{"lab": {"correspondence": [{"contact_email": "aHVhaXlpbnpAYW5kcmV3LmNtdS5lZHU=", "@id": "/users/c55c809d-8ffc-42f9-a528-12db012f846f/", "display_title": "Huaiying Zhang"}], "uuid": "3525602a-81b2-4db9-92bd-858ef82e1b53", "status": "current", "display_title": "Huaiying Zhang, CMU", "@type": ["Lab", "Item"], "title": "Huaiying Zhang, CMU", "@id": "/labs/huaiying-zhang-lab/", "pi": {"error": "no view permissions"}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin", "role.lab_submitter", "submits_for.3525602a-81b2-4db9-92bd-858ef82e1b53"]}}, "award": {"display_title": "PHASE SEPARATION-INDUCED NUCLEAR ORGANIZATION IN ALT CANCER", "status": "current", "name": "1U01CA260851-01", "project": "4DN", "description": "NI-OFHHD: All cancer cells need to maintain telomere length for immortality. While most cancer cells reactivate telomerase, a reverse transcriptase, to elongates telomere from an RNA template, about 10-15% of cancer cells are telomerase-negative and adopt a homologous-recombination based alternative lengthening of telomeres (ALT) pathway. ALT cells exhibit many abnormalities in nuclear organization, including the formation of nuclear bodies called APBs for ALT telomere-associated promyelocytic leukemia nuclear bodies, clustering of telomeres within APBs, and the formation of RNA foci on telomeres with a long non-coding RNA called telomere repeat-containing RNA (TERRA). These unique features are used as biomarkers for ALT diagnosis and can be attractive therapeutic targets because of reduced side effects on healthy cells that do not share these features. However, how these features contribute to telomere maintenance and ALT cancer cell growth remain elusive, due to the lack of conceptual model as well as experimental tools to monitor and control their assembly and function in live cells. Based on our observation that APBs exhibit liquid behavior and long non- coding RNAs can phase separate with RNA-binding proteins, we propose a liquid-liquid phase separation model for the assmembly and function of these ALT specific features. We hypothesize TERRA phase separates with its interacting proteins to nucleate APB liquid droplets. The liquid nature of APBs droplets (also called condensates) would promote coalescence of APBs to drive telomere clustering. Meanwhile, condensation of APB droplets can concentrate DNA repair factors, providing opportunities for telomeres to use one another as repair templates to elongate within APBs. To test our hypothesis, we developed a state-of-the- art optogenetic approach to control APB assembly. We demonstrate that liquid phase separation underlies APB assembly and coalescence of APB droplets indeed drives telomere clustering. Building on our ability to control telomere clustering and APB assembly and by collaborating with experts in super resolution microscopy, nuclear mechanics, chromosome organization and ALT cancer, we will investigate how DNA repair factors are recruited to and organized in APB condensates for ALT telomere DNA synthesis (Aim 1) and how telomere clustering leads to unique genome organization and gene expression in ALT cells (Aim 2). We will then extend our optogenetic tools to control RNA and dissect TERRA contributions in ALT (Aim 3). Results obtained by manipulating cultured ALT cells will be confirmed by characterizing ALT tissue or creating de novo ALT phonotypes in primary human cells. Our results will provide mechanistic understanding on how protein and/or RNA phase separation contributes to ALT cancer, which will offer the potential to develop strategies specifically targeting these unique phase separation processes, rather than the existing molecules that shared by heathy cells, for ALT cancer treatment. 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(2024) PMID:38461301", "@type": ["Publication", "Item"], "title": "TERRA-LSD1 phase separation promotes R-loop formation for telomere maintenance in  ALT cancer cells.", "status": "current", "journal": "Nature communications", "uuid": "7ba475b0-cc62-4723-93eb-390269c0f3ed", "@id": "/publications/7ba475b0-cc62-4723-93eb-390269c0f3ed/", "abstract": "The telomere repeat-containing RNA (TERRA) forms R-loops to promote  homology-directed DNA synthesis in the alternative lengthening of telomere (ALT)  pathway. Here we report that TERRA contributes to ALT via interacting with the  lysine-specific demethylase 1A (LSD1 or KDM1A). We show that LSD1 localizes to  ALT telomeres in a TERRA dependent manner and LSD1 function in ALT is largely  independent of its demethylase activity. Instead, LSD1 promotes TERRA recruitment  to ALT telomeres via RNA binding. In addition, LSD1 and TERRA undergo phase  separation, driven by interactions between the RNA binding properties of LSD1 and  the G-quadruplex structure of TERRA. Importantly, the formation of TERRA-LSD1  condensates enriches the R-loop stimulating protein Rad51AP1 and increases  TERRA-containing R-loops at telomeres. Our findings suggest that LSD1-TERRA phase  separation enhances the function of R-loop regulatory molecules for ALT telomere  maintenance, providing a mechanism for how the biophysical properties of histone  modification enzyme-RNA interactions impact chromatin function.", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}], "number_of_experiments": 3, "imaging_paths": [{"path": {"display_title": "Chromosomes targeted by DAPI", "labeled_probe": "DAPI", "@id": "/imaging-paths/357362cd-99f2-436f-a8f5-27f06d305f29/", "@type": ["ImagingPath", "Item"], "status": "released", "uuid": "357362cd-99f2-436f-a8f5-27f06d305f29", "target": [{"cellular_structure": "Chromosomes", "organism_name": "unspecified", "uuid": "655728f9-d4ab-4dd2-971c-1627ab2c8d33", "status": "released", "@id": "/bio-features/655728f9-d4ab-4dd2-971c-1627ab2c8d33/", "@type": ["BioFeature", "Item"], "display_title": "Chromosomes", "feature_type": {"uuid": "e47e2bea-bc1e-4989-881c-521fc0b33529", "status": "released", "display_title": "cellular_component", "@type": ["OntologyTerm", "Item"], "term_id": "GO:0005575", "preferred_name": "cellular_component", "term_name": "cellular_component", "term_url": "http://purl.obolibrary.org/obo/GO_0005575", "@id": "/ontology-terms/GO:0005575/", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}], "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "channel": "ch00"}, {"path": {"display_title": "Telomere DNA (human) targeted by Alexa Fluor 488", "@id": "/imaging-paths/2e0c2a51-ad2d-4f8d-8761-4c190bd54e9a/", "@type": ["ImagingPath", "Item"], "status": "released", "uuid": "2e0c2a51-ad2d-4f8d-8761-4c190bd54e9a", "target": [{"organism_name": "human", "uuid": "3b4b9c25-3e1c-4e14-89db-8a37c95ca899", "status": "released", "@id": "/bio-features/3b4b9c25-3e1c-4e14-89db-8a37c95ca899/", "@type": ["BioFeature", "Item"], "preferred_label": "Telomere DNA (human)", "display_title": "Telomere DNA (human)", "feature_type": {"uuid": "41b7a2c5-f2d9-46b4-a49d-51502fe61bb3", "status": "released", "display_title": "region", "@type": ["OntologyTerm", "Item"], "term_id": "SO:0000001", "preferred_name": "region", "term_name": "region", "term_url": "http://purl.obolibrary.org/obo/SO_0000001", "@id": "/ontology-terms/SO:0000001/", "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}], "principals_allowed": {"view": ["system.Everyone"], "edit": ["group.admin"]}}, "channel": "ch01"}], "@context": "/terms/", "aggregated-items": {"badges": [{"parent": "/biosamples/4DNBSD87XWGC/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing Cell Culture Details"], "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/4DNBSLR1881C/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing Cell Culture Details"], "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/4DNBS1Q27TQE/", "embedded_path": "experiments_in_set.biosample.badges", "item": {"messages": ["Biosample missing Cell Culture Details"], "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": []}