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This proposal is significant because we propose a novel model forthe role of nuclear organization in regulating gene expression: 1) Nuclear speckles and still unknown nuclearcompartments/bodies help organize other phase-separated condensates to modulate gene expression; 2)Nuclear speckles together with surrounding nuclear compartments/bodies and associated phase-separatedcondensates together represent active nuclear niches which may have different functional properties; 3) Smalldistances matter: gene movements of only a few hundred nm between repressive and these different activenuclear niches may differentially regulate gene expression; 4) Action-at-a distance: component flux into andout of these nuclear compartments will have global effects on gene expression; 5) These same nuclearcompartments/bodies may similarly modulate RNA processing and organize nuclear export. Here we propose to: 1) Identify multiple components of known and still unknown nuclear \u201cactiveniches\u201d; 2) Map genome-wide the positions and predicted movements of genes relative to these active nichesduring physiological transitions; 3) Visualize nuclear body/compartment dynamics and fluxes of proteinsbetween nuclear bodies in steady-state and through physiological transitions; 4) Visualize movements ofreporter transgenes, endogenous genes, and rewired chromosome loci relative to these nuclearbodies/compartments and temporally correlate changes in gene expression with their dynamic movements andcompartment associations; 5) Visualize movements of pre-mRNAs and nuclear mRNAs during RNAprocessing and export; 6) Measure fluxes of nuclear body components to and from adjacent transcribingchromatin. Additionally, we propose developing relatively low-cost, novel microscope platforms and softwarespecifically designed to facilitate these live-cell imaging goals in our laboratories as well as others. Our Aims will be to: 1. Map proteins, genes, RNAs relative to active nuclear compartment(s) usingiterative rounds of TSA-MS-Ratio, validation by light microscopy, and TSA-Seq; 2. Measure dynamics ofbodies, components of nuclear bodies using live-cell imaging; 3. Measure temporal correlation betweenchanges in gene expression and gene movement relative to nuclear bodies and visualize the export path ofexpressed transcripts; 4. Design and deliver two novel microscopes designed to facilitate Aims 1-3 at amodest cost. Successful completion of these Aims should significantly change our current understanding of therole of nuclear organization in regulating gene expression with impact across a wide range of research fields.\"", "@id": "/awards/1U01DK127422-01/", "status": "current", "project": "4DN", "uuid": "ef637c2d-7e48-4e26-aad2-eac1f680d4e8", "center_title": "Belmont", "@type": ["Award", "Item"], "display_title": "IDENTIFICATION OF THE ACTIVE NUCLEAR NICHE(S) USING NOVEL PROTEOMIC, GENOMIC, TRANSGENIC, AND LIVE-CELL MICROSCOPY TECHNOLOGIES", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "md5sum": "e01368f3cc74d5f8fc1e965b8ddb97cf", "status": "released", "aliases": ["belmont-lab:U2OS_SON_2xDTT_Input_bio1_ATGTCAGA_L001_R1_001.fastq"], "filename": "U2OS_SON_2xDTT_Input_bio1_ATGTCAGA_L001_R1_001.fastq.gz", "accession": "4DNFIR4QVKOR", "file_size": 1718141585, "file_type": "reads", "instrument": "NovaSeq6000", "file_format": {"status": "released", "display_title": "fastq", "@id": "/file-formats/fastq/", "uuid": "c13d06cf-218e-4f61-aaf0-91f226248b2c", "@type": ["FileFormat", "Item"], "file_format": "fastq", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "read_length": 100, "date_created": "2024-12-19T15:40:02.663803+00:00", "submitted_by": {"error": "no view permissions"}, "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2025-09-05T17:06:31.720137+00:00"}, "content_md5sum": "5a5321290a195af0232eb4f29d35fad9", "public_release": "2025-08-27", "quality_metric": {"display_title": "QualityMetricFastqc from 2024-12-20", "@type": ["QualityMetricFastqc", "QualityMetric", "Item"], "@id": "/quality-metrics-fastqc/d58ee5e2-1882-4a1d-a091-5eb983a9f6d7/", "Total Sequences": 36634348, "status": "released", "overall_quality_status": "PASS", "uuid": "d58ee5e2-1882-4a1d-a091-5eb983a9f6d7", "Sequence length": "100", "url": "https://s3.amazonaws.com/elasticbeanstalk-fourfront-webprod-wfoutput/4DNFIR4QVKOR/fastqc_report.html", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "schema_version": "2", "file_first_line": "@A00419:487:HL25CDRXY:1:2101:1163:1000 1:N:0:ATGTCAGA", "project_release": "2025-08-27", "flowcell_details": [{"barcode": "ATGTCAGA", "machine": "NovaSeq6000"}], "file_classification": "raw file", "@id": "/files-fastq/4DNFIR4QVKOR/", "@type": ["FileFastq", "File", "Item"], "uuid": "a6fe5828-7372-4643-8710-80777c95e98e", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}, "display_title": "4DNFIR4QVKOR.fastq.gz", "external_references": [], "experiments": [{"display_title": "TSA-seq against SON protein on U2OS (Tier 2) - 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