Homologous recombination protects mammalian cells from replication-associated DNA double-strand breaks arising in response to methyl methanesulfonate

Nikolova, Teodora; Ensminger, Michael; Löbrich, Markus; Kaina, Bernd

doi:10.1016/j.dnarep.2010.07.005

Cited by 66 publications

(59 citation statements)

References 55 publications

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“…Since all these effects were abolished following the BRCA2-and XRCC3-defective cells with chromatid exchanges and complex rearrangements involving two or more chromosomes. This is similar to our previous data on HR mutants treated with MMS 29 and indicates misrepair of transiently formed DSB by HR, probably through defective resolution of Holiday junctions. This observation is in agreement with reports on resolvase activity associated with the RAD51C-XRCC3 complex in human cells.…”

Section: Discussionsupporting

confidence: 81%

“…36,37 Recently, another NHEJ protein, ligase 4, was found to be a molecular target that was related to ACNU sensitivity in human glioblastoma cells. 38 As outlined above, in unsynchronized cells, we observed a high degree of colocalization of γH2AX and 53BP1 in nuclear foci, which is in agreement with our previous data with MMS-induced replication-dependent DSB, 29 supporting the notion that 53BP1 localization and retention to sites of DSBs requires phosphorylation of H2AX. 39 A comparison of the rate of foci disappearance between 8 and 32 h post-exposure revealed that cells lacking either BRCA2, XRCC3 or Rad51D showed a highly elevated foci level up to 32 h, whereas in wt cells, the frequency was declining with time, which supports the view that ACNU-induced DSB are primarily repaired by XRCC3-, Rad51D-and BRCA2-dependent functions.…”

Section: Discussionsupporting

confidence: 81%

“…Similar data were previously obtained with methyl methanesulfonate, against which DNAPKcs did not provoke resistance. 29 Overall, the data support and extend recent reports showing that HR is involved in the repair of DNA damage induced by CNU. 36,37 Recently, another NHEJ protein, ligase 4, was found to be a molecular target that was related to ACNU sensitivity in human glioblastoma cells.…”

Section: Discussionsupporting

confidence: 79%

“…The BRCA2, XRCC3 and Rad51D mutants are cytogenetically quite unstable, exhibiting a high spontaneous frequency of aberrations. 29 In the present experiments, we observed a spontaneous frequency of ≈0.2 aberrations per cell for these mutants. After ACNU treatment, the yield of chromosomal aberrations increased 15-30 times compared with the corresponding wt.…”

Section: Experiments With Synchronized Cellsmentioning

confidence: 48%

“…Slide preparation was done as previously described. 29 The following aberrations were counted: chromatid breaks, chromatid translocations (triradials, quadriradials) and intercalary deletions. Gaps were not included in the final evaluation.…”

Section: Acknowledgmentsmentioning

confidence: 99%

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Chloroethylnitrosourea-induced cell death and genotoxicity: Cell cycle dependence and the role of DNA double-strand breaks, HR and NHEJ

et al. 2012

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C hloroethylnitrosureas (CNUs) are powerful DNA-reactive alkylating agents used in cancer therapy. Here, we analyzed cyto-and genotoxicity of nimustine (ACNU), a representative of CNUs, in synchronized cells and in cells deficient in repair proteins involved in homologous recombination (HR) or nonhomologous end-joining (NHEJ). We show that HR mutants are extremely sensitive to ACNU, as measured by colony formation, induction of apoptosis and chromosomal aberrations. The NHEJ mutants differed in their sensitivity, with Ku80 mutants being moderately sensitive and DNA-PKcs mutated cells being resistant. HR mutated cells displayed a sustained high level of γH2AX foci , which co-stained with Rad51 and 53BP1, indicating DNA double-strand breaks (DSB) to be formed. Using synchronized cells, we analyzed whether DSB formation after ACNU treatment was replication-dependent. We show that γH2AX foci were not induced in G 1 but increased significantly in S phase and remained at a high level in G 2 , where a fraction of cells became arrested and underwent, with a delay of > 12 h, cell death by apoptosis and necrosis. Rad51, ATM, MDC-1 and RPA-2 foci were also formed and shown to co-localize with γH2AX foci induced in S phase, indicating that the DNA damage response was activated. All effects observed were abrogated by MGMT, which repairs O 6 -chloroethylguanine that is converted into DNA cross-links. We deduce that the major genotoxic and killing lesion induced by CNUs are O 6 -chloroethylguanine-triggered

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Section: Discussionsupporting

confidence: 81%

Section: Discussionsupporting

confidence: 81%

Section: Discussionsupporting

confidence: 79%

Section: Experiments With Synchronized Cellsmentioning

confidence: 48%

Section: Acknowledgmentsmentioning

confidence: 99%

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Chloroethylnitrosourea-induced cell death and genotoxicity: Cell cycle dependence and the role of DNA double-strand breaks, HR and NHEJ

et al. 2012

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Creation of a Yeast Strain with Co‐Translationally Acylated Nucleosomes

Liu

Zhang

et al. 2022

Angewandte Chemie

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Structurally diverse acylations have been identified as post-translational modifications (PTMs) on histone lysine residues, but their functions and regulations remain largely unknown. Interestingly, in nature, a lysine acylation analog, pyrrolysine, is introduced as a co-translational modification (CTM) through genetic encoding. To explore this alternative life form, we created a model organism Saccharomyces cerevisiae containing site-specific lysine CTMs (acetyllysine, crotonyl-lysine, or another synthetic analog) at histone H3K56 using non-canonical amino acid mutagenesis to afford a chemically modified nucleosome in lieu of their own in vivo. We further demonstrated that acetylation of histone H3K56 partly tends to provide a more favorable chromatin environment for DNA repair in yeast compared to crotonylation and crosstalk with other PTMs differently. This study provides a potentially universal approach to decipher the consequences of different histone lysine PTMs in eukaryotes.

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