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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.\"", "uuid": "ef637c2d-7e48-4e26-aad2-eac1f680d4e8", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "status": "released", "aliases": ["belmont-lab:thrap3-tsa-seq-v2-u2os-cond3primeG"], "accession": "4DNESYQ24WZ9", "condition": "condition3enhancementG - anti-THRAP3", "description": "Replicates of THRAP3 TSA-seq version 2 Reaction Condition 3' (PBS 50% Sucrose plus 1.5 mM DTT) 30 minute Reaction on U2OS cells", "date_created": "2024-12-19T15:40:56.368980+00:00", "submitted_by": {"error": "no view permissions"}, "dataset_label": "TSA-seq v2 THRAP3 in U2OS", "last_modified": {"modified_by": {"error": "no view permissions"}, "date_modified": "2025-08-29T06:34:29.598692+00:00"}, "public_release": "2025-08-27", 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Genes within these regions amplify their expression only with NS contact. However, gene expression differences inversely correlate with differences in NS distance, genome-wide. We hypothesized the existence of additional gene expression niches away from, but spatially correlated with, NS. Here we report the identification of two dynamic perispeckle patterns of protein concentrations extending outwards from NS and persisting even after NS are eliminated. Highly active chromosome regions which weakly associate with NS instead show close, NS-independent association with these perispeckle patterns. Additionally, transcripts from model intron-containing versus intronless genes associate differentially with these two patterns. While genes within NS-associated genomic regions are predominantly downregulated upon NS depletion, genes associated with perispeckle patterns are biased towards upregulation. We suggest the interchromatin space is partitioned into additional gene expression niches- surrounding and extending from NS - that may be involved in mRNA and gene dynamics.", "uuid": "e2ff1ddb-d194-462c-b74d-4987a1b74350", "status": "current", "url": "http://biorxiv.org/lookup/doi/10.1101/2025.08.23.671945", "@type": ["Publication", "Item"], "display_title": "Venkata NC et al. (2025) doi:10.1101/2025.08.23.671945", "@id": "/publications/e2ff1ddb-d194-462c-b74d-4987a1b74350/", "authors": ["Venkata NC", "Kim J", "Faber G", "Misra S", "Bektash A", "Chaturvedi P", "Hernanadez G", "Dopie J", "Kanemaki M", "Han KY", "Shav-Tal Y", "Belmont AS"], "journal": "bioRxiv", "short_attribution": "Venkata NC et al. (2025)", "title": "Highly active chromosome regions preferentially associate with two perispeckle networks that partition the interchromatin space", "ID": "doi:10.1101/2025.08.23.671945", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "pubs_using": [], "publications_of_set": [{"journal": "bioRxiv", "authors": ["Venkata NC", "Kim J", "Faber G", "Misra S", "Bektash A", "Chaturvedi P", "Hernanadez G", "Dopie J", "Kanemaki M", "Han KY", "Shav-Tal Y", "Belmont AS"], "title": "Highly active chromosome regions preferentially associate with two perispeckle networks that partition the interchromatin space", "uuid": "e2ff1ddb-d194-462c-b74d-4987a1b74350", "abstract": "A subset of highly active chromosomal hot zones reproducibly positions adjacent to nuclear speckles (NS). Genes within these regions amplify their expression only with NS contact. However, gene expression differences inversely correlate with differences in NS distance, genome-wide. We hypothesized the existence of additional gene expression niches away from, but spatially correlated with, NS. Here we report the identification of two dynamic perispeckle patterns of protein concentrations extending outwards from NS and persisting even after NS are eliminated. Highly active chromosome regions which weakly associate with NS instead show close, NS-independent association with these perispeckle patterns. Additionally, transcripts from model intron-containing versus intronless genes associate differentially with these two patterns. While genes within NS-associated genomic regions are predominantly downregulated upon NS depletion, genes associated with perispeckle patterns are biased towards upregulation. We suggest the interchromatin space is partitioned into additional gene expression niches- surrounding and extending from NS - that may be involved in mRNA and gene dynamics.", "ID": "doi:10.1101/2025.08.23.671945", "display_title": "Venkata NC et al. (2025) doi:10.1101/2025.08.23.671945", "@id": "/publications/e2ff1ddb-d194-462c-b74d-4987a1b74350/", "status": "current", "@type": ["Publication", "Item"], "date_published": "2025-08-26", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}], "number_of_experiments": 3, "@context": "/terms/", "aggregated-items": {"badges": [{"parent": "/biosamples/4DNBS61886MX/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing doubling_number"], "badge": {"commendation": null, "warning": "Biosample Metadata Incomplete", "uuid": "2b2cc7ff-b7a8-4138-9a6c-22884fc71690", "@id": "/badges/biosample-metadata-incomplete/", "badge_icon": "/static/img/badges/biosample-icon.svg", "description": "Biosample is missing metadata information required as part of the standards implemented by the 4DN Samples working group."}}}, {"parent": "/biosamples/4DNBSKR9NL37/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing doubling_number"], "badge": {"commendation": null, "warning": "Biosample Metadata Incomplete", "uuid": "2b2cc7ff-b7a8-4138-9a6c-22884fc71690", "@id": "/badges/biosample-metadata-incomplete/", "badge_icon": "/static/img/badges/biosample-icon.svg", "description": "Biosample is missing metadata information required as part of the standards implemented by the 4DN Samples working group."}}}, {"parent": "/biosamples/4DNBS5PTAHOL/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing doubling_number", "Tiered cell line cultured after 2022-05-10 missing 4DN specified FBS vendor and lot info"], "badge": {"commendation": null, "warning": "Biosample Metadata Incomplete", "uuid": "2b2cc7ff-b7a8-4138-9a6c-22884fc71690", "@id": "/badges/biosample-metadata-incomplete/", "badge_icon": "/static/img/badges/biosample-icon.svg", "description": "Biosample is missing metadata information required as part of the standards implemented by the 4DN Samples working group."}}}]}, "validation-errors": []}