{"lab": {"uuid": "53f8d746-0c48-4151-92a0-2d24ad28cb5a", "display_title": "Christine Disteche, UW", "@type": ["Lab", "Item"], "title": "Christine Disteche, UW", "status": "current", "correspondence": [{"contact_email": "Y2Rpc3RlY2hAdS53YXNoaW5ndG9uLmVkdQ==", "@id": "/users/d89fb49f-f068-43c3-8e3c-6b1c68757a53/", "display_title": "Christine Disteche"}], "@id": "/labs/christine-disteche-lab/", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.lab_submitter", "submits_for.53f8d746-0c48-4151-92a0-2d24ad28cb5a"]}}, "award": {"uuid": "fcc7f634-9252-499f-b79c-380795af2ddd", "center_title": "NOFIC - Shendure", "project": "4DN", "description": "NOFIC: A current grand challenge in genomics involves accurately assaying, at all relevant scales, the 3D conformation of DNA in vivo and then linking conformational changes to dynamic processes such as the cell cycle, differentiation and disease. Here we propose to create the University of Washington Center for Nuclear Organization and Function, bringing together an interdisciplinary team of investigators whose diverse areas of expertise - technology development, computational modeling, and mouse and human biology - make them ideally suited to this challenge. Our overall hypothesis is that characterizing and understanding changes in genome architecture over time (the 4D nucleome) will lead to fundamental insights into human biology and disease. We will address this hypothesis by developing a combination of experimental and computational methods development, coupled with their systematic biological validation and application to development- and disease-relevant systems. On the experimental side, we will further optimize our recently developed DNase Hi- C assay, including combinatorial methods for single cells, ultimately aiming to concurrently assay nuclear architecture and gene expression within each of many single cells. On the computational side, we will extend our existing 3D modeling algorithms to account for diploidy, cell-to-cell variabilit, the hierarchical nature of genome architecture, and to explicitly model architectural changes over cell cycle and cell differentiation time scales. We will then employ several complementary computational methods to link our 4D nucleome models to existing, 1D genomics data sets. The outputs of these new experimental and computational technologies will be subjected to orthogonal validation in several well-understood model systems: human cell lines, in vivo tissues from interspecific F1 hybrid mice, mouse embryonic stem cells (ESCs) and skeletal myoblasts. We will also test specific predictions of the models in response to targeted (genome editing) or large-scale (chromosome silencing) perturbations. After initial validation and in parallel with further methods development, we will apply our new tools to the analysis of three biological systems: we will characterize the dynamics of nuclear architecture during the directed differentiation of na\u00efve human ESCs into cardiomyocytes and endothelial cells; we will test the hypothesis that cardiomyopathy-inducing mutations in the nuclear scaffolding protein, lamin A, are associated with derangements in cardiomyocyte nuclear architecture; and we will determine the changes in human cardiomyocyte nuclear architecture induced by trisomy 21. The proposed center will produce new experimental protocols for ascertaining 4D nucleome architecture, two new software toolkits for modeling the 4D nucleome and linking features of the nucleome to other types of genomic data, a variety of publicly available, large-scale 4D nucleome data sets in mouse and human systems, and fundamental insights into human biology and disease. In all of this work, we will work closely and openly with NOFIC and the 4DN Network to maximize the impact of our center and the overall program.", "@id": "/awards/1U54DK107979-01/", "@type": ["Award", "Item"], "display_title": "UNIVERSITY OF WASHINGTON CENTER FOR NUCLEAR ORGANIZATION AND FUNCTION", "status": "current", "name": "1U54DK107979-01", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "status": "released", "aliases": ["disteche_lab:customset_sci_hic_all"], "dbxrefs": ["GEO:GSE185608"], "accession": "4DNESB7XYI9V", "description": "sci-Hi-C experiments on F121-6, F123 or ES_Tsix-stop mESC cells differentiated to embryoid body (time-course)", "date_created": "2021-09-03T12:24:08.070343+00:00", "submitted_by": {"error": "no view permissions"}, "dataset_label": "sci-Hi-C on mESCs differentiated to embryoid body", "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2024-02-23T15:59:47.455899+00:00"}, "public_release": "2021-09-13", "schema_version": "2", "static_headers": [{"lab": {"@id": "/labs/4dn-dcic-lab/", "display_title": "4DN DCIC, HMS", "@type": ["Lab", "Item"], "status": "current", "uuid": "828cd4fe-ebb0-4b36-a94a-d2e3a36cc989", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.lab_submitter", "submits_for.828cd4fe-ebb0-4b36-a94a-d2e3a36cc989"]}}, "body": "**Sci-Hi-C**\n\n\nSci-Hi-C is a method to detect and quantify chromatin interactions in large numbers of single cells. This massively multiplexed method was developed in 2017 and it enables characterization of 3D genome architecture in thousands of single cells in parallel without requiring compartmentalization of each cell or microfluidic manipulation.\n\nThis protocol involves cross-linking the cells with formaldehyde. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme DpnII is used to fragment the chromatin. Nuclei are then distributed to 96 wells, wherein the first barcode is introduced through ligation of barcoded biotinylated double-stranded bridge adaptors. Intact nuclei are then pooled and subjected to proximity ligation, followed by dilution and redistribution to a second 96-well plate. Importantly, this dilution is carried out such that each well in this second plate contains at most 25 nuclei. Following lysis, a second barcode is introduced through ligation of barcoded Y-adaptors. As the number of barcode combinations (96 \u00d7 96) exceeds the number of nuclei (96 \u00d7 25), the vast majority of single nuclei are tagged by a unique combination of barcodes. All material is once again pooled, and biotinylated junctions are purified with streptavidin beads, restriction digested, and further processed to Illumina sequencing libraries. Sequencing these molecules with relatively long paired-end reads (i.e., 2 \u00d7 250 base pairs) allows one to identify not only the genome-derived fragments of conventional Hi-C, but also external and internal barcodes,  which enable decomposition of the Hi-C data into single-cell contact probability maps.\n\n\nSee [Ramani et al 2017](https://www.nature.com/articles/nmeth.4155) for more details on in sci-Hi-C.\n\n<dl>\n<a href=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" target=\"_blank\">\n<img src=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" style=\"width:1000px\" />  \n</a>\n<br/><br/>\n<em>Image source: Ramani et. al. Nature Methods 2017, Figure 1a </em>\n</dl>", "name": "item-page-headers.ExperimentType.sciHiC", "award": {"status": "current", "uuid": "b0b9c607-f8b4-4f02-93f4-9895b461334b", "display_title": "4D NUCLEOME NETWORK DATA COORDINATION AND INTEGRATION CENTER - PHASE I", "@type": ["Award", "Item"], "@id": "/awards/1U01CA200059-01/", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "title": "Assay Description", "status": "released", "aliases": ["4dn-dcic-lab:experiment_infobox_sciHiC"], "options": {"filetype": "md", "collapsible": false, "default_open": false}, "date_created": "2018-10-10T14:42:45.156523+00:00", "section_type": "Item Page Header", "submitted_by": {"error": "no view permissions"}, "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2019-06-14T14:04:34.975645+00:00"}, "schema_version": "2", "@id": "/static-sections/ae5a6470-0694-4ba3-893a-40b170401bc0/", "@type": ["StaticSection", "UserContent", "Item"], "uuid": "ae5a6470-0694-4ba3-893a-40b170401bc0", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.owner", "userid.56c9c683-bb11-471b-b590-c656f7dc03c1"]}, "display_title": "Assay Description", "external_references": [], "content": "**Sci-Hi-C**\n\n\nSci-Hi-C is a method to detect and quantify chromatin interactions in large numbers of single cells. This massively multiplexed method was developed in 2017 and it enables characterization of 3D genome architecture in thousands of single cells in parallel without requiring compartmentalization of each cell or microfluidic manipulation.\n\nThis protocol involves cross-linking the cells with formaldehyde. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme DpnII is used to fragment the chromatin. Nuclei are then distributed to 96 wells, wherein the first barcode is introduced through ligation of barcoded biotinylated double-stranded bridge adaptors. Intact nuclei are then pooled and subjected to proximity ligation, followed by dilution and redistribution to a second 96-well plate. Importantly, this dilution is carried out such that each well in this second plate contains at most 25 nuclei. Following lysis, a second barcode is introduced through ligation of barcoded Y-adaptors. As the number of barcode combinations (96 \u00d7 96) exceeds the number of nuclei (96 \u00d7 25), the vast majority of single nuclei are tagged by a unique combination of barcodes. All material is once again pooled, and biotinylated junctions are purified with streptavidin beads, restriction digested, and further processed to Illumina sequencing libraries. Sequencing these molecules with relatively long paired-end reads (i.e., 2 \u00d7 250 base pairs) allows one to identify not only the genome-derived fragments of conventional Hi-C, but also external and internal barcodes,  which enable decomposition of the Hi-C data into single-cell contact probability maps.\n\n\nSee [Ramani et al 2017](https://www.nature.com/articles/nmeth.4155) for more details on in sci-Hi-C.\n\n<dl>\n<a href=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" target=\"_blank\">\n<img src=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" style=\"width:1000px\" />  \n</a>\n<br/><br/>\n<em>Image source: Ramani et. al. Nature Methods 2017, Figure 1a </em>\n</dl>", "filetype": "md", "content_as_html": "<div class=\"markdown-container\"><p><strong>Sci-Hi-C</strong></p>\n<p>Sci-Hi-C is a method to detect and quantify chromatin interactions in large numbers of single cells. This massively multiplexed method was developed in 2017 and it enables characterization of 3D genome architecture in thousands of single cells in parallel without requiring compartmentalization of each cell or microfluidic manipulation.</p>\n<p>This protocol involves cross-linking the cells with formaldehyde. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme DpnII is used to fragment the chromatin. Nuclei are then distributed to 96 wells, wherein the first barcode is introduced through ligation of barcoded biotinylated double-stranded bridge adaptors. Intact nuclei are then pooled and subjected to proximity ligation, followed by dilution and redistribution to a second 96-well plate. Importantly, this dilution is carried out such that each well in this second plate contains at most 25 nuclei. Following lysis, a second barcode is introduced through ligation of barcoded Y-adaptors. As the number of barcode combinations (96 \u00d7 96) exceeds the number of nuclei (96 \u00d7 25), the vast majority of single nuclei are tagged by a unique combination of barcodes. All material is once again pooled, and biotinylated junctions are purified with streptavidin beads, restriction digested, and further processed to Illumina sequencing libraries. Sequencing these molecules with relatively long paired-end reads (i.e., 2 \u00d7 250 base pairs) allows one to identify not only the genome-derived fragments of conventional Hi-C, but also external and internal barcodes,  which enable decomposition of the Hi-C data into single-cell contact probability maps.</p>\n<p>See <a href=\"https://www.nature.com/articles/nmeth.4155\" rel=\"noopener noreferrer\" target=\"_blank\">Ramani et al 2017</a> for more details on in sci-Hi-C.</p>\n<dl>\n<a href=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" rel=\"noopener noreferrer\" target=\"_blank\">\n<img src=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/sciHiC.jpg\" style=\"width:1000px\"/>\n</a>\n<br/><br/>\n<em>Image source: Ramani et. al. 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(2021)", "title": "Single-cell landscape of nuclear configuration and gene expression during stem cell differentiation and X inactivation.", "uuid": "59d7080d-7ce9-4ade-b35b-5b8adcc80402", "abstract": "BACKGROUND: Mammalian development is associated with extensive changes in gene expression, chromatin accessibility, and nuclear structure. Here, we follow such  changes associated with mouse embryonic stem cell differentiation and X inactivation by integrating, for the first time, allele-specific data from these  three modalities obtained by high-throughput single-cell RNA-seq, ATAC-seq, and Hi-C. RESULTS: Allele-specific contact decay profiles obtained by single-cell Hi-C clearly show that the inactive X chromosome has a unique profile in differentiated cells that have undergone X inactivation. Loss of this inactive X-specific structure at mitosis is followed by its reappearance during the cell cycle, suggesting a \"bookmark\" mechanism. Differentiation of embryonic stem cells to follow the onset of X inactivation is associated with changes in contact decay profiles that occur in parallel on both the X chromosomes and autosomes. Single-cell RNA-seq and ATAC-seq show evidence of a delay in female versus male cells, due to the presence of two active X chromosomes at early stages of differentiation. The onset of the inactive X-specific structure in single cells occurs later than gene silencing, consistent with the idea that chromatin compaction is a late event of X inactivation. Single-cell Hi-C highlights evidence of discrete changes in nuclear structure characterized by the acquisition of very long-range contacts throughout the nucleus. Novel computational approaches allow for the effective alignment of single-cell gene expression, chromatin accessibility, and 3D chromosome structure. CONCLUSIONS: Based on trajectory analyses, three distinct nuclear structure states are detected reflecting discrete and profound simultaneous changes not only to the structure of the X chromosomes, but also to that of autosomes during differentiation. Our study reveals that long-range structural changes to chromosomes appear as discrete events, unlike progressive changes in gene expression and chromatin accessibility.", "display_title": "Bonora G et al. (2021) PMID:34579774", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "pubs_using": [], "publications_of_set": [{"title": "Single-cell landscape of nuclear configuration and gene expression during stem cell differentiation and X inactivation.", "uuid": "59d7080d-7ce9-4ade-b35b-5b8adcc80402", "display_title": "Bonora G et al. (2021) PMID:34579774", "@type": ["Publication", "Item"], "journal": "Genome biology", "abstract": "BACKGROUND: Mammalian development is associated with extensive changes in gene expression, chromatin accessibility, and nuclear structure. Here, we follow such  changes associated with mouse embryonic stem cell differentiation and X inactivation by integrating, for the first time, allele-specific data from these  three modalities obtained by high-throughput single-cell RNA-seq, ATAC-seq, and Hi-C. RESULTS: Allele-specific contact decay profiles obtained by single-cell Hi-C clearly show that the inactive X chromosome has a unique profile in differentiated cells that have undergone X inactivation. Loss of this inactive X-specific structure at mitosis is followed by its reappearance during the cell cycle, suggesting a \"bookmark\" mechanism. Differentiation of embryonic stem cells to follow the onset of X inactivation is associated with changes in contact decay profiles that occur in parallel on both the X chromosomes and autosomes. Single-cell RNA-seq and ATAC-seq show evidence of a delay in female versus male cells, due to the presence of two active X chromosomes at early stages of differentiation. The onset of the inactive X-specific structure in single cells occurs later than gene silencing, consistent with the idea that chromatin compaction is a late event of X inactivation. Single-cell Hi-C highlights evidence of discrete changes in nuclear structure characterized by the acquisition of very long-range contacts throughout the nucleus. Novel computational approaches allow for the effective alignment of single-cell gene expression, chromatin accessibility, and 3D chromosome structure. CONCLUSIONS: Based on trajectory analyses, three distinct nuclear structure states are detected reflecting discrete and profound simultaneous changes not only to the structure of the X chromosomes, but also to that of autosomes during differentiation. Our study reveals that long-range structural changes to chromosomes appear as discrete events, unlike progressive changes in gene expression and chromatin accessibility.", "ID": "PMID:34579774", "@id": "/publications/59d7080d-7ce9-4ade-b35b-5b8adcc80402/", "status": "current", "authors": ["Bonora G", "Ramani V", "Singh R", "Fang H", "Jackson DL", "Srivatsan S", "Qiu R", "Lee C", "Trapnell C", "Shendure J", "Duan Z", "Deng X", "Noble WS", "Disteche CM"], "date_published": "2021-09-27", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}], "number_of_experiments": 23, "@context": "/terms/", "aggregated-items": {"badges": [{"parent": "/biosamples/4DNBSREAXXIJ/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing morphology_image", "Biosample is a stem cell line over 10 passages but missing karyotype"], "badge": {"commendation": null, "warning": "Biosample Metadata Incomplete", "uuid": 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