Caffeine impairs resection during DNA break repair by reducing the levels of nucleases Sae2 and Dna2

Tsabar, Michael; Eapen, Vinay V.; Mason, Jennifer M.; Memisoglu, Gonen; Waterman, David P.; Long, Marcus J. C.; Bishop, Douglas K.; Haber, James E.

doi:10.1093/nar/gkv520

Cited by 47 publications

(38 citation statements)

References 89 publications

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“…This sensitivity is apparently achieved by regulating the level of Rad53 phosphorylation in response to more DSBs (Mantiero et al 2007). Maintenance of γ-H2AX in the 50-kb region around each of two DSBs is regulated independently so that it is removed around a repaired DSB even as it persists around an unrepaired DSB (Tsabar et al 2015). Here we provide the first evidence that cells can sense the number of repairable DSBs even if one of these DSBs repairs rapidly enough so that it does not by itself elicit detectable Rad53 phosphorylation.…”

Section: Discussionmentioning

confidence: 73%

Asf1 facilitates dephosphorylation of Rad53 after DNA double-strand break repair

et al. 2016

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To allow for sufficient time to repair DNA double-stranded breaks (DSBs), eukaryotic cells activate the DNA damage checkpoint. In budding yeast, Rad53 (mammalian Chk2) phosphorylation parallels the persistence of the unrepaired DSB and is extinguished when repair is complete in a process termed recovery or when the cells adapt to the DNA damage checkpoint. A strain containing a slowly repaired DSB does not require the histone chaperone Asf1 to resume cell cycle progression after DSB repair. When a second, rapidly repairable DSB is added to this strain, Asf1 becomes required for recovery. Recovery from two repairable DSBs also depends on the histone acetyltransferase Rtt109 and the cullin subunit Rtt101, both of which modify histone H3 that is associated with Asf1. We show that dissociation of histone H3 from Asf1 is required for efficient recovery and that Asf1 is required for complete dephosphorylation of Rad53 when the upstream DNA damage checkpoint signaling is turned off. Our data suggest that the requirements for recovery from the DNA damage checkpoint become more stringent with increased levels of damage and that Asf1 plays a histone chaperone-independent role in facilitating complete Rad53 dephosphorylation following repair.

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Section: Discussionmentioning

confidence: 73%

Asf1 facilitates dephosphorylation of Rad53 after DNA double-strand break repair

et al. 2016

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“…Other cell cytotoxicity data indicate that SCR-7 may interfere with cancer growth and proliferation by blocking the sealing of double strand breaks 22–28. Additionally, SCR-7 and other small-molecule inhibitors to DSB repair have been reported as viable options in primary and adjuvant chemotherapies for several cancer types.…”

Section: Resultsmentioning

confidence: 99%

Non-homologous End Joining Inhibitor SCR-7 to Exacerbate Low-dose Doxorubicin Cytotoxicity in HeLa Cells

et al. 2017

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Among the genotoxic drug regimens, doxorubicin (DOX) is known for its high-dose side effects in several carcinomas, including cervical cancer. This study reports on testing the combined use of a DOX genotoxic drug and SCR-7 non-homologous end joining (NHEJ) inhibitor for HeLa cells. An in vitro DNA damaging assay of DOX was performed on plasmid and genomic DNA substrate. In vitro cytotoxicity was investigated using trypan blue dye exclusion, DNA metabolizing, and propidium iodide-based flow cytometric assays. DOX (between 20–100 μM) displayed clear DNA binding and interaction, such as the shearing and smearing of plasmid and genomic DNA. DNA metabolizing assay data indicate that HeLa lysate with DOX and SCR-7 treatment exhibited better in vitro plasmid DNA stability compared with DOX treatment alone. SCR-7 augmented the effects of low-dose DOX by demonstrating enhanced cell death from 15% to 50%. The flow cytometric data also supported that the combination of SCR-7 with DOX lead to a 23% increase in propidium iodide-based HeLa staining, thus indicating enhanced death. In summary, the inhibition of NHEJ DNA repair pathway can potentiate low-dose DOX to produce appreciable cytotoxicity in HeLa cells.

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“…As previously shown [30], in a rad52 Δ strain, where recombination is absent (Figure 1D), there was no re-establishment of nucleosome protection (Figure 2A and S2C). The loss of nucleosome protection is gradual over the first 3 h, consistent with removal of sequences by 5’ to 3’ resection [4, 31] (Figure S2C). Because the unrepaired locus showed no protection of the 5S region, we hypothesize that the reestablishment of nucleosome protection reflects the formation of the repaired product.…”

Section: Resultsmentioning

confidence: 68%

“…Similarly, sequences near the DSB are only fully resected approximately 2 h to 4 h after DSB induction [4, 31]. It is possible that resection leads to the eviction of the 5S nucleosome 2 h after DSB induction, although previous studies show that the RSC complex is responsible for repositioning nucleosomes around an irreparable DSB [7, 8].…”

Section: Discussionmentioning

confidence: 99%

Re-establishment of nucleosome occupancy during double-strand break repair in budding yeast

et al. 2016

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Homologous recombination (HR) is an evolutionary conserved pathway in eukaryotes that repairs a double-strand break (DSB) by copying homologous sequences from a sister chromatid, a homologous chromosome or an ectopic location. Recombination is challenged by the packaging of DNA into nucleosomes, which may impair the process at many steps, from resection of the DSB ends to the re-establishement of nucleosomes after repair. However, nucleosome dynamics during DSB repair have not been well described, primarily because of a lack of well-ordered nucleosomes around a DSB. We designed a system in budding yeast Saccharomyces cerevisiae to monitor nucleosome dynamics during repair of an HO endonuclease-induced DSB. Nucleosome occupancy around the break is lost following DSB formation, by 5’ to 3’ resection of the DSB end. Soon after repair is complete, nucleosome occupancy is partially restored in a repair-dependent but cell cycle-independent manner. Full re-establishment of nucleosome protection back to the level prior to DSB induction is achieved when the cell cycle resumes following repair. These findings may have implications to the mechanisms by which cells sense the completion of repair.

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Caffeine impairs resection during DNA break repair by reducing the levels of nucleases Sae2 and Dna2

Cited by 47 publications

References 89 publications

Asf1 facilitates dephosphorylation of Rad53 after DNA double-strand break repair

Asf1 facilitates dephosphorylation of Rad53 after DNA double-strand break repair

Non-homologous End Joining Inhibitor SCR-7 to Exacerbate Low-dose Doxorubicin Cytotoxicity in HeLa Cells

Re-establishment of nucleosome occupancy during double-strand break repair in budding yeast

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