Drosha drives the formation of DNA:RNA hybrids around DNA break sites to facilitate DNA damage repair
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE97648 | GEO | 2018/02/18
REPOSITORIES: GEO
Similar Datasets
Project description:Drosha drives the formation of DNA:RNA hybrids around DNA break sites to facilitate DNA damage repair
| PRJNA382559 | ENA
Project description:The DNA damage response (DDR) is the signaling cascade that recognizes DNA double-strand breaks (DSB) and promotes their resolution via the DNA repair pathways of Non-Homologous End Joining (NHEJ) or Homologous Recombination (HR). We and others have shown that DDR activation requires DROSHA. However, whether DROSHA exerts its functions by associating with damage sites, what controls its recruitment and how DROSHA influences DNA repair, remains poorly understood. Here we show that DROSHA associates to DSBs independently from transcription. Neither H2AX, nor ATM nor DNA-PK kinase activities are required for its recruitment to break site. Rather, DROSHA interacts with RAD50 and inhibition of MRN by Mirin treatment abolishes this interaction. MRN inactivation by RAD50 knockdown or mirin treatment prevents DROSHA recruitment to DSB and, as a consequence, also 53BP1 recruitment. During DNA repair, DROSHA inactivation reduces NHEJ and boosts HR frequency. Indeed, DROSHA knockdown also increase the association of downstream HR factors such as RAD51 to DNA ends. Overall, our results demonstrate that DROSHA is recruited at DSBs by the MRN complex and direct DNA repair toward NHEJ.
2021-03-15 | GSE166631 | GEO
Project description:Homologous recombination (HR) is crucial for genetic exchange, accurate repair of DNA double-strand breaks and pivotal for genome integrity. HR uses homologous sequences for repair, but how homology search, the exploration of the genome for homologous DNA sequences, is conducted in the nucleus remains poorly understood. Here, we use time-resolved chromatin immunoprecipitations of repair proteins to monitor homology search in vivo. We found that homology search proceeds by a probing mechanism, which commences around the break and samples preferentially on the broken chromosome. However, elements thought to instruct chromosome loops mediate homology search shortcuts, and centromeres, which cluster within the nucleus, may facilitate homology search on other chromosomes. Our study thus revealed crucial parameters for homology search in vivo and emphasizes the importance of linear distance, chromosome architecture and proximity for recombination efficiency. 2 new custom ChIP-chip platforms used; both Nimblegen; differ in oligo density: (platform 1: 2006-07-18_Scerevisiae_ChIP_Stefan Jentsch MPI Biochemistry S.cerevisiae 385K Tiling Array Version 1) ( platform 2: 100304_Scer2_MS_Chip_Stefan Jentsch MPI Biochemistry S.cerevisiae 135K Tiling Array Version 2) ChIP-chip profiling of DSB repair factors (Rad51, Rad52, RPA, gamma-H2A) upon single inducible DSBs in S.cerevisiae
2013-03-21 | E-GEOD-44844 | biostudies-arrayexpress
Project description:Recent observations show that the single-cell response of p53 to ionizing radiation (IR) is “digital” in that it is the number of oscillations rather than the amplitude of p53 that shows dependence on the radiation dose. We present a model of this phenomenon. In our model, double-strand break (DSB) sites induced by IR interact with a limiting pool of DNA repair proteins, forming DSB–protein complexes at DNA damage foci. The persisting complexes are sensed by ataxia telangiectasia mutated (ATM), a protein kinase that activates p53 once it is phosphorylated by DNA damage. The ATM-sensing module switches on or off the downstream p53 oscillator, consisting of a feedback loop formed by p53 and its negative regulator, Mdm2. In agreement with experiments, our simulations show that by assuming stochasticity in the initial number of DSBs and the DNA repair process, p53 and Mdm2 exhibit a coordinated oscillatory dynamics upon IR stimulation in single cells, with a stochastic number of oscillations whose mean increases with IR dose. The damped oscillations previously observed in cell populations can be explained as the aggregate behavior of single cell
2020-05-13 | MODEL2005130001 | BioModels
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:RNA-DNA hybrids are a major internal cause of DNA damage within cells, and their degradation by RNAse H enzymes is important for maintaining genomic stability. Here, we identified an unexpected role for RNA-DNA hybrids and RNase H enzymes in DNA repair. Using a site-specific DNA double-stranded break (DSB) system in Schizosaccharomyces pombe, we showed that RNA-DNA hybrids form as part of the homologous recombination (HR)-mediated DSB repair process and RNase H enzymes are essential for their degradation and efficient completion of DNA repair. Deleting RNase H stabilizes RNA-DNA hybrids around DSB sites and strongly impairs recruitment of the ssDNA-binding RPA complex. In contrast, overexpressing RNase H1 destabilizes these hybrids, leading to excessive strand resection and RPA recruitment, and to severe loss of repeat regions around DSBs. Our study challenges the existing model of HR-mediated DSB repair, and reveals a surprising role for RNA-DNA hybrids in maintaining genomic stability.
2016-10-28 | GSE84882 | GEO
Project description:RNA-DNA hybrids are a major internal cause of DNA damage within cells, and their degradation by RNAse H enzymes is important for maintaining genomic stability. Here, we identified an unexpected role for RNA-DNA hybrids and RNase H enzymes in DNA repair. Using a site-specific DNA double-stranded break (DSB) system in Schizosaccharomyces pombe, we showed that RNA-DNA hybrids form as part of the homologous recombination (HR)-mediated DSB repair process and RNase H enzymes are essential for their degradation and efficient completion of DNA repair. Deleting RNase H stabilizes RNA-DNA hybrids around DSB sites and strongly impairs recruitment of the ssDNA-binding RPA complex. In contrast, overexpressing RNase H1 destabilizes these hybrids, leading to excessive strand resection and RPA recruitment, and to severe loss of repeat regions around DSBs. Our study challenges the existing model of HR-mediated DSB repair, and reveals a surprising role for RNA-DNA hybrids in maintaining genomic stability.
2016-10-28 | GSE80398 | GEO
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. Drosophila melanogaster S2 cells were cultured and transfected with reporter gene plasmids that were either circular or modified by restriction digest prior to transfection. Following transfection, total RNA was isolated from the cells and gel-purified for size selection (~18-30 nt). Digested plasmid samples were compared to those of circular plasmids and a nontransfected control.
2012-08-03 | E-GEOD-38967 | biostudies-arrayexpress
Project description:Homologous recombination (HR) is crucial for genetic exchange, accurate repair of DNA double-strand breaks and pivotal for genome integrity. HR uses homologous sequences for repair, but how homology search, the exploration of the genome for homologous DNA sequences, is conducted in the nucleus remains poorly understood. Here, we use time-resolved chromatin immunoprecipitations of repair proteins to monitor homology search in vivo. We found that homology search proceeds by a probing mechanism, which commences around the break and samples preferentially on the broken chromosome. However, elements thought to instruct chromosome loops mediate homology search shortcuts, and centromeres, which cluster within the nucleus, may facilitate homology search on other chromosomes. Our study thus revealed crucial parameters for homology search in vivo and emphasizes the importance of linear distance, chromosome architecture and proximity for recombination efficiency.
2013-03-21 | GSE44844 | GEO
Project description:Chromatin relaxation is a prerequisite step that allows the DNA repair machinery to access double-strand breaks (DSBs), and local histones around the DNA breaks suffer from prompt acetylation changes. However, an intriguing question remains as to where the large demands of acetyl-CoA is precisely produced promptly. Here, we report that pyruvate dehydrogenase 1α (PDHE1α) catalyzes pyruvate metabolism to provide acetyl-CoA promptly in response to DNA damage. PDHE1α was quickly recruited to chromatin in a PARylation-dependent manner, which further drove acetyl-CoA generation to support local chromatin acetylation around the DSB regions. In turn, this process increased the formation of relaxed chromatin to benefit repair factor loading, thus promoting DSB repair to ultimately maintain genome stability and contribute to the resistance of cancer cells to DNA-damaging treatments in vitro and in vivo. In accordance with this, blocking PARylation-based PDHE1α chromatin recruitment markedly attenuated chromatin relaxation and the DSB repair efficiency, resulting in genome instability and restoration of radiosensitivity. Collectively, these findings reveal an undescribed mechanism that underlies site-directed acetyl-CoA generation involving chromatin-associated PDHE1α and its instrumental function in DNA repair and regulating local chromatin acetylation around DSB sites.
2023-07-27 | PXD040676 | Pride