Ontology highlight
ABSTRACT:
PROVIDER: PRJNA881334 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Other | ||||
| SRR21607735.fastq.gz | Fastqsanger.gz | |||
| SRR21607736.fastq.gz | Fastqsanger.gz | |||
| SRR21607737.fastq.gz | Fastqsanger.gz | |||
| SRR21607738.fastq.gz | Fastqsanger.gz |
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Project description:DNA polymerase theta (Pol-theta)-mediated end-joining (TMEJ) repairs DNA double-strand breaks and confers resistance to genotoxic agents. How Pol-theta is regulated at the molecular level to exert TMEJ remains unknown. We find that Pol-theta interacts with and is PARylated by PARP1 in vitro and in cells. PARP1 recruits Pol-theta to the vicinity of DNA damage via PARylation dependent liquid demixing, however, PARylated Pol-theta (PAR-Pol-theta) cannot perform TMEJ due to its inability to bind DNA. PARG-mediated de-PARylation of Pol-theta reactivates its DNA binding and end-joining activities in vitro. Consistent with this, PARG is essential for TMEJ in cells and the temporal recruitment of PARG to DNA damage corresponds with TMEJ activation and dissipation of PARP1 and PAR. These studies support a two-step spatiotemporal mechanism of TMEJ regulation. First, PARP1 PARylates Pol-theta and facilitates its recruitment to DNA damage sites in an inactivated state. PARG subsequently activates TMEJ by removing repressive PAR marks on Pol-theta.
| MSV000092839 | MassIVE
Project description:In homologous recombination deficient cells, polymerase theta (Pol θ)-mediated end-joining (TMEJ) repairs replicative DNA breaks during mitosis through interactions with RHINO and TOPBP1. V(D)J recombination and class switch recombination (CSR) rely on DNA breakage in G1-phase B lymphocytes and repair by non-homologous end-joining (NHEJ). In this study, we assayed CSR in primary B cells deficient in XRCC4, SHLD1, and Pol θ. We found that B cells deficient in both XRCC4 and Pol θ, or SHLD1 and Pol θ, exhibit an increased rate of IgH breaks, while retaining CSR levels similar to those of B cells deficient in XRCC4 or SHLD1. We show that in XRCC4- or SHLD1-deficient B cells, Pol θ promotes the formation of aberrant recombination products characterized by exacerbated double-strand break (DSB) end resection, inversion and microhomology, and that these end-joining events occur independently of RHINO. Furthermore, we find that TMEJ of RAG-generated DSBs is also independent of RHINO and concomitant to S/G2 entry. Based on these results, we propose that in the absence of NHEJ, TMEJ repairs persistent G1-phase DSBs in S/G2, rather than in mitosis.
2025-08-26 | GSE306291 | GEO
Project description:Copy number variation (CNV) is a form of genetic alteration that has been strongly implicated in the pathogenesis of a wide range of neurological disorders. Replication stress is a common cause of CNV. Despite the prevailing model that CNV arises from DNA double strand breaks (DSBs), there has been no assay that directly perturbs presumed DSB sources and measures CNV output. In this article, we identified a subset of recurrent DNA break clusters (RDC) as a causal factor for CNV formation. In murine neural progenitor cells under replication stress, mapping the formation of CNVs revealed their exclusive location in RDC regions that contain actively transcribed genes. CRISPR/Cas9-mediated transcriptional suppression demonstrated that CNV formation depends on transcription-mediated RDCs, but is independent of elevated replication timing. We found that polymerase theta (Pol θ), a protector against CNV formation, plays a critical but context dependent role upstream of RDC. Chemically inhibiting the polymerase activity of Pol θ reduced end filling and micro-homology-mediated joining in XRCC4/P53-deficient cells. Conversely, Pol θ inhibition led to elevated DSB density at RDC-containing loci in wild-type neural stem and progenitor cells, suggesting its role in preventing transcription-replication conflict. These results underscore the role of RDCs in driving genome heterogeneity, and suggest their potential involvement in the pathogenesis of various conditions including neuropsychiatric disorders and cancer.
2026-03-03 | GSE305346 | GEO
Project description:Copy number variation (CNV) is a form of genetic alteration that has been strongly implicated in the pathogenesis of a wide range of neurological disorders. Replication stress is a common cause of CNV. Despite the prevailing model that CNV arises from DNA double strand breaks (DSBs), there has been no assay that directly perturbs presumed DSB sources and measures CNV output. In this article, we identified a subset of recurrent DNA break clusters (RDC) as a causal factor for CNV formation. In murine neural progenitor cells under replication stress, mapping the formation of CNVs revealed their exclusive location in RDC regions that contain actively transcribed genes. CRISPR/Cas9-mediated transcriptional suppression demonstrated that CNV formation depends on transcription-mediated RDCs, but is independent of elevated replication timing. We found that polymerase theta (Pol θ), a protector against CNV formation, plays a critical but context dependent role upstream of RDC. Chemically inhibiting the polymerase activity of Pol θ reduced end filling and micro-homology-mediated joining in XRCC4/P53-deficient cells. Conversely, Pol θ inhibition led to elevated DSB density at RDC-containing loci in wild-type neural stem and progenitor cells, suggesting its role in preventing transcription-replication conflict. These results underscore the role of RDCs in driving genome heterogeneity, and suggest their potential involvement in the pathogenesis of various conditions including neuropsychiatric disorders and cancer.
2026-03-03 | GSE305347 | GEO
Project description:Genome/chromosome organization is highly ordered and controls nuclear events. Here, we show that the TATA box-binding protein (TBP) interacts with the Cnd2 kleisin subunit of condensin to mediate interphase and mitotic chromosome organization in fission yeast. TBP recruits condensin onto RNA polymerase III-transcribed (Pol III) genes and highly transcribed Pol II genes; condensin in turn associates these genes with centromeres. Inhibition of the Cnd2-TBP interaction disrupts condensin localization across the genome and the proper assembly of mitotic chromosomes, leading to severe defects in chromosome segregation and eventually causing cellular lethality. We propose that the Cnd2-TBP interaction coordinates transcription with chromosomal architecture by linking dispersed gene loci with centromeres. This chromosome arrangement can contribute to the efficient transmission of physical force at the kinetochore to chromosomal arms, thereby supporting the fidelity of chromosome segregation. Genome-wide distributions of condensin and Pol III factors in fission yeast.
2015-08-12 | E-GEOD-60273 | biostudies-arrayexpress
Project description:Genetic separation of Brca1 functions reveal mutation-dependent Pol theta vulnerabilities
| PRJNA1025027 | ENA
Project description:Adaptive immune responses to infection result in the formation of memory T and B cells that respond more rapidly and robustly to reinfections, providing the basis of the immunological memory targeted by vaccines. Underlying the enhanced responsiveness of memory cells is their ability to rapidly up-regulate the transcription of key effector genes at a higher level compared to naïve cells (termed transcriptional memory). While transcriptionally permissive histone modifications are known to provide chromatin structures that facilitate transcriptional memory, the molecular mechanisms that underpin this process still remain elusive. The role of the cytoplasmic signalling kinase, PKC-θ in T cell signalling cascades is well established, however PKC-θ has also recently been described as a chromatin-modifying enzyme. We have previously demonstrated that PKC-θ forms a part of an active transcription complex that docks at the IL2 gene - a gene which displays transcriptional memory. In this study, we perturbed PKC-θ expression using approaches such as siRNA knockdown in primary human naïve and memory CD4+ T cells to show that transcriptional memory is highly dependent on PKC-θ, particularly in memory CD4+ T cells. Chromatin-tethered PKC-θ was immuno-precipitated with high levels of the activating marks, H3K4me3 and H3K9ac and low levels of the repressive mark, H3K27me3 at the IL2 promoter in ex vivo-derived, activated memory CD4+ T cells. Further ChIP-sequencing analysis revealed that PKC-θ was prevalent at transcriptionally permissive domains such as promoters, upstream regions and exons but also at intronic regions containing putative enhancers annotated to a subset of genes preferentially expressed in memory CD4+ T cells. Collectively, these data argue that chromatin-tethered PKC-θ can directly regulate genes to establish and maintain a permissive chromatin state that facilitates transcriptional memory in human CD4+ T cells.
2016-05-01 | GSE61241 | GEO
Project description:DNA polymerase iota (Pol ι) is a Y-family lesion bypass polymerases, characterized by its remarkably low fidelity. While Pol ι is ubiquitously expressed in nearly all cell types, its function remains largely undefined. Previous studies in cellular and mouse models deficient in Pol ι have revealed no discernible phenotype. In this report, we show that Pol ι plays an important role in mitotic DNA synthesis. Cells lacking Pol ι exhibit heightened vulnerability to replication stress, increased formation of micronuclei, and accumulation of chromosomal breaks. The protein level of Pol ι is tightly regulated and restricted to G2/M phase cells via CRL7/FBXW11-mediated proteasomal degradation. The absence of Pol ι during S phase ensures that its error-prone DNA synthesis activity is excluded from interfering with normal DNA replication. These findings suggest that Pol ι function is critical in resolving DNA damage-induced replication stress prior to the completion of mitosis.
2025-07-01 | PXD065672 |
Project description:The human A-family DNA polymerase θ (Pol q) is a large, multidomain enzyme whose physiological function is still unclear despite its in vitro translesion synthesis capacity in front of DNA damage and its involvement in some features of DNA repair after external stress. Here we present evidence that Pol q holds a novel role in the absence of external stress as a critical determinant of the replication timing program in human cells. Pol q binds to chromatin at early G1 and is required for proper formation of pre-replicative complexe and replication origin activation. Pol q-depleted cells show modified spatial organization of chromatin-loop structures at replication factories. Genome-wide analysis of replication timing shows delayed replication of a part of early replicating domains and advanced replication of a part of late replicating domains following Pol q depletion. Our results identify Pol q as one of the first critical human factors discovered in the replication timing programme.
2014-10-28 | GSE53070 | GEO