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Nucleosomes protect DNA from irradiation-induced double strand breaks in living cells [BLISS human]

ABSTRACT: The number and position of breaks using BLISS-seq was measured in MCF-7 cells upon irradiation with 10 Gy and with or without the use of the antioxidant (N-acetylcysteine, NAC, 10mM).

ORGANISM(S): Homo sapiens

PROVIDER: GSE150016 | GEO | 2020/07/07

REPOSITORIES: GEO

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Nucleosomes protect DNA from irradiation-induced double strand breaks in living cells (BLISS)

Project description:The depletion of HMGB1 reduces the deposition of histones on naked DNA (Celona et al, 2011). in primary MEFs and yeast. We verified that the downmodulation of HMGB1 decreases histone level also in a model of murine mesothelioma. HMGB1 was KD in Murine Mesothelioma AB22 cells by the expression of a shRNA. Nucleosome positioning and occupancy were tested by ATAC-seq. Since a reduced amount of nucleosomes increase the cell sensitivity to gamma irradiatio, we tested the number and position of breaks using BLISS-seq.

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Nucleosomes protect DNA from irradiation-induced double strand breaks in living cells (BLISS)

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Nucleosomes protect DNA from irradiation-induced double strand breaks in living cells

Project description:This SuperSeries is composed of the SubSeries listed below.

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Nucleosomes protect DNA from irradiation-induced double strand breaks in living cells

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Genome-wide detection of DNA double-strand breaks by in-suspension BLISS

Project description:sBLISS (in-suspension Break Labeling In Situ and Sequencing) is a versatile and widely applicable method for identification of endogenous and induced DNA double-strand breaks (DSBs), in any cell type that can be brought into suspension. After in situ labeling, DSB ends are linearly amplified followed by next-generation sequencing and DSB landscape analysis. Here, we present a step-by-step experimental protocol for sBLISS, followed by a basic computational analysis. The main advantages of sBLISS are (i) the suspension setup, which circumvents the need to work with delicate coverslips as in the original BLISS, (ii) the possibility for adaptation to a high-throughput robotics or single-cell workflow, and (iii) its flexibility and applicability to virtually every cell type, including patient-derived cells, organoids, and isolated nuclei. The wet-lab protocol can be completed in 1.5 weeks, and it is suitable for researchers with intermediate expertise in molecular biology and genomics. For the computational analyses, basic-to-intermediate bioinformatics expertise is required.

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