Dissection of a CTCF insulator and its impact on chromosome topology and oncogene expression in gastrointestinal stromal tumor (RNA-Seq)
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE241926 | GEO | 2024/03/06
REPOSITORIES: GEO
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Project description:Enhancer-Promoter communication is dependent on the topological organization of genomic DNA, including topologically-associated domains (TADs) and boundaries enforced by the CTCF insulator protein. Here, we characterized the structure and function of the FGF3/4 locus and the boundary element that insulates the region from an enhancer in the adjacent TAD.
2024-03-06 | GSE241925 | GEO
Project description:Enhancer-Promoter communication is dependent on the topological organization of genomic DNA, including topologically-associated domains (TADs) and boundaries enforced by the CTCF insulator protein. Here, we characterized the structure and function of the FGF3/4 locus and the boundary element that insulates the region from an enhancer in the adjacent TAD.
2024-03-06 | GSE241924 | GEO
Project description:Gain-of-function IDH mutations define major clinical and prognostic classes of gliomas. Mutant IDH protein produces a novel onco-metabolite, 2-hydroxyglutarate (2-HG), that interferes with iron-dependent hydroxylase enzymes, including the TET family of 5'-methylcytosine hydroxylases. TET enzymes are critical for the dynamic regulation of DNA methylation. IDH mutant gliomas thus manifest a CpG island methylator phenotype (G-CIMP), though the functional significance of this altered epigenetic state remains unclear. Here we show that IDH1 mutant gliomas exhibit hyper-methylation at CTCF binding sites, leading to reduced binding of this methylation-sensitive insulator protein. Loss of CTCF binding is associated with a loss of insulation between topological domains and aberrant gene activation. We specifically demonstrate that loss of CTCF at a domain boundary permits a constitutive enhancer to aberrantly interact with the receptor tyrosine kinase gene PDGFRA, a prominent glioma oncogene. Treatment of IDH mutant gliomaspheres with demethylating agent partially restores insulator function and reduces PDGFRA expression. Conversely, CRISPR-mediated disruption of the CTCF binding sequence in IDH wildtype gliomaspheres induces PDGFRA expression and increases proliferation. Our study suggests that IDH mutations promote gliomagenesis by disrupting chromosomal topology and allowing aberrant regulatory interactions that induce oncogene expression. CTCF occupancy characterization and histone H3K27 acetylation profiling in IDH1 mutant and wild-type glioma patient specimens and culture models. ChIP-seq raw data is to be made available through dbGaP (controlled access) due to patient privacy concerns.
2015-12-23 | E-GEOD-70991 | biostudies-arrayexpress
Project description:Genome organization is driven by forces affecting transcriptional state, but the relationship between transcription and genome architecture remains unclear. Here, we identified the Drosophila transcription factor Motif 1 Binding Protein (M1BP) in physical association with the gypsy chromatin insulator core complex, including the universal insulator protein CP190. M1BP is required for enhancer-blocking and barrier activities of the gypsy insulator as well as its proper nuclear localization. Genome-wide, M1BP specifically colocalizes with CP190 at Motif 1-containing promoters, which are enriched at topologically associating domain (TAD) borders. M1BP facilitates CP190 chromatin binding at many shared sites and vice versa. Both factors promote Motif 1-dependent gene expression and transcription near TAD borders genome-wide. Finally, loss of M1BP reduces chromatin accessibility and increases both inter- and intra-TAD local genome compaction. Our results reveal physical and functional interaction between CP190 and M1BP to activate transcription at TAD borders and mediate chromatin insulator-dependent genome organization.
2021-06-07 | PXD026497 | Pride
Project description:Gain-of-function IDH mutations define major clinical and prognostic classes of gliomas. Mutant IDH protein produces a novel onco-metabolite, 2-hydroxyglutarate (2-HG), that interferes with iron-dependent hydroxylase enzymes, including the TET family of 5'-methylcytosine hydroxylases. TET enzymes are critical for the dynamic regulation of DNA methylation. IDH mutant gliomas thus manifest a CpG island methylator phenotype (G-CIMP), though the functional significance of this altered epigenetic state remains unclear. Here we show that IDH1 mutant gliomas exhibit hyper-methylation at CTCF binding sites, leading to reduced binding of this methylation-sensitive insulator protein. Loss of CTCF binding is associated with a loss of insulation between topological domains and aberrant gene activation. We specifically demonstrate that loss of CTCF at a domain boundary permits a constitutive enhancer to aberrantly interact with the receptor tyrosine kinase gene PDGFRA, a prominent glioma oncogene. Treatment of IDH mutant gliomaspheres with demethylating agent partially restores insulator function and reduces PDGFRA expression. Conversely, CRISPR-mediated disruption of the CTCF binding sequence in IDH wildtype gliomaspheres induces PDGFRA expression and increases proliferation. Our study suggests that IDH mutations promote gliomagenesis by disrupting chromosomal topology and allowing aberrant regulatory interactions that induce oncogene expression.
2015-12-23 | GSE70991 | GEO
Project description:<p>Metabolic lesions with pleiotropic effects on epigenetic regulation and other cellular processes are widely implicated in cancer, yet their oncogenic mechanisms remain poorly understood. Succinate dehydrogenase (SDH) deficiency causes a subset of gastrointestinal stromal tumors (GISTs) with DNA hyper-methylation. Here we associate this hyper-methylation with changes in chromosome topology that activate oncogenic programs. To investigate epigenetic alterations in this disease, we systematically mapped DNA methylation, CTCF insulators, enhancers and chromosome topology in KIT-mutant, PDGFRA-mutant and SDH-deficient GISTs. Although these respective subtypes share similar enhancer landscapes, we identified hundreds of putative insulators where DNA methylation replaced CTCF binding in SDH-deficient GISTs. We focused on disrupted insulators that partitions super-enhancers from FGF3, FGF4 and the KIT oncogene. Recurrent loss of this insulator alters locus topology in SDH-deficient GISTs, allowing aberrant physical interaction between enhancers and oncogenes. CRISPR-mediated excision of the corresponding CTCF motif in an SDH-intact model disrupted the boundary and up-regulated FGFs and KIT expression. Our findings reveal how a metabolic lesion destabilizes chromatin structure to facilitate the initiation and selection of epigenetic alterations that drive oncogenic programs in the absence of canonical mutations.</p>
| phs001906 | dbGaP
Project description:Chromatin insulators organize the genome into distinct transcriptional domains and contribute to cell type-specific chromatin organization. However, factors regulating tissue-specific insulator function have not yet been discovered. Here we identify the RNA recognition motif-containing protein, Shep, as a direct interactor of two individual components of the gypsy insulator complex in Drosophila. Mutation of shep improves gypsy-dependent enhancer blocking, indicating a role as a negative regulator of insulator activity. Unlike ubiquitously expressed core gypsy insulator proteins, Shep is highly expressed in the central nervous system (CNS) with lower expression in other tissues. We developed a novel, quantitative tissue-specific barrier assay to demonstrate that Shep functions as a negative regulator of insulator activity in the CNS but not in muscle tissue. Additionally, mutation of shep alters insulator complex nuclear localization in the CNS but not other tissues. Consistent with negative regulatory activity, ChIP-seq analysis of Shep in a CNS-derived cell line indicates substantial genome-wide colocalization with a single gypsy insulator component but limited overlap with intact insulator complexes. Taken together, these data reveal a novel, tissue-specific mode of regulation of a chromatin insulator. ChIP-seq of Shep, Su(Hw), and Mod(mdg4)2.2 in Drosophila BG3 cells along with alternate antibodies
2012-11-30 | E-GEOD-40797 | biostudies-arrayexpress
Project description:Chromatin insulators are DNA-protein complexes that can prevent the spread of repressive chromatin and block communication between enhancers and promoters to regulate gene expression. In Drosophila, the gypsy chromatin insulator complex consists of three core proteins: CP190, Su(Hw), and Mod(mdg4)67.2. These factors concentrate at nuclear foci termed insulator bodies, and their normal localization is correlated with proper insulator function. Here, we identified NURF301/E(bx), a nucleosome remodeling factor, as a novel regulator of gypsy insulator body localization through a high-throughput RNAi imaging screen. NURF301 promotes gypsy-dependent insulator barrier activity and physically interacts with gypsy insulator proteins. Using ChIP-seq, we found that NURF301 co-localizes with insulator proteins genome-wide, and NURF301 promotes chromatin association of Su(Hw) and CP190 at gypsy insulator binding sites. These effects correlate with NURF301-dependent nucleosome repositioning. At the same time, CP190 and Su(Hw) are also required for recruitment of NURF301 to chromatin. Finally, Oligopaint FISH combined with immunofluorescence revealed that NURF301 promotes 3D contact between insulator bodies and gypsy binding site DNA, and NURF301 is required for proper nuclear positioning of gypsy binding sites. Our data provide new insights into how a nucleosome remodeling factor and insulator proteins cooperatively contribute to nuclear organization.
2022-10-14 | PXD032165 | Pride
Project description:Chromatin insulators organize the genome into distinct transcriptional domains and contribute to cell type-specific chromatin organization. However, factors regulating tissue-specific insulator function have not yet been discovered. Here we identify the RNA recognition motif-containing protein, Shep, as a direct interactor of two individual components of the gypsy insulator complex in Drosophila. Mutation of shep improves gypsy-dependent enhancer blocking, indicating a role as a negative regulator of insulator activity. Unlike ubiquitously expressed core gypsy insulator proteins, Shep is highly expressed in the central nervous system (CNS) with lower expression in other tissues. We developed a novel, quantitative tissue-specific barrier assay to demonstrate that Shep functions as a negative regulator of insulator activity in the CNS but not in muscle tissue. Additionally, mutation of shep alters insulator complex nuclear localization in the CNS but not other tissues. Consistent with negative regulatory activity, ChIP-seq analysis of Shep in a CNS-derived cell line indicates substantial genome-wide colocalization with a single gypsy insulator component but limited overlap with intact insulator complexes. Taken together, these data reveal a novel, tissue-specific mode of regulation of a chromatin insulator.
2012-11-30 | GSE40797 | GEO
Project description:Chromatin insulators are DNA-protein complexes situated throughout the genome capable of demarcating independent transcriptional domains. Previous studies point to an important role for RNA in gypsy chromatin insulator function in Drosophila; however, the identity of these putative insulator-associated RNAs is not currently known. Here we utilize RNA-immunoprecipitation and high throughput sequencing (RIP-seq) to isolate RNAs stably associated with gypsy insulator complexes. Strikingly, these RNAs correspond to specific sense-strand, spliced, and polyadenylated mRNAs, including two insulator protein transcripts. In order to assess the functional significance of these associated mRNAs independent of their coding function, we expressed untranslatable versions of these transcripts in developing flies and observed both alteration of insulator complex nuclear localization as well as improvement of enhancer-blocking activity. Together these data suggest a novel, noncoding mechanism by which certain mRNAs contribute to chromatin insulator function. RIP-seq of insulator proteins with different library preparations and multiple biological replicates
2013-08-06 | E-GEOD-29991 | biostudies-arrayexpress