{"lab": {"@id": "/labs/john-lis-lab/", "uuid": "c0bc993a-b7e2-4342-8523-6922c3c4dc35", "display_title": "John Lis, CORNELL", "@type": ["Lab", "Item"], "correspondence": [{"contact_email": "anRsMTBAY29ybmVsbC5lZHU=", "@id": "/users/b7fcb0e6-b184-4bba-966a-0fb4f8364529/", "display_title": "John Lis"}], "status": "current", "title": "John Lis, CORNELL", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.lab_submitter", "submits_for.c0bc993a-b7e2-4342-8523-6922c3c4dc35"]}}, "tags": ["stress_collection"], "award": {"description": "NT: Nuclear organization of DNA is complex and consists of multiple layers. At the lowest resolution, large sections of chromosomes are packed in territories, at a higher level chromosomal regions are organized in topologically-associated domains that provide a framework for interactions of transcription factors (TFs) bound to promoters and distal regulatory elements (enhancers), and finally the structural interplay of regulators, RNA polymerase II, and chromatin then lead to regulated gene expression. In recent years, a number of methods called chromatin (conformation) capture (CC) have been developed and used to capture dynamic and stable DNA contacts that constitute genome architecture and potential regulatory interactions. A major limitation of these methods is that they all depend on a single crosslinker, formaldehyde, which crosslinks DNA to proteins as well as proteins to other proteins. This complicates the interpretation of the observed 'DNA-DNA' contacts and it lacks distance information. Here we propose an orthogonal strategy, named Distance-Hi-C (D-Hi-C), where we design, test, and apply a battery of photo-activated crosslinkers, designed to directly measure distances between interacting sites genome-wide. These bivalent crosslinkers consist of two reactive groups separated by a linker. DNA intercalaters that can be crosslinked by photo-activation (i.e. Psoralen) will be used as the reactive groups. Linkers will be of varying precise lengths, and either flexible or rigid in nature so that the 3-dimensional distance between crosslinked loci can be inferred. Such DNA-specific bivalent crosslinking reagents, when substituted for formaldehyde in Hi-C protocol, produces space constraints revealing enhancer- promoter interactions and potentially allowing the inference of the 3D arrangement of the nuclear genome with unprecedented precision. Moreover, D-Hi-C is expected to lower backgrounds and allow examination of short and moderate range interactions, which are obscured by high backgrounds of current methods. Addition of groups such as digoxigenin to the bivalent crosslinkers, in addition to biotin incorporated to the ligation products, will enable better purification and thus deeper examination of genomic interactions. Finally, sampling DNA distances in time following gene activation provides a means of exploring the 4D architecture and setting critical limits in evaluating mechanisms of gene activation. The specific aims are to 1) synthesize a battery of bivalent crosslinkers and evaluate their ability to crosslink DNA in vitro; 2) test D-Hi-C crosslinkers relative to formaldehyde in the Drosophila nuclei model; 3) apply new crosslinkers to tier 1, ENCODE GM12878 and K562 cell lines to test their efficacy in human cells; and 4) test locus-specific photo-crosslinking that will allow a more focused and thorough examination of locus-specific interactions with the genome in a time course of gene activation. The development of these methodologies will be broadly useful in providing critical insights into gene regulatory mechanisms that are operative in normal animal development and homeostasis and that go awry in diseases like cancer.", "name": "1U01HL129958-01", "@type": ["Award", "Item"], "status": "current", "@id": "/awards/1U01HL129958-01/", "project": "4DN", "center_title": "NT - Lis", "uuid": "d52d2d3e-a491-436f-9150-ee670b1fe3a4", "display_title": "DISTANCE-HI-C: CREATING PHOTO ACTIVATED X-LINKERS TO DEFINE NUCLEAR ARCHITECTURE", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "study": "Heat Shock", "status": "released", "aliases": ["lis-lab:ExperimentSet_K562_InSituHiC_NHS_MboI_20170718"], "dbxrefs": ["GEO:GSE130758", "SRA:SRP195614"], "accession": "4DNESU95RUNO", "condition": "Control", "description": "in situ Hi-C on non-heat treated K562 cells with MboI", "study_group": "Disrupted or Atypical Cells", "date_created": "2018-10-11T20:56:05.073145+00:00", "submitted_by": {"error": "no view permissions"}, "dataset_label": "HiC on K562 cells +/- Heat Shock", "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2024-07-10T06:45:56.161808+00:00"}, "public_release": "2019-01-07", "replicate_exps": [{"bio_rep_no": 1, "tec_rep_no": 1, "replicate_exp": {"uuid": "23354847-f40b-4068-8e01-7aa78b33037d", "display_title": "in situ Hi-C on K562 (Tier 2) with MboI - 4DNEXYOJPVND", "accession": "4DNEXYOJPVND", "@id": "/experiments-hi-c/4DNEXYOJPVND/", "status": "released", "@type": ["ExperimentHiC", "Experiment", "Item"], "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}}, {"bio_rep_no": 2, "tec_rep_no": 1, "replicate_exp": {"uuid": "56612106-397c-4880-bc39-4b7acc9ce58c", "display_title": "in situ Hi-C on K562 (Tier 2) with MboI - 4DNEX15IP9D9", "accession": "4DNEX15IP9D9", "@id": "/experiments-hi-c/4DNEX15IP9D9/", "status": "released", "@type": ["ExperimentHiC", "Experiment", "Item"], "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}}], "schema_version": "2", "static_content": [{"content": {"display_title": "4DNESU95RUNO - Processed files", "title": "4DNESU95RUNO - Processed files", "@type": ["HiglassViewConfig", "UserContent", "Item"], "award": {"@type": ["Award", "Item"], "uuid": "d52d2d3e-a491-436f-9150-ee670b1fe3a4", "display_title": "DISTANCE-HI-C: CREATING PHOTO ACTIVATED X-LINKERS TO DEFINE NUCLEAR ARCHITECTURE", "@id": "/awards/1U01HL129958-01/", "status": "current", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "description": "4DNESU95RUNO (in situ Hi-C on non-heat treated K562 cells with MboI): 4DNFI244AS29, 4DNFI4SRNKNX, 4DNFIA7AR7WB, 4DNFI5WH9HQX", "filetype": "HiglassViewConfig", "uuid": "827cacab-ce17-4929-99e4-0556c5ec6c76", "contributing_labs": [], "lab": {"display_title": "John Lis, CORNELL", "@id": "/labs/john-lis-lab/", "status": "current", "@type": ["Lab", "Item"], "uuid": "c0bc993a-b7e2-4342-8523-6922c3c4dc35", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.lab_submitter", "submits_for.c0bc993a-b7e2-4342-8523-6922c3c4dc35"]}}, "name": "827cacab-ce17-4929-99e4-0556c5ec6c76", "status": "released", "@id": "/higlass-view-configs/827cacab-ce17-4929-99e4-0556c5ec6c76/", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.owner", "userid.986b362f-4eb6-4a9c-8173-3ab267307e3a"]}}, "location": "tab:processed-files", "description": "auto_generated_higlass_view_config"}], "static_headers": [{"lab": {"@id": "/labs/4dn-dcic-lab/", "@type": ["Lab", "Item"], "display_title": "4DN DCIC, HMS", "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": "<b>In Situ Hi-C</b>\n\n<p>\n   In situ Hi-C is a method to detect and quantify the pairwise interactions between chromosome regions across the entire genome. It was developed in 2014 as an improvement over the dilution Hi-C method. Compared to standard dilution Hi-C, this technique reduces the frequency of random ligation because the ligation is performed <i>in situ</i> inside the nucleus, a constrained space, instead of in solution, where DNA fragments are floating freely. In addition, this protocol can be done more quickly in the lab and was the first to introduce the use of 4-cutter restriction enzymes as opposed to the previous 6-cutters, providing higher resolution.\n</p>\n<p>\nThe protocol involves cross-linking the cells with formaldehyde to form links between physically adjacent DNA regions. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme is used to digest the chromatin into multiple DNA fragments. The resulting fragments are biotinylated by end filling of the fragments ends. The fragments are then ligated and the DNA is purified and sheared. The biotinylated fragments are pulled down from the solution with streptavidin beads and a library is constructed and sequenced. Analysis of the resulting paired-end short read sequences produces a matrix that shows the number of interactions between different DNA regions.\n</p>\n<p>\nSee <a href=\"https://www.sciencedirect.com/science/article/pii/S0092867414014974?via%3Dihub\">Rao et al., 2014</a> for more details.\n</p>\n\n<div>\n<img style=\"width: 600px;max-width:100%;\" src=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/IsHC_fig1.png\"/>\n  <br/><br/>\n  <em>Image source: Rao et al., 2014, Figure 1A</em>\n</div>", "name": "item-page-headers.ExperimentType.insituhic", "award": {"status": "current", "@type": ["Award", "Item"], "@id": "/awards/1U01CA200059-01/", "display_title": "4D NUCLEOME NETWORK DATA COORDINATION AND INTEGRATION CENTER - PHASE I", "uuid": "b0b9c607-f8b4-4f02-93f4-9895b461334b", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "title": "Assay Description", "status": "released", "aliases": ["4dn-dcic-lab:experiment_infobox_hic"], "options": {"filetype": "html", "collapsible": false, "default_open": false, "convert_ext_links": true}, "date_created": "2018-09-07T18:19:43.777198+00:00", "section_type": "Item Page Header", "submitted_by": {"error": "no view permissions"}, "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2023-11-09T14:48:01.220007+00:00"}, "schema_version": "2", "@id": "/static-sections/298554ad-20e2-4449-a752-ac190123dab7/", "@type": ["StaticSection", "UserContent", "Item"], "uuid": "298554ad-20e2-4449-a752-ac190123dab7", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.owner", "userid.56c9c683-bb11-471b-b590-c656f7dc03c1"]}, "display_title": "Assay Description", "external_references": [], "content": "<b>In Situ Hi-C</b>\n\n<p>\n   In situ Hi-C is a method to detect and quantify the pairwise interactions between chromosome regions across the entire genome. It was developed in 2014 as an improvement over the dilution Hi-C method. Compared to standard dilution Hi-C, this technique reduces the frequency of random ligation because the ligation is performed <i>in situ</i> inside the nucleus, a constrained space, instead of in solution, where DNA fragments are floating freely. In addition, this protocol can be done more quickly in the lab and was the first to introduce the use of 4-cutter restriction enzymes as opposed to the previous 6-cutters, providing higher resolution.\n</p>\n<p>\nThe protocol involves cross-linking the cells with formaldehyde to form links between physically adjacent DNA regions. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme is used to digest the chromatin into multiple DNA fragments. The resulting fragments are biotinylated by end filling of the fragments ends. The fragments are then ligated and the DNA is purified and sheared. The biotinylated fragments are pulled down from the solution with streptavidin beads and a library is constructed and sequenced. Analysis of the resulting paired-end short read sequences produces a matrix that shows the number of interactions between different DNA regions.\n</p>\n<p>\nSee <a href=\"https://www.sciencedirect.com/science/article/pii/S0092867414014974?via%3Dihub\">Rao et al., 2014</a> for more details.\n</p>\n\n<div>\n<img style=\"width: 600px;max-width:100%;\" src=\"https://s3.amazonaws.com/4dn-dcic-public/static-pages/InfoBoxes/IsHC_fig1.png\"/>\n  <br/><br/>\n  <em>Image source: Rao et al., 2014, Figure 1A</em>\n</div>", "filetype": "html", "content_as_html": "<div class=\"html-container\"><b>In Situ Hi-C</b>\n<p>\n   In situ Hi-C is a method to detect and quantify the pairwise interactions between chromosome regions across the entire genome. It was developed in 2014 as an improvement over the dilution Hi-C method. Compared to standard dilution Hi-C, this technique reduces the frequency of random ligation because the ligation is performed <i>in situ</i> inside the nucleus, a constrained space, instead of in solution, where DNA fragments are floating freely. In addition, this protocol can be done more quickly in the lab and was the first to introduce the use of 4-cutter restriction enzymes as opposed to the previous 6-cutters, providing higher resolution.\n</p>\n<p>\nThe protocol involves cross-linking the cells with formaldehyde to form links between physically adjacent DNA regions. The cells are then permeabilized with their nuclei intact. A 4-cutter restriction enzyme is used to digest the chromatin into multiple DNA fragments. The resulting fragments are biotinylated by end filling of the fragments ends. The fragments are then ligated and the DNA is purified and sheared. The biotinylated fragments are pulled down from the solution with streptavidin beads and a library is constructed and sequenced. 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This duplication does not affect the higlass display of these files, howevver, downstream analyses using this file may encounter issues due to this pixel duplication.  The counts from the duplicate pixels can be aggregated to determine the correct count value at that location. If this issue is problematic for your needs you should consider regenerating the matrices from the merged pairs file of the associated dataset using a more recent version of cooler.  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Our results suggest that the chromatin conformation necessary for a robust HS response is preestablished in NHS cells of diverse metazoan species.", "authors": ["Ray J", "Munn PR", "Vihervaara A", "Lewis JJ", "Ozer A", "Danko CG", "Lis JT"], "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "pubs_using": [], "publications_of_set": [{"@id": "/publications/5b2a068c-8de5-4d6f-8ac7-6a274444bfb2/", "authors": ["Ray J", "Munn PR", "Vihervaara A", "Lewis JJ", "Ozer A", "Danko CG", "Lis JT"], "ID": "PMID:31506350", "date_published": "2019-09-10", "abstract": "Heat shock (HS) initiates rapid, extensive, and evolutionarily conserved changes  in transcription that are accompanied by chromatin decondensation and nucleosome  loss at HS loci. Here we have employed in situ Hi-C to determine how heat stress  affects long-range chromatin conformation in human and Drosophila cells. We found that compartments and topologically associating domains (TADs) remain unchanged by an acute HS. Knockdown of Heat Shock Factor 1 (HSF1), the master transcriptional regulator of the HS response, identified HSF1-dependent genes and revealed that up-regulation is often mediated by distal HSF1 bound enhancers. HSF1-dependent genes were usually found in the same TAD as the nearest HSF1 binding site. Although most interactions between HSF1 binding sites and target promoters were established in the nonheat shock (NHS) condition, a subset increased contact frequency following HS. Integrating information about HSF1 binding strength, RNA polymerase abundance at the HSF1 bound sites (putative enhancers), and contact frequency with a target promoter accurately predicted which up-regulated genes were direct targets of HSF1 during HS. Our results suggest that the chromatin conformation necessary for a robust HS response is preestablished in NHS cells of diverse metazoan species.", "title": "Chromatin conformation remains stable upon extensive transcriptional changes driven by heat shock.", "@type": ["Publication", "Item"], "journal": "Proceedings of the National Academy of Sciences of the United States of America", "uuid": "5b2a068c-8de5-4d6f-8ac7-6a274444bfb2", "status": "current", "display_title": "Ray J et al. 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