proteins may have a role to play in DNA repair distinct from NER, since fibroblasts deficient in CSA or CSB have been shown to exhibit sensitivity to ionising radiation [45]–[49]. The results presented here would suggest that fibroblasts defective in CSA or CSB proteins do have reduced ability to repair strand breaks and DNA damage induced by ionising radiation.…”

Section: Discussionmentioning

confidence: 55%

Use of the Comet-FISH Assay to Compare DNA Damage and Repair in p53 and hTERT Genes following Ionizing Radiation

et al. 2012

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The alkaline single cell gel electrophoresis (comet) assay can be combined with fluorescent in situ hybridisation (FISH) methodology in order to investigate the localisation of specific gene domains within an individual cell. The number and position of the fluorescent signal(s) provides information about the relative damage and subsequent repair that is occurring in the targeted gene domain(s). In this study, we have optimised the comet-FISH assay to detect and compare DNA damage and repair in the p53 and hTERT gene regions of bladder cancer cell-lines RT4 and RT112, normal fibroblasts and Cockayne Syndrome (CS) fibroblasts following γ-radiation. Cells were exposed to 5Gy γ-radiation and repair followed for up to 60 minutes. At each repair time-point, the number and location of p53 and hTERT hybridisation spots was recorded in addition to standard comet measurements. In bladder cancer cell-lines and normal fibroblasts, the p53 gene region was found to be rapidly repaired relative to the hTERT gene region and the overall genome, a phenomenon that appeared to be independent of hTERT transcriptional activity. However, in the CS fibroblasts, which are defective in transcription coupled repair (TCR), this rapid repair of the p53 gene region was not observed when compared to both the hTERT gene region and the overall genome, proving the assay can detect variations in DNA repair in the same gene. In conclusion, we propose that the comet-FISH assay is a sensitive and rapid method for detecting differences in DNA damage and repair between different gene regions in individual cells in response to radiation. We suggest this increases its potential for measuring radiosensitivity in cells and may therefore have value in a clinical setting.

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Impaired Repair of Ionizing Radiation-Induced DNA Damage in Cockayne Syndrome Cells

Cited by 17 publications

References 55 publications

Cockayne syndrome pathogenesis: Lessons from mouse models

Cockayne syndrome pathogenesis: Lessons from mouse models

DNA damage during the G0/G1 phase triggers RNA-templated, Cockayne syndrome B-dependent homologous recombination

Use of the Comet-FISH Assay to Compare DNA Damage and Repair in p53 and hTERT Genes following Ionizing Radiation

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