Differential Contribution of Steady-State RNA and Active Transcription in Chromatin Organization
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE114337 | GEO | 2019/04/01
REPOSITORIES: GEO
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Project description:Higher order chromatin structure establishes domains that organize the genome and coordinate gene expression. However, the molecular mechanisms controlling transcription of individual loci within a topological domain (TAD) are not fully understood. The cystic fibrosis transmembrane conductance regulator (CFTR) gene provides a paradigm for investigating these mechanisms. CFTR occupies a TAD bordered by CTCF/cohesin binding sites within which are cell-type-selective cis-regulatory elements for the locus. We showed previously that intronic and extragenic enhancers, when occupied by specific transcription factors, are recruited to the CFTR promoter by a looping mechanism to drive gene expression. Here we use a combination of CRISPR/Cas9 editing of cis-regulatory elements and siRNA-mediated depletion of architectural proteins to determine the relative contribution of structural elements and enhancers to the higher order structure and expression of the CFTR locus. We found the boundaries of the CFTR TAD are conserved among diverse cell types and are dependent on CTCF and cohesin complex. Removal of an upstream CTCF-binding insulator alters the interaction profile, but has little effect on CFTR expression. Within the TAD, intronic enhancers recruit cell-type selective transcription factors and deletion of a pivotal enhancer element dramatically decreases CFTR expression, but has minor effect on its 3D structure. Examination of open chromatin region in Caco2 (colorectal adenocarcinoma cells), HBE (primary human bronchial epithelial cells), and primary adult human epididymis cells
2016-01-07 | E-GEOD-74709 | biostudies-arrayexpress
Project description:The packaging of DNA into chromatin plays an important role in transcriptional regulation and nuclear processes. Brahma related gene-1 SMARCA4 (also known as BRG1), the essential ATPase subunit of the mammalian SWI/SNF chromatin remodeling complex, uses the energy from ATP hydrolysis to disrupt nucleosomes at target regions. Although the transcriptional role of SMARCA4 at gene promoters is well-studied, less is known about its role in higher-order genome organization. SMARCA4 knockdown in human mammary epithelial MCF-10A cells resulted in 176 up-regulated genes, including many related to lipid and calcium metabolism, and 1292 down-regulated genes, some of which encode extracellular matrix (ECM) components that can exert mechanical forces and affect nuclear structure. ChIP-seq analysis of SMARCA4 localization and SMARCA4-bound super-enhancers demonstrated extensive binding at intergenic regions. Furthermore, Hi-C analysis showed extensive SMARCA4-mediated alterations in higher-order genome organization at multiple resolutions. First, SMARCA4 knockdown resulted in clustering of intra- and inter- sub-telomeric regions, demonstrating a novel role for SMARCA4 in telomere organization. SMARCA4 binding was enriched at TAD (Topologically Associating Domain) boundaries, and SMARCA4 knockdown resulted in weakening of TAD boundary strength. Taken together, these findings provide a dynamic view of SMARCA4-dependent changes in higher-order chromatin organization and gene expression, identifying SMARCA4 as a novel component of chromatin organization. Hi-C and RNA-seq experiments were conducted in MCF-10A shSCRAM and shSMARCA4 cells. SMARCA4 ChIP-seq was conducted in wildtype MCF-10A cells.
2016-06-02 | E-GEOD-74716 | biostudies-arrayexpress
Project description:The spatial arrangement of interphase chromosomes in the nucleus is important for gene expression and genome function in animals and in plants. The recently developed Hi-C technology is an efficacious method to investigate genome packing. Here we present a detailed Hi-C map of the three-dimensional genome organization of the plant Arabidopsis thaliana. We find that local chromatin packing differs from the patterns seen in animals, with kilobasepair-sized segments that have much higher intra-chromosome interaction rates than neighboring regions and which represent a dominant local structural feature of genome conformation in A. thaliana. These regions appear as positive strips on two-dimensional representations of chromatin interaction and they are enriched in epigenetic marks H3K27me3, H3.1 and H3.3. We also identify over 400 insulator-like regions. Furthermore, although topologically associating domains (TADs), which are prominent in animals, are not the dominant feature of A. thaliana genome packing, we found over 1,000 regions that have properties of TAD boundaries, and a similar number of regions similar to the interior of TADs. These insulator-like, TAD-boundary-like, and TAD-interior-like regions show strong enrichment for distinct epigenetic marks, and correlate with gene transcription levels. We conclude that epigenetic modifications, gene density, and transcriptional activity all contribute to shaping the local structure of the A. thaliana nuclear genome.
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Project description:Higher order chromatin structure establishes domains that organize the genome and coordinate gene expression. However, the molecular mechanisms controlling transcription of individual loci within a topological domain (TAD) are not fully understood. The cystic fibrosis transmembrane conductance regulator (CFTR) gene provides a paradigm for investigating these mechanisms. CFTR occupies a TAD bordered by CTCF/cohesin binding sites within which are cell-type-selective cis-regulatory elements for the locus. We showed previously that intronic and extragenic enhancers, when occupied by specific transcription factors, are recruited to the CFTR promoter by a looping mechanism to drive gene expression. Here we use a combination of CRISPR/Cas9 editing of cis-regulatory elements and siRNA-mediated depletion of architectural proteins to determine the relative contribution of structural elements and enhancers to the higher order structure and expression of the CFTR locus. We found the boundaries of the CFTR TAD are conserved among diverse cell types and are dependent on CTCF and cohesin complex. Removal of an upstream CTCF-binding insulator alters the interaction profile, but has little effect on CFTR expression. Within the TAD, intronic enhancers recruit cell-type selective transcription factors and deletion of a pivotal enhancer element dramatically decreases CFTR expression, but has minor effect on its 3D structure.
2016-01-07 | GSE74709 | GEO
Project description:Recent advances enabled by Hi-C technique have unraveled principles of chromosomal folding, which were since linked to many genomic processes. In particular, Hi-C revealed that chromosomes of animals are organized into Topologically Associating Domains (TADs), evolutionary conserved compact chromatin domains that influence gene expression. However, mechanisms that underlie partitioning of the genome into TADs remain poorly understood. To explore principals of TAD folding in Drosophila melanogaster, we performed Hi-C and PolyA+ RNA-seq in four cell lines of various origins (S2, Kc167, DmBG3-c2, and OSC). Contrary to previous studies, we find that regions between TADs (i.e. the inter-TADs and TAD boundaries) in Drosophila are only weakly enriched with the insulator protein dCTCF, while another insulator protein Su(Hw) is preferentially present within TADs. However, Drosophila inter-TADs harbor active chromatin and constitutively transcribed (housekeeping) genes. Accordingly, we find that binding of insulator proteins dCTCF and Su(Hw) predict TAD boundaries much worse than the active chromatin marks (in the minimal case, H3K4me3 and total RNA) do. Moreover, inter-TADs correspond to decompacted interbands of polytene chromosomes, whereas TADs mostly correspond to densely packed bands. Collectively, our results suggest that TADs are condensed chromatin domains depleted in active chromatin marks, separated by regions of active chromatin that cannot be organized into compact structures, possibly due to high levels of histone acetylation. Finally, we test this hypothesis by polymer simulations, and find that TAD partitioning can be explained by different modes of inter-nucleosomal interactions for active and inactive chromatin. Hi-C experiments, PolyA+ RNA profiling and mapping of chromosomal rearrangements in four Drosophila melanogaster cell lines.
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Project description:Pervasive RNA Binding Protein Enrichment on TAD Boundaries Regulates TAD Organization [Hi-C]
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Project description:Topological associating domains (TADs) are self-interacting genomic units crucial for shaping gene regulation patterns. Despite their importance, the extent of their evolutionary conservation and its functional implications remain largely unknown. In this study, we generate Hi-C and ChIP-seq data and compare TAD organization across four primate and four rodent species, and characterize the genetic and epigenetic properties of TAD boundaries in correspondence to their evolutionary conservation. We find 14% of all human TAD boundaries to be shared among all eight species (ultraconserved), while 15% are human-specific. Ultraconserved TAD boundaries have stronger insulation strength, CTCF binding, and enrichment of older retrotransposons, compared to species-specific boundaries. CRISPR-Cas9 knockouts of two ultraconserved boundaries in mouse models leads to tissue-specific gene expression changes and morphological phenotypes. Deletion of a human-specific boundary near the autism-related AUTS2 gene results in upregulation of this gene in neurons. Overall, our study provides pertinent TAD boundary evolutionary conservation annotations, and showcase the functional importance of TAD evolution.
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Project description:The relationship between evolutionary genome remodeling and the three-dimensional structure of the genome remain largely unexplored. Here we use the heavily rearranged gibbon genome to examine how evolutionary chromosomal rearrangements impact genome-wide chromatin interactions, topologically associating domains (TADs), and their epigenetic landscape. We use high-resolution maps of gibbon-human breaks of synteny (BOS), apply Hi-C in gibbon, measure an array of epigenetic features, and perform cross-species comparisons. We find that gibbon rearrangements occur at TAD boundaries, independent of the parameters used to identify TADs. This overlap is supported by a remarkable genetic and epigenetic similarity between BOS and TAD boundaries, namely presence of CpG islands and SINE elements, and enrichment in CTCF and H3K4me3 binding. Cross-species comparisons reveal that regions orthologous to BOS also correspond with boundaries of large (400-600kb) TADs in human and other mammalian species. The co-localization of rearrangement breakpoints and TAD boundaries may be due to higher chromatin fragility at these locations and/or increased selective pressure against rearrangements that disrupt TAD integrity. We also examine the small portion of BOS that did not overlap with TAD boundaries and gave rise to novel TADs in the gibbon genome. We postulate that these new TADs generally lack deleterious consequences. Lastly, we show that limited epigenetic homogenization occurs across breakpoints, irrespective of their time of occurrence in the gibbon lineage. Overall, our findings demonstrate remarkable conservation of chromatin interactions and epigenetic landscape in gibbons, in spite of extensive genomic shuffling.
2018-05-30 | GSE115064 | GEO
Project description:The relationship between evolutionary genome remodeling and the three-dimensional structure of the genome remain largely unexplored. Here we use the heavily rearranged gibbon genome to examine how evolutionary chromosomal rearrangements impact genome-wide chromatin interactions, topologically associating domains (TADs), and their epigenetic landscape. We use high-resolution maps of gibbon-human breaks of synteny (BOS), apply Hi-C in gibbon, measure an array of epigenetic features, and perform cross-species comparisons. We find that gibbon rearrangements occur at TAD boundaries, independent of the parameters used to identify TADs. This overlap is supported by a remarkable genetic and epigenetic similarity between BOS and TAD boundaries, namely presence of CpG islands and SINE elements, and enrichment in CTCF and H3K4me3 binding. Cross-species comparisons reveal that regions orthologous to BOS also correspond with boundaries of large (400-600kb) TADs in human and other mammalian species. The co-localization of rearrangement breakpoints and TAD boundaries may be due to higher chromatin fragility at these locations and/or increased selective pressure against rearrangements that disrupt TAD integrity. We also examine the small portion of BOS that did not overlap with TAD boundaries and gave rise to novel TADs in the gibbon genome. We postulate that these new TADs generally lack deleterious consequences. Lastly, we show that limited epigenetic homogenization occurs across breakpoints, irrespective of their time of occurrence in the gibbon lineage. Overall, our findings demonstrate remarkable conservation of chromatin interactions and epigenetic landscape in gibbons, in spite of extensive genomic shuffling.
2018-05-30 | GSE115063 | GEO
Project description:The relationship between evolutionary genome remodeling and the three-dimensional structure of the genome remain largely unexplored. Here we use the heavily rearranged gibbon genome to examine how evolutionary chromosomal rearrangements impact genome-wide chromatin interactions, topologically associating domains (TADs), and their epigenetic landscape. We use high-resolution maps of gibbon-human breaks of synteny (BOS), apply Hi-C in gibbon, measure an array of epigenetic features, and perform cross-species comparisons. We find that gibbon rearrangements occur at TAD boundaries, independent of the parameters used to identify TADs. This overlap is supported by a remarkable genetic and epigenetic similarity between BOS and TAD boundaries, namely presence of CpG islands and SINE elements, and enrichment in CTCF and H3K4me3 binding. Cross-species comparisons reveal that regions orthologous to BOS also correspond with boundaries of large (400-600kb) TADs in human and other mammalian species. The co-localization of rearrangement breakpoints and TAD boundaries may be due to higher chromatin fragility at these locations and/or increased selective pressure against rearrangements that disrupt TAD integrity. We also examine the small portion of BOS that did not overlap with TAD boundaries and gave rise to novel TADs in the gibbon genome. We postulate that these new TADs generally lack deleterious consequences. Lastly, we show that limited epigenetic homogenization occurs across breakpoints, irrespective of their time of occurrence in the gibbon lineage. Overall, our findings demonstrate remarkable conservation of chromatin interactions and epigenetic landscape in gibbons, in spite of extensive genomic shuffling.
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