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ABSTRACT:
PROVIDER: PRJNA1005701 | ENA |
REPOSITORIES: ENA
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Project description:Background: Of the many types of DNA damage, DNA double-strand breaks (DSB) are probably the most deleterious. Mounting evidence points to an intricate relationship between DSBs and transcription. A cell system in which the impact on transcription can be investigated at precisely mapped genomic DSBs is essential to study this relationship. Methods: Here in a human cell line, we map genome-wide and at high-resolution the DSBs induced by a restriction enzyme and we characterize their impact on gene expression by four independent approaches by monitoring steady-state RNA levels, rates of RNA synthesis, transcription initiation and RNA polymerase II elongation. Results: We consistently observe transcriptional repression in proximity to DSBs. Downregulation of transcription depends on ATM kinase activity and it is sensitive to the distance from the DSB. Conclusions: Our study couples for the first time high-resolution mapping of DSBs with multilayered transcriptomics to dissect the events shaping gene expression after DSB induction at multiple endogenous sites.
2017-06-01 | GSE97589 | GEO
Project description:DNA Double Strand Breaks (DSBs) are a deleterious product of genotoxic agents or radiotherapy. Despite the inherent chromatin localization of DSBs, the literature has largely ignored the effect of local chromatin or histone post translational modifications (PTMs) on DSB induction or recognition especially in a therapy-relevant setting. Analysis of epigenetic linkages to the DNA Damage Response are challenging owing to the random nature of exogenous DSB induction. Here, we directly measure stochastic DNA damage induced by ionizing radiation (IR) and infer relationships between basal epigenetic states and cellular ability to detect DSBs. Contrary to the expected uniform localization of DSBs and γH2AX, we show that DSB recognition is separate from DSB induction and that both processes are dependent on the local chromatin milieu. In general, actively transcribed regions contain more γH2AX than heterochromatic regions suggesting a link between transcription and the DDR. In contrast to previous studies, we suggest that transcription proximal to DNA damage is necessary for early DSB detection and suggest a requirement for transcription mediated repair in genome maintenance. Finally, we reveal a dual effect of the repressive mark H3K27me3 on the DNA damage response. While basal H3K27me3 attenuates γH2AX deposition, transcribed regions recruit H3K27me3 following DSB induction possibly as an obligate step in DSB recognition. We uncover epigenetic determinants of DSB recognition and suggest new mechanisms by which the epigenome direct DNA repair. Our findings suggest new targets for radio-adjuvant therapy and reframe the current stochastic model of radiotherapy induced damage.
2021-12-31 | GSE181934 | GEO
Project description:DSB-induced transcription of pLac-Tet plasmid in cell-free extracts.
| PRJNA304447 | ENA
Project description:Small RNAs have been implicated in numerous cellular processes, including effects on chromatin structure and the repression of transposons. We describe the generation of a small RNA response at DNA ends in Drosophila that is analogous to the recently reported DSB-induced RNAs (diRNAs) or Dicer and Drosha dependent small RNAs (ddRNAs) in Arabidopsis and vertebrates. Active transcription in the vicinity of the break amplifies this small RNA response, demonstrating that the normal mRNA contributes to the endo-siRNA precursor. The double-stranded RNA precursor forms with an antisense transcript that initiates at the DNA break. Breaks are thus sites of transcription initiation, a novel aspect of the cellular DSB response. This response is specific to a double-strand break since nicked DNA structures do not trigger small RNA production. The small RNAs are generated independently of the exact end structure (blunt, 3'- or 5'-overhang), can repress homologous sequences in trans and may therefore - in addition to putative roles in repair - exert a quality control function by clearing potentially truncated messages from genes in the vicinity of the break.
2012-08-03 | GSE38967 | GEO
Project description:Meiotic recombination is initiated by developmentally programmed DNA double-strand breaks (DSBs). In S. cerevisiae, the vast majority of DSBs occur in the nucleosome-depleted regions at gene promoters, where transcription factors (TFs) bind. It has been proposed that TF binding can stimulate DSB formation nearby by modulating local chromatin structure. However, a prior study in TF bas1 mutant suggested that the role of TF binding in determining break formation is complex. Here, we examined fine-scale DSB distributions in TF mutant (bas1Δ and ino4Δ) strains. In bas1Δ mutants, 239 out of the 2468 hotspots showed reduced DSB activity, whereas 87 hotspots showed increased DSB activity. Similarly, in ino4Δ mutant, 415 out of the 2468 hotspots showed reduced DSB activity, whereas 322 hotspots showed increased DSB activity. We also mapped Bas1 and Ino4 binding sites in meiosis and found that only a small portion of the affected hotspots contained TF binding sites. This indicates that TF can influence DSB distribution both directly and indirectly. Surprisingly, these DSB changes in TF mutants did not correlate with change in chromatin structure and histone H3K4me3 modification, suggesting that the role of TF on DSB distribution cannot be simply explained by affecting local chromatin status. Nine samples total: two wild type, four bas1 mutant and three ino4 mutant (each an independent culture)
2015-08-21 | E-GEOD-67910 | biostudies-arrayexpress
Project description:Meiotic recombination is initiated by developmentally programmed DNA double-strand breaks (DSBs). In S. cerevisiae, the vast majority of DSBs occur in the nucleosome-depleted regions at gene promoters, where transcription factors (TFs) bind. It has been proposed that TF binding can stimulate DSB formation nearby by modulating local chromatin structure. However, a prior study in TF bas1 mutant suggested that the role of TF binding in determining break formation is complex. Here, we examined fine-scale DSB distributions in TF mutant (bas1Δ and ino4Δ) strains. In bas1Δ mutants, 239 out of the 2468 hotspots showed reduced DSB activity, whereas 87 hotspots showed increased DSB activity. Similarly, in ino4Δ mutant, 415 out of the 2468 hotspots showed reduced DSB activity, whereas 322 hotspots showed increased DSB activity. We also mapped Bas1 and Ino4 binding sites in meiosis and found that only a small portion of the affected hotspots contained TF binding sites. This indicates that TF can influence DSB distribution both directly and indirectly. Surprisingly, these DSB changes in TF mutants did not correlate with change in chromatin structure and histone H3K4me3 modification, suggesting that the role of TF on DSB distribution cannot be simply explained by affecting local chromatin status. Twelve samples total: four wild type, four bas1 mutant and four ino4 mutant (each an independent culture has one H3K4me3 ChIP sample and one H3 ChIP sample)
2015-08-21 | E-GEOD-67907 | biostudies-arrayexpress
Project description:Meiotic recombination is initiated by developmentally programmed DNA double-strand breaks (DSBs). In S. cerevisiae, the vast majority of DSBs occur in the nucleosome-depleted regions at gene promoters, where transcription factors (TFs) bind. It has been proposed that TF binding can stimulate DSB formation nearby by modulating local chromatin structure. However, a prior study in TF bas1 mutant suggested that the role of TF binding in determining break formation is complex. Here, we examined fine-scale DSB distributions in TF mutant (bas1Δ and ino4Δ) strains. In bas1Δ mutants, 239 out of the 2468 hotspots showed reduced DSB activity, whereas 87 hotspots showed increased DSB activity. Similarly, in ino4Δ mutant, 415 out of the 2468 hotspots showed reduced DSB activity, whereas 322 hotspots showed increased DSB activity. We also mapped Bas1 and Ino4 binding sites in meiosis and found that only a small portion of the affected hotspots contained TF binding sites. This indicates that TF can influence DSB distribution both directly and indirectly. Surprisingly, these DSB changes in TF mutants did not correlate with change in chromatin structure and histone H3K4me3 modification, suggesting that the role of TF on DSB distribution cannot be simply explained by affecting local chromatin status. Four samples total: Bas1-Myc (ChIP and input samples), Ino4-Myc (ChIP and input samples)
2015-08-21 | E-GEOD-67912 | biostudies-arrayexpress
Project description:The DSB-machinery, which induces the programmed DNA double-strand breaks (DSBs) in leptotene and zygotene stages during meiosis, needs to be kept in silence after the initiation of pachytene stage to prevent the activation of DSB checkpoint that may lead to meiotic arrest or apoptosis of germ cells. However, the mechanisms underlying this repression remain largely unknown. Here, we report that ZFP541, a germ cell-specific zinc finger protein, is responsible for the suppression of DSBs formation at late pachytene. Lack of Zfp541 in mice leads to generation of DSBs in late pachytene spermatocytes by DSB formation related-proteins and causes male infertility due to meiotic failure. Plated-based scRNA-seq of Zfp541-/- spermatocytes revealed that ZFP541 negatively regulates many meiotic prophase genes, including genes for DSB formation and their upstream transcriptional regulators, in late pachytene spermatocytes. These results were confirmed by 10x single-cell RNA-seq data on spermatogenesis of Zfp541-/- testes, which suggested that Zfp541 is required for repressing the activation of pre-pachytene gene expression programs from early to late pachytene. ZFP541 ChIP-seq on pachytene and diplotene spermatocytes demonstrated that ZFP541 occupies the promoters of meiosis initiators (e.g., Meiosin and Rxra) and a subset of their downstream genes to repress their transcription, and thus prevent the reactivation of pre-pachytene gene expression programs in pachytene spermatocytes. Thus, our results not only revealed the role of ZFP541 in maintaining the repression of pre-pachytene transcriptional programs in pachytene spermatocytes but also provide new insight into the regulation of meiotic progression by timely turning off pre-pachytene genes.
2021-04-16 | GSE172160 | GEO
Project description:AT-rich interactive domain-containing protein 1A (ARID1A), a SWI/SNF chromatin remodeling complex subunit, is frequently mutated across various cancer entities. ARID1A contributes to DNA damage response pathways, and we previously reported that ARID1A loss is associated with mutational signatures linked to DNA repair defects. Here we show that ARID1A is promoting both DNA double-strand breaks (DSBs) repair pathways, non-homologous end-joining (NHEJ) and homologous recombination (HR). ARID1A is accumulated at DSBs after DNA damage and regulates the chromatin loop formation at DSB sites by interacting with the cohesin subunit RAD21 and CTCF insulator protein promoting their enrichment around the DSBs. Further analysis demonstrates that ARID1A’s involvement in DSB repair is accompanied by alterations of chromatin accessibility and distribution of activating histone marks at DSBs. ARID1A binds to and promotes the recruitment of the HDAC1 to control transcription repression at these DSBs. Altogether, ARID1A simultaneously regulates DSB repair and transcription silencing in transcriptionally active chromatin. ARID1A depletion resulted in defective DSB repair, which subsequently led to accumulation of micronuclei, activation of cGAS-STING pathway and an increased expression of immunomodulatory cytokines and chemokines upon IR treatment. Furthermore, low ARID1A expression in cancer patients receiving radiotherapy was associated with higher infiltration of several immune cells. The high mutation rate of ARID1A in various cancer types highlights its clinical relevance as a promising biomarker that correlates with the level of immune regulatory cytokines and can estimate the levels of tumor-infiltrating immune cells and response to combination of radio- and immunotherapy.
2024-06-22 | PXD047225 | Pride
Project description:We have investigated whether components of Polycomb repressive complex 1 (PRC1) are recruited to double-strand breaks (DSBs) generated by inducible expression of the AsiSI restriction enzyme in normal human fibroblasts. Using chromatin immunoprecipitation and deep sequencing (ChIP-seq), which detects PRC1 proteins at common sites throughout the genome, we did not find evidence for recruitment of PRC1 components to AsiSI-induced DSBs. In contrast, the S2056 phosphorylated form of DNA-PKcs and other DNA repair proteins were detected at a subset of AsiSI sites that are predominantly at the 5’ ends of transcriptionally active genes. Our data question the idea that PcG protein recruitment provides a link between DSB repairs and transcriptional repression. Single chromatin preparations from treated and untreated cells (+/- OHT), immunoprecipitated with two antibodies (pDNA-PKcs and MEL18) were sequenced as technical replicates, along with the corresponding input DNAs
2014-07-06 | E-GEOD-55605 | biostudies-arrayexpress