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Fujiwara N, Ueno T, Yamazaki T, Hirose T. Unraveling architectural RNAs: Structural and functional blueprints of membraneless organelles and strategies for genome-scale identification. Biochim Biophys Acta Gen Subj 2025; 1869:130815. [PMID: 40348038 DOI: 10.1016/j.bbagen.2025.130815] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2025] [Revised: 04/25/2025] [Accepted: 05/06/2025] [Indexed: 05/14/2025]
Abstract
Architectural RNAs (arcRNAs) are long noncoding RNAs that serve as structural scaffolds for membraneless organelles (MLOs), facilitating cellular organization and dynamic responses to stimuli. Acting as blueprints for MLO assembly, arcRNAs recruit specific proteins and nucleic acids to establish and maintain the internal structure of MLOs while coordinating their spatial relationships with other organelles. This organized framework enables precise spatiotemporal regulation, allowing for targeted control of transcription, RNA processing, and cellular responses to stress. Notably, arcRNAs exhibit the "semi-extractable" feature, a property derived from their stable binding to cellular structures, making them partially resistant to conventional RNA extraction methods. This unique feature serves as a useful criterion for identifying novel arcRNAs, providing an opportunity to accelerate research in long noncoding RNAs and deepen our understanding of their functional roles in cellular processes.
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Affiliation(s)
- Naoko Fujiwara
- Graduate School of Frontier Biosciences, The University of Osaka, Suita 565-0871, Japan
| | - Tsuyoshi Ueno
- Graduate School of Frontier Biosciences, The University of Osaka, Suita 565-0871, Japan
| | - Tomohiro Yamazaki
- Graduate School of Frontier Biosciences, The University of Osaka, Suita 565-0871, Japan
| | - Tetsuro Hirose
- Graduate School of Frontier Biosciences, The University of Osaka, Suita 565-0871, Japan.
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2
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Zhai PL, Chen MM, Wang Q, Zhao JJ, Tang XM, Lu CN, Liu J, Yang QX, Xiang ML, Tang QH, Gu B, Zhang SP, Tang SP, Fu D. Multi-omics analysis identifies a liquid-liquid phase separation-related subtypes in head and neck squamous cell carcinoma. Front Oncol 2025; 15:1509810. [PMID: 40078192 PMCID: PMC11897011 DOI: 10.3389/fonc.2025.1509810] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2024] [Accepted: 02/11/2025] [Indexed: 03/14/2025] Open
Abstract
Background Growing evidence indicates that abnormal liquid-liquid phase separation (LLPS) can disrupt biomolecular condensates, contributing to cancer development and progression. However, the influence of LLPS on the prognosis of head and neck squamous cell carcinoma (HNSCC) patients and its effects on the tumor immune microenvironment (TIME) are not yet fully understood. Therefore, we aimed to categorize patients with HNSCC based on LLPS-related genes and explored their multidimensional heterogeneity. Methods We integrated the transcriptomic data of 3,541 LLPS-related genes to assess the LLPS patterns in 501 patients with HNSCC within The Cancer Genome Atlas cohort. Subsequently, we explored the differences among the three LLPS subtypes using multi-omics analysis. We also developed an LLPS-related prognostic risk signature (LPRS) to facilitate personalized and integrative assessments and then screened and validated potential therapeutic small molecule compounds targeting HNSCC via experimental analyses. Result By analyzing the expression profiles of 85 scaffolds, 355 regulators, and 3,101 clients of LLPS in HNSCC, we identified three distinct LLPS subtypes: LS1, LS2, and LS3. We confirmed notable differences among these subtypes in terms of prognosis, functional enrichment, genomic alterations, TIME patterns, and responses to immunotherapy. Additionally, we developed the LPRS, a prognostic signature for personalized integrative assessments, which demonstrated strong predictive capability for HNSCC prognosis across multiple cohorts. The LPRS also showed significant correlations with the clinicopathological features and TIME patterns in HNSCC patients. Furthermore, the LPRS effectively predicted responses to immune checkpoint inhibitor therapy and facilitated the screening of potential small-molecule compounds for treating HNSCC patients. Conclusion This study presents a new classification system for HNSCC patients grounded in LLPS. The LPRS developed in this research offers improved personalized prognosis and could optimize immunotherapy strategies for HNSCC.
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Affiliation(s)
- Peng-Lei Zhai
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Meng-Min Chen
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- Research Institute of Pancreatic Diseases, Shanghai Key Laboratory of Translational Research for Pancreatic Neoplasms, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- State Key Laboratory of Oncogenes and Related Genes, Institute of Translational Medicine, Shanghai Jiao Tong University, Shanghai, China
| | - Qi Wang
- Department of Urology, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Jing-Jun Zhao
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
| | - Xiao-Mei Tang
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Cui-Ni Lu
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Jia Liu
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Qin-Xin Yang
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Ming-Liang Xiang
- Department of Otolaryngology & Head and Neck Surgery, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Qing-Hai Tang
- Hunan Key Laboratory for Conservation and Utilization of Biological Resources in the Nanyue Mountainous Region, College of Life Sciences, Hengyang Normal University, Hengyang, China
| | - Biao Gu
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
| | - Shu-Ping Zhang
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
| | - Si-Ping Tang
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
| | - Da Fu
- Key Laboratory of Functional Organometallic Materials of College of Hunan Province, College of Chemistry and Materials Science, Hengyang Normal University, Hengyang, China
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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Arias Escayola D, Zhang C, Nischwitz E, Schärfen L, Dörner K, Straube K, Kutay U, Butter F, Neugebauer KM. Identification of coilin interactors reveals coordinated control of Cajal body number and structure. J Cell Biol 2025; 224:e202305081. [PMID: 39602297 PMCID: PMC11602656 DOI: 10.1083/jcb.202305081] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2023] [Revised: 10/13/2024] [Accepted: 11/04/2024] [Indexed: 11/29/2024] Open
Abstract
The cell nucleus contains distinct biomolecular condensates that form at specific genetic loci, organize chromosomes in 3D space, and regulate RNA processing. Among these, Cajal bodies (CBs) require key "scaffolding" proteins for their assembly, which is not fully understood. Here, we employ proximity biotinylation, mass spectrometry, and functional screening to comprehensively identify and test the functions of CB components. We document 144 protein interactors of coilin, of which 70 were newly detected, and establish 25 players needed for CB assembly and/or maintenance. Surprisingly, the depletion of nine coilin interactors-mostly constituents of the 60S ribosome (RPLs)-increased CB number and caused subdomains defined by coilin and the survival motor neuron protein (SMN) to merge. These phenotypes were traceable to altered nuclear levels of dimethylarginine. Our data implicate RPL24 and other players in the regulation of CBs by modulating posttranslational modifications. Moreover, the prevalence of transcription factors among the identified components highlights roles for gene activity in CB assembly and nuclear positioning.
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Affiliation(s)
- Dahyana Arias Escayola
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Chuyue Zhang
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | | | - Leonard Schärfen
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Kerstin Dörner
- Department of Biology, Institute of Biochemistry, ETH Zürich, Zürich, Switzerland
| | - Korinna Straube
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Ulrike Kutay
- Department of Biology, Institute of Biochemistry, ETH Zürich, Zürich, Switzerland
| | - Falk Butter
- Institute of Molecular Biology (IMB), Mainz, Germany
| | - Karla M. Neugebauer
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
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Zhang H, Wang Z, Zhang J, Li Z, Liu J, Yu J, Zhao Y, Guo F, Chen WD, Wang YD. A MYC-STAMBPL1-TOE1 positive feedback loop mediates EGFR stability in hepatocellular carcinoma. Cell Rep 2024; 43:114812. [PMID: 39388352 DOI: 10.1016/j.celrep.2024.114812] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2023] [Revised: 08/24/2024] [Accepted: 09/17/2024] [Indexed: 10/12/2024] Open
Abstract
The role of STAM binding protein-like 1 (STAMBPL1), a Lys-63 linkage-specific deubiquitinase, in hepatocellular carcinoma has remained elusive. Here, we report the functions of STAMBPL1 in modulating the stability of the protein and mRNA of the epidermal growth factor receptor (EGFR). STAMBPL1 deficiency attenuates liver tumorigenesis in vitro and in vivo. STAMBPL1 removes K63-linked ubiquitin chains from EGFR to avoid lysosome degradation upon EGF stimulation. STAMBPL1 augments RNA efficient splicing of EGFR to avoid intron retention by activating cleavage of the K63-linked ubiquitin chain on the target of EGR1 protein 1 (TOE1). Moreover, the EGFR-MYC axis has a positive feedback regulation on the transcription of STAMBPL1, and depletion of STAMBPL1 in vivo blunts MYC-driven liver tumorigenesis. Inhibition of STAMBPL1 or TOE1 synergistically improves the antitumor activity of lenvatinib. Our work shows the mechanism of STAMBPL1 in liver cancer and suggests it as a potential therapeutic target for liver cancer treatment.
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Affiliation(s)
- Hongli Zhang
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China
| | - Zixuan Wang
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China
| | - Jian Zhang
- Department of Clinical Pathology, Nanyang Central Hospital, Nanyang, P.R. China
| | - Zhengtai Li
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China
| | - Jiaxuan Liu
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China
| | - Jingwen Yu
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China; Key Laboratory of Receptors-Mediated Gene Regulation and Drug Discovery, School of Basic Medical Science, Inner Mongolia Medical University, Hohhot, Inner Mongolia, P. R. China
| | - Yiqi Zhao
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China; Key Laboratory of Receptors-Mediated Gene Regulation and Drug Discovery, School of Basic Medical Science, Inner Mongolia Medical University, Hohhot, Inner Mongolia, P. R. China
| | - Fan Guo
- Key Laboratory of Receptors-Mediated Gene Regulation and Drug Discovery, School of Basic Medical Science, Inner Mongolia Medical University, Hohhot, Inner Mongolia, P. R. China
| | - Wei-Dong Chen
- Key Laboratory of Receptors-Mediated Gene Regulation and Drug Discovery, School of Basic Medical Science, Inner Mongolia Medical University, Hohhot, Inner Mongolia, P. R. China; Key Laboratory of Receptors-Mediated Gene Regulation, Hebi Key Laboratory of Liver Disease, The People's Hospital of Hebi, School of Medicine, Henan University, Henan, P.R. China.
| | - Yan-Dong Wang
- State Key Laboratory of Chemical Resource Engineering, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, P.R. China.
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Thivierge C, Bellefeuille M, Diwan SS, Dyakov BJA, Leventis R, Perron G, Najafabadi HS, Gravel SP, Gingras AC, Duchaine TF. Paraspeckle-independent co-transcriptional regulation of nuclear microRNA biogenesis by SFPQ. Cell Rep 2024; 43:114695. [PMID: 39250314 DOI: 10.1016/j.celrep.2024.114695] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2024] [Revised: 06/28/2024] [Accepted: 08/13/2024] [Indexed: 09/11/2024] Open
Abstract
MicroRNAs (miRNAs) play crucial roles in physiological functions and disease, but the regulation of their nuclear biogenesis remains poorly understood. Here, BioID on Drosha, the catalytic subunit of the microprocessor complex, reveals its proximity to splicing factor proline- and glutamine (Q)-rich (SFPQ), a multifunctional RNA-binding protein (RBP) involved in forming paraspeckle nuclear condensates. SFPQ depletion impacts both primary and mature miRNA expression, while other paraspeckle proteins (PSPs) or the paraspeckle scaffolding RNA NEAT1 do not, indicating a paraspeckle-independent role. Comprehensive transcriptomic analyses show that SFPQ loss broadly affects RNAs and miRNA host gene (HG) expression, influencing both their transcription and the stability of their products. Notably, SFPQ protects the oncogenic miR-17∼92 polycistron from degradation by the nuclear exosome targeting (NEXT)-exosome complex and is tightly linked with its overexpression across a broad variety of cancers. Our findings reveal a dual role for SFPQ in regulating miRNA HG transcription and stability, as well as its significance in cancers.
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Affiliation(s)
- Caroline Thivierge
- Rosalind and Morris Goodman Cancer Institute, McGill Centre for RNA Sciences & Department of Biochemistry, McGill University, Montréal, QC H3G 1Y6, Canada
| | - Maxime Bellefeuille
- Rosalind and Morris Goodman Cancer Institute, McGill Centre for RNA Sciences & Department of Biochemistry, McGill University, Montréal, QC H3G 1Y6, Canada
| | - Sarah-Slim Diwan
- Rosalind and Morris Goodman Cancer Institute, McGill Centre for RNA Sciences & Department of Biochemistry, McGill University, Montréal, QC H3G 1Y6, Canada
| | - Boris J A Dyakov
- Lunenfeld-Tanenbaum Research Institute, Sinai Health System & Department of Molecular Genetics, University of Toronto, Toronto, ON M5G 1X5, Canada
| | - Rania Leventis
- Rosalind and Morris Goodman Cancer Institute, McGill Centre for RNA Sciences & Department of Biochemistry, McGill University, Montréal, QC H3G 1Y6, Canada
| | - Gabrielle Perron
- McGill Genome Centre & Department of Human Genetics, McGill University, Montréal, QC H3A 0G1, Canada
| | - Hamed S Najafabadi
- McGill Genome Centre & Department of Human Genetics, McGill University, Montréal, QC H3A 0G1, Canada
| | | | - Anne-Claude Gingras
- Lunenfeld-Tanenbaum Research Institute, Sinai Health System & Department of Molecular Genetics, University of Toronto, Toronto, ON M5G 1X5, Canada
| | - Thomas F Duchaine
- Rosalind and Morris Goodman Cancer Institute, McGill Centre for RNA Sciences & Department of Biochemistry, McGill University, Montréal, QC H3G 1Y6, Canada.
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Gilloteaux J, Jamison JM, Summers JL, Taper HS. Reactivation of nucleases with peroxidation damages induced by a menadione: ascorbate combination devastates human prostate carcinomas: ultrastructural aspects. Ultrastruct Pathol 2024; 48:378-421. [PMID: 39105605 DOI: 10.1080/01913123.2024.2379300] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2024] [Revised: 06/27/2024] [Accepted: 07/09/2024] [Indexed: 08/07/2024]
Abstract
INTRODUCTION Xenografts of androgen-independent human DU145 prostate metastatic carcinomas implanted in nu/nu male mice have revealed a significant survival after a prooxidant anticancer treatment consisting of a combination of menadione bisulfite and sodium ascorbate (VK3:VC). METHODS Implanted samples of diaphragm carcinomas from longest survived mice from either oral, intraperitoneal (IP), or both oral and IP treatment groups were assessed with light, scanning, and transmission electron microscopy to analyze morphologic damages. RESULTS Compared with previous fine structure data of in vitro untreated carcinomas, the changes induced by oral, IP, and oral with IP VK3:VC treatment dismantled those xenografts with autoschizis, and necrotic atrophy was accomplished by cell's oxidative stress whose injuries were consequent to reactivated deoxyribonucleases and ribonucleases. Tumor destructions resulted from irreversible damages of nucleus components, endoplasmic reticulum, and mitochondria there. Other alterations included those of the cytoskeleton that resulted in characteristic self-excisions named " autoschizis." All these injuries lead resilient cancer cells to necrotic cell death. CONCLUSION The fine structure damages caused by VK3:VC prooxidant combination in the human DU145 prostate xenografts confirmed those shown in vitro and of other cell lines with histochemistry and biomolecular investigations. These devastations incurred without damage to normal tissues; thus, our data brought support for the above combination to assist in the treatment of prostate cancers and other cancers.
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Affiliation(s)
- Jacques Gilloteaux
- Department of Anatomical Sciences, St Georges' University International School of Medicine, Newcastle upon Tyne, UK
- Department of Anatomical Sciences, NEOMed (NEOUCOM), Rootstown, Ohio, USA
- Department of Medicine, Unit of Research in Molecular Physiology (URPhyM), NARILIS, Université de Namur, Namur, Belgium
| | - James M Jamison
- Department of Urology, Summa Health System, Akron, Ohio, USA
- St Thomas Hospital, The Apatone Development Center, Summa Research Fondation, Akron Ohio, USA
| | - Jack L Summers
- Department of Urology, Summa Health System, Akron, Ohio, USA
- St Thomas Hospital, The Apatone Development Center, Summa Research Fondation, Akron Ohio, USA
| | - Henryk S Taper
- Département des Sciences Pharmaceutiques, Unité de Pharmacocinétique, Métabolisme, Nutrition et Toxicologie, Université Catholique de Louvain, Brussels, Belgium
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7
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Toya H, Okamatsu-Ogura Y, Yokoi S, Kurihara M, Mito M, Iwasaki S, Hirose T, Nakagawa S. The essential role of architectural noncoding RNA Neat1 in cold-induced beige adipocyte differentiation in mice. RNA (NEW YORK, N.Y.) 2024; 30:1011-1024. [PMID: 38692841 PMCID: PMC11251523 DOI: 10.1261/rna.079972.124] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/30/2024] [Accepted: 04/08/2024] [Indexed: 05/03/2024]
Abstract
Neat1 is an architectural RNA that provides the structural basis for nuclear bodies known as paraspeckles. Although the assembly processes by which Neat1 organizes paraspeckle components are well-documented, the physiological functions of Neat1 are not yet fully understood. This is partly because Neat1 knockout (KO) mice, lacking paraspeckles, do not exhibit overt phenotypes under normal laboratory conditions. During our search for conditions that elicit clear phenotypes in Neat1 KO mice, we discovered that the differentiation of beige adipocytes-inducible thermogenic cells that emerge upon cold exposure-is severely impaired in these mutant mice. Neat1_2, the architectural isoform of Neat1, is transiently upregulated during the early stages of beige adipocyte differentiation, coinciding with increased paraspeckle formation. Genes with altered expression during beige adipocyte differentiation typically cluster at specific chromosomal locations, some of which move closer to paraspeckles upon cold exposure. These observations suggest that paraspeckles might coordinate the regulation of these gene clusters by controlling the activity of certain transcriptional condensates that coregulate multiple genes. We propose that our findings highlight a potential role for Neat1 and paraspeckles in modulating chromosomal organization and gene expression, potentially crucial processes for the differentiation of beige adipocytes.
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Affiliation(s)
- Hikaru Toya
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Yuko Okamatsu-Ogura
- Laboratory of Biochemistry, Faculty of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan
| | - Saori Yokoi
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Misuzu Kurihara
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Mari Mito
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
| | - Shintaro Iwasaki
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
| | - Tetsuro Hirose
- RNA Biofunction Laboratory, Graduate School of Frontier Biosciences, Osaka University, Suita 565-0871, Japan
| | - Shinichi Nakagawa
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
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Kehinde IO, Oduro-Kwateng E, Soliman MES. Allosteric covalent inhibition of TOE1 as potential unexplored anti-cancer target: structure-based virtual screening and covalent molecular dynamics analysis. J Recept Signal Transduct Res 2024; 44:97-106. [PMID: 39377533 DOI: 10.1080/10799893.2024.2411690] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2024] [Revised: 09/24/2024] [Accepted: 09/28/2024] [Indexed: 10/09/2024]
Abstract
Cancer remains a formidable challenge in therapeutic development owing to its complex molecular mechanisms and resistance to conventional treatments. Recent evidence suggests that TOE1 may play a role in cancer progression, making it an attractive target for therapeutic interventions, nevertheless, very limited research in literature has explored the potential of TOE1 inhibitors as anti-cancer. Herein, by exploring a library of 13,900 cysteine-targeted covalent inhibitors via a comprehensive virtual screening process, we sought to identify potential compounds that could be developed into effective cancer therapies against TOE1. The compounds were first screened based on their binding affinity, followed by their compliance with drug-like properties, and finally, by their effective covalent modeling to a reactive cysteine (Cys80). A total of 66 compounds, 28 compounds, and 3 compounds were found to have higher binding affinities, optimum drug-likeness, and higher covalent docking scores, respectively, than the reference compound. The top three screened compounds, 0462, 2204, and 7034, demonstrated favorable interaction profiles, covalent binding dynamics, free binding energetics, and per-residue energy contributions as compared to the reference compound. Notably, compound 0462 contributed to the highest free binding energy and significantly enhanced the stability and rigidity of TOE1, while restricting residue flexibility. This study provides an account of the molecular mechanics underpinning the covalent inhibition of TOE1, while providing a compelling case for further investigation and translation of the screened TOE1 inhibitors, particularly compound 0462, as novel therapeutics against cancer.
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Affiliation(s)
- Ibrahim Oluwatobi Kehinde
- Molecular Bio-computation and Drug Design Laboratory, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
| | - Ernest Oduro-Kwateng
- Molecular Bio-computation and Drug Design Laboratory, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
| | - Mahmoud E S Soliman
- Molecular Bio-computation and Drug Design Laboratory, School of Health Sciences, University of KwaZulu-Natal, Durban, South Africa
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9
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Deng B, Wan G. Technologies for studying phase-separated biomolecular condensates. ADVANCED BIOTECHNOLOGY 2024; 2:10. [PMID: 39883284 PMCID: PMC11740866 DOI: 10.1007/s44307-024-00020-0] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/23/2024] [Revised: 02/26/2024] [Accepted: 02/27/2024] [Indexed: 01/31/2025]
Abstract
Biomolecular condensates, also referred to as membrane-less organelles, function as fundamental organizational units within cells. These structures primarily form through liquid-liquid phase separation, a process in which proteins and nucleic acids segregate from the surrounding milieu to assemble into micron-scale structures. By concentrating functionally related proteins and nucleic acids, these biomolecular condensates regulate a myriad of essential cellular processes. To study these significant and intricate organelles, a range of technologies have been either adapted or developed. In this review, we provide an overview of the most utilized technologies in this rapidly evolving field. These include methods used to identify new condensates, explore their components, investigate their properties and spatiotemporal regulation, and understand the organizational principles governing these condensates. We also discuss potential challenges and review current advancements in applying the principles of biomolecular condensates to the development of new technologies, such as those in synthetic biology.
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Affiliation(s)
- Boyuan Deng
- Guangdong Provincial Key Laboratory of Pharmaceutical Functional Genes, MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, GuangZhou, GuangDong, China
| | - Gang Wan
- Guangdong Provincial Key Laboratory of Pharmaceutical Functional Genes, MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-Sen University, GuangZhou, GuangDong, China.
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10
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Ledoux N, Lelong EIJ, Simard A, Hussein S, Adjibade P, Lambert JP, Mazroui R. The Identification of Nuclear FMRP Isoform Iso6 Partners. Cells 2023; 12:2807. [PMID: 38132127 PMCID: PMC10742089 DOI: 10.3390/cells12242807] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2023] [Revised: 12/02/2023] [Accepted: 12/05/2023] [Indexed: 12/23/2023] Open
Abstract
A deficiency of FMRP, a canonical RNA-binding protein, causes the development of Fragile X Syndrome (FXS), which is characterised by multiple phenotypes, including neurodevelopmental disorders, intellectual disability, and autism. Due to the alternative splicing of the encoding FMR1 gene, multiple FMRP isoforms are produced consisting of full-length predominantly cytoplasmic (i.e., iso1) isoforms involved in translation and truncated nuclear (i.e., iso6) isoforms with orphan functions. However, we recently implicated nuclear FMRP isoforms in DNA damage response, showing that they negatively regulate the accumulation of anaphase DNA genomic instability bridges. This finding provided evidence that the cytoplasmic and nuclear functions of FMRP are uncoupled played by respective cytoplasmic and nuclear isoforms, potentially involving specific interactions. While interaction partners of cytoplasmic FMRP have been reported, the identity of nuclear FMRP isoform partners remains to be established. Using affinity purification coupled with mass spectrometry, we mapped the nuclear interactome of the FMRP isoform iso6 in U2OS. In doing so, we found FMRP nuclear interaction partners to be involved in RNA processing, pre-mRNA splicing, ribosome biogenesis, DNA replication and damage response, chromatin remodeling and chromosome segregation. By comparing interactions between nuclear iso6 and cytoplasmic iso1, we report a set of partners that bind specifically to the nuclear isoforms, mainly proteins involved in DNA-associated processes and proteasomal proteins, which is consistent with our finding that proteasome targets the nuclear FMRP iso6. The specific interactions with the nuclear isoform 6 are regulated by replication stress, while those with the cytoplasmic isoform 1 are largely insensitive to such stress, further supporting a specific role of nuclear isoforms in DNA damage response induced by replicative stress, potentially regulated by the proteasome.
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Affiliation(s)
- Nassim Ledoux
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
| | - Emeline I. J. Lelong
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
| | - Alexandre Simard
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
| | - Samer Hussein
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
| | - Pauline Adjibade
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
| | - Jean-Philippe Lambert
- Centre de Recherche du CHU de Québec—Université Laval, Axe Endocrinologie et néphrologie, Département de Médecine Moléculaire, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada;
- PROTEO, Le Regroupement Québécois De Recherche Sur La Fonction, L’ingénierie et Les Applications des Protéines, Université Laval, Québec, QC G1V 0A6, Canada
| | - Rachid Mazroui
- Centre de Recherche du CHU de Québec—Université Laval, Axe Oncologie, Département de Biologie Moléculaire, Biochimie Médicale et Pathologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada; (N.L.); (E.I.J.L.); (A.S.); (S.H.); (P.A.)
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11
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Lettin L, Erbay B, Blair GE. Viruses and Cajal Bodies: A Critical Cellular Target in Virus Infection? Viruses 2023; 15:2311. [PMID: 38140552 PMCID: PMC10747631 DOI: 10.3390/v15122311] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2023] [Revised: 11/22/2023] [Accepted: 11/23/2023] [Indexed: 12/24/2023] Open
Abstract
Nuclear bodies (NBs) are dynamic structures present in eukaryotic cell nuclei. They are not bounded by membranes and are often considered biomolecular condensates, defined structurally and functionally by the localisation of core components. Nuclear architecture can be reorganised during normal cellular processes such as the cell cycle as well as in response to cellular stress. Many plant and animal viruses target their proteins to NBs, in some cases triggering their structural disruption and redistribution. Although not all such interactions have been well characterised, subversion of NBs and their functions may form a key part of the life cycle of eukaryotic viruses that require the nucleus for their replication. This review will focus on Cajal bodies (CBs) and the viruses that target them. Since CBs are dynamic structures, other NBs (principally nucleoli and promyelocytic leukaemia, PML and bodies), whose components interact with CBs, will also be considered. As well as providing important insights into key virus-host cell interactions, studies on Cajal and associated NBs may identify novel cellular targets for development of antiviral compounds.
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Affiliation(s)
- Lucy Lettin
- School of Molecular and Cellular Biology, University of Leeds, Leeds LS2 9JT, UK (B.E.)
| | - Bilgi Erbay
- School of Molecular and Cellular Biology, University of Leeds, Leeds LS2 9JT, UK (B.E.)
- Moleküler Biyoloji ve Genetik Bölümü, Fen Fakültesi, Van Yuzuncu Yil University, Van 65140, Türkiye
| | - G. Eric Blair
- School of Molecular and Cellular Biology, University of Leeds, Leeds LS2 9JT, UK (B.E.)
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12
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Zhu J, Yang J, Wen H, Wang M, Zheng X, Zhao J, Sun X, Yang P, Mao Q, Li Y, Xia H. Expression and functional analysis of fam76b in zebrafish. FISH & SHELLFISH IMMUNOLOGY 2023; 142:109161. [PMID: 37838209 DOI: 10.1016/j.fsi.2023.109161] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/29/2023] [Revised: 10/08/2023] [Accepted: 10/11/2023] [Indexed: 10/16/2023]
Abstract
FAM76B is nuclear speckle-localized protein with a molecular weight of 39 kDa. The amino sequence of FAM76B protein is highly conserved among species, suggesting that FAM76B has important biological functions. However, the biological function of FAM76B is currently still unclear. To explore the biological function of FAM76B, we firstly used zebrafish as the experimental model to study the distribution and expression level of Fam76b. The results indicated that fam76b is highly expressed in hematopoiesis and immune systems of zebrafish by real-time quantitative PCR, in situ hybridization and Tg(fam76b: eGFP) transgenic zebrafish. Then, the fam76b gene was knocked out by CRISPR/Cas9 in zebrafish and fam76b rescue in fam76b-/- zebrafish was performed using the TOL2 transposable system. fam76b gene knockout zebrafish exhibit reduced thymus, excessive inflammatory response, and increased mortality. FAM76B was further found to be involved in regulating the development of hematopoiesis and immune system, and participate in the process of inflammatory response. Our findings in the study lay the groundwork for elucidating the function of the new molecule Fam76b and provide new insights into the development of zebrafish hematopoietic and immune system.
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Affiliation(s)
- Jiuling Zhu
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China; School of Basic Medical Sciences, Wannan Medical College, 22 Wenchang West Road, Wuhu, 241002, Anhui, PR China
| | - Jiahang Yang
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - He Wen
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Mengtian Wang
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Xiaojing Zheng
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Junli Zhao
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Xiaohong Sun
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Peiyan Yang
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China
| | - Qinwen Mao
- Department of Pathology, University of Utah, 2000 Circle of Hope Drive, Salt Lake City, UT, 84112, USA
| | - Yu Li
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China.
| | - Haibin Xia
- Laboratory of Gene Therapy, Department of Biochemistry, College of Life Sciences, Shaanxi Normal University, 199 South Chang'an Road, Xi'an, 710062, Shaanxi, PR China.
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13
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Valverde JM, Dubra G, Phillips M, Haider A, Elena-Real C, Fournet A, Alghoul E, Chahar D, Andrés-Sanchez N, Paloni M, Bernadó P, van Mierlo G, Vermeulen M, van den Toorn H, Heck AJR, Constantinou A, Barducci A, Ghosh K, Sibille N, Knipscheer P, Krasinska L, Fisher D, Altelaar M. A cyclin-dependent kinase-mediated phosphorylation switch of disordered protein condensation. Nat Commun 2023; 14:6316. [PMID: 37813838 PMCID: PMC10562473 DOI: 10.1038/s41467-023-42049-0] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2023] [Accepted: 09/28/2023] [Indexed: 10/11/2023] Open
Abstract
Cell cycle transitions result from global changes in protein phosphorylation states triggered by cyclin-dependent kinases (CDKs). To understand how this complexity produces an ordered and rapid cellular reorganisation, we generated a high-resolution map of changing phosphosites throughout unperturbed early cell cycles in single Xenopus embryos, derived the emergent principles through systems biology analysis, and tested them by biophysical modelling and biochemical experiments. We found that most dynamic phosphosites share two key characteristics: they occur on highly disordered proteins that localise to membraneless organelles, and are CDK targets. Furthermore, CDK-mediated multisite phosphorylation can switch homotypic interactions of such proteins between favourable and inhibitory modes for biomolecular condensate formation. These results provide insight into the molecular mechanisms and kinetics of mitotic cellular reorganisation.
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Affiliation(s)
- Juan Manuel Valverde
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, 3584 CH, Utrecht, Netherlands
- Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, Netherlands
| | - Geronimo Dubra
- IGMM, CNRS, University of Montpellier, INSERM, Montpellier, France
- Equipe Labellisée LIGUE 2018, Ligue Nationale Contre le Cancer, Paris, France
| | - Michael Phillips
- Department of Physics and Astronomy, University of Denver, Denver, Co, 80208, USA
| | - Austin Haider
- Department of Molecular and Cellular Biophysics, University of Denver, 80208, Denver, Co, USA
| | | | - Aurélie Fournet
- CBS, CNRS, University of Montpellier, INSERM, Montpellier, France
| | - Emile Alghoul
- IGH, CNRS, University of Montpellier, Montpellier, France
| | - Dhanvantri Chahar
- IGMM, CNRS, University of Montpellier, INSERM, Montpellier, France
- Equipe Labellisée LIGUE 2018, Ligue Nationale Contre le Cancer, Paris, France
| | - Nuria Andrés-Sanchez
- IGMM, CNRS, University of Montpellier, INSERM, Montpellier, France
- Equipe Labellisée LIGUE 2018, Ligue Nationale Contre le Cancer, Paris, France
| | - Matteo Paloni
- Department of Physics and Astronomy, University of Denver, Denver, Co, 80208, USA
| | - Pau Bernadó
- CBS, CNRS, University of Montpellier, INSERM, Montpellier, France
| | - Guido van Mierlo
- Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Oncode Institute, Radboud University Nijmegen, Nijmegen, 6525 GA, The Netherlands
| | - Michiel Vermeulen
- Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Oncode Institute, Radboud University Nijmegen, Nijmegen, 6525 GA, The Netherlands
| | - Henk van den Toorn
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, 3584 CH, Utrecht, Netherlands
- Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, Netherlands
| | - Albert J R Heck
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, 3584 CH, Utrecht, Netherlands
- Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, Netherlands
| | | | | | - Kingshuk Ghosh
- Department of Physics and Astronomy, University of Denver, Denver, Co, 80208, USA
- Department of Molecular and Cellular Biophysics, University of Denver, 80208, Denver, Co, USA
| | - Nathalie Sibille
- CBS, CNRS, University of Montpellier, INSERM, Montpellier, France
| | - Puck Knipscheer
- Oncode Institute, Hubrecht Institute-KNAW and University Medical Center, Utrecht, 3584 CT, Netherlands
| | - Liliana Krasinska
- IGMM, CNRS, University of Montpellier, INSERM, Montpellier, France
- Equipe Labellisée LIGUE 2018, Ligue Nationale Contre le Cancer, Paris, France
| | - Daniel Fisher
- IGMM, CNRS, University of Montpellier, INSERM, Montpellier, France.
- Equipe Labellisée LIGUE 2018, Ligue Nationale Contre le Cancer, Paris, France.
| | - Maarten Altelaar
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, 3584 CH, Utrecht, Netherlands.
- Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, Netherlands.
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14
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Huynh TN, Parker R. The PARN, TOE1, and USB1 RNA deadenylases and their roles in non-coding RNA regulation. J Biol Chem 2023; 299:105139. [PMID: 37544646 PMCID: PMC10493513 DOI: 10.1016/j.jbc.2023.105139] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2023] [Revised: 07/28/2023] [Accepted: 07/29/2023] [Indexed: 08/08/2023] Open
Abstract
The levels of non-coding RNAs (ncRNAs) are regulated by transcription, RNA processing, and RNA degradation pathways. One mechanism for the degradation of ncRNAs involves the addition of oligo(A) tails by non-canonical poly(A) polymerases, which then recruit processive sequence-independent 3' to 5' exonucleases for RNA degradation. This pathway of decay is also regulated by three 3' to 5' exoribonucleases, USB1, PARN, and TOE1, which remove oligo(A) tails and thereby can protect ncRNAs from decay in a manner analogous to the deubiquitination of proteins. Loss-of-function mutations in these genes lead to premature degradation of some ncRNAs and lead to specific human diseases such as Poikiloderma with Neutropenia (PN) for USB1, Dyskeratosis Congenita (DC) for PARN and Pontocerebellar Hypoplasia type 7 (PCH7) for TOE1. Herein, we review the biochemical properties of USB1, PARN, and TOE1, how they modulate ncRNA levels, and their roles in human diseases.
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Affiliation(s)
- Thao Ngoc Huynh
- Department of Biochemistry, University of Colorado Boulder, Boulder, Colorado, USA
| | - Roy Parker
- Department of Biochemistry, University of Colorado Boulder, Boulder, Colorado, USA; Howard Hughes Medical Institute, Chevy Chase, Maryland, USA.
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15
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Fierro C, Gatti V, La Banca V, De Domenico S, Scalera S, Corleone G, Fanciulli M, De Nicola F, Mauriello A, Montanaro M, Calin GA, Melino G, Peschiaroli A. The long non-coding RNA NEAT1 is a ΔNp63 target gene modulating epidermal differentiation. Nat Commun 2023; 14:3795. [PMID: 37365156 PMCID: PMC10293300 DOI: 10.1038/s41467-023-39011-5] [Citation(s) in RCA: 13] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2022] [Accepted: 05/25/2023] [Indexed: 06/28/2023] Open
Abstract
The transcription factor ΔNp63 regulates epithelial stem cell function and maintains the integrity of stratified epithelial tissues by acting as transcriptional repressor or activator towards a distinct subset of protein-coding genes and microRNAs. However, our knowledge of the functional link between ∆Np63 transcriptional activity and long non-coding RNAs (lncRNAs) expression is quite limited. Here, we show that in proliferating human keratinocytes ∆Np63 represses the expression of the lncRNA NEAT1 by recruiting the histone deacetylase HDAC1 to the proximal promoter of NEAT1 genomic locus. Upon induction of differentiation, ∆Np63 down-regulation is associated by a marked increase of NEAT1 RNA levels, resulting in an increased assembly of paraspeckles foci both in vitro and in human skin tissues. RNA-seq analysis associated with global DNA binding profile (ChIRP-seq) revealed that NEAT1 associates with the promoter of key epithelial transcription factors sustaining their expression during epidermal differentiation. These molecular events might explain the inability of NEAT1-depleted keratinocytes to undergo the proper formation of epidermal layers. Collectively, these data uncover the lncRNA NEAT1 as an additional player of the intricate network orchestrating epidermal morphogenesis.
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Affiliation(s)
- Claudia Fierro
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
- Translational Pediatrics and Clinical Genetics Research Division, Bambino Gesù Children's Hospital, IRCSS, Piazza Sant'Onofrio, 4, Rome, Italy
| | - Veronica Gatti
- Institute of Translational Pharmacology (IFT), CNR, Via Fosso del Cavaliere 100, 00133, Rome, Italy
| | - Veronica La Banca
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
| | - Sara De Domenico
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
| | - Stefano Scalera
- UOSD SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, IRCCS Regina Elena National Cancer Institute, Rome, Italy
| | - Giacomo Corleone
- UOSD SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, IRCCS Regina Elena National Cancer Institute, Rome, Italy
| | - Maurizio Fanciulli
- UOSD SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, IRCCS Regina Elena National Cancer Institute, Rome, Italy
| | - Francesca De Nicola
- UOSD SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, IRCCS Regina Elena National Cancer Institute, Rome, Italy
| | - Alessandro Mauriello
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
| | - Manuela Montanaro
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
| | - George A Calin
- Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, 77030, USA
- Center for RNA Interference and Non-Coding RNAs, The University of Texas MD Anderson Cancer Center, Houston, TX, 77030, USA
| | - Gerry Melino
- Department of Experimental Medicine, Tor Vergata Oncoscience Research (TOR), University of Rome "Tor Vergata", Via Montpellier 1, 00133, Rome, Italy
| | - Angelo Peschiaroli
- Institute of Translational Pharmacology (IFT), CNR, Via Fosso del Cavaliere 100, 00133, Rome, Italy.
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16
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Choi HJ, Lee JY, Kim K. Glutathionylation on RNA-binding proteins: a regulator of liquid‒liquid phase separation in the pathogenesis of amyotrophic lateral sclerosis. Exp Mol Med 2023; 55:735-744. [PMID: 37009800 PMCID: PMC10167235 DOI: 10.1038/s12276-023-00978-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Revised: 01/09/2023] [Accepted: 01/11/2023] [Indexed: 04/04/2023] Open
Abstract
RNA-binding proteins (RBPs) containing low-sequence complexity domains mediate the formation of cellular condensates and membrane-less organelles with biological functions via liquid‒liquid phase separation (LLPS). However, the abnormal phase transition of these proteins induces the formation of insoluble aggregates. Aggregates are pathological hallmarks of neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS). The molecular mechanisms underlying aggregate formation by ALS-associated RPBs remain largely unknown. This review highlights emerging studies on various posttranslational modifications (PTMs) related to protein aggregation. We begin with the introduction of several ALS-associated RBPs that form aggregates induced by phase separation. In addition, we highlight our recent discovery of a new PTM involved in the phase transition during the pathogenesis of fused-in-sarcoma (FUS)-associated ALS. We suggest a molecular mechanism through which LLPS mediates glutathionylation in FUS-linked ALS. This review aims to provide a detailed overview of the key molecular mechanisms of LLPS-mediated aggregate formation by PTMs, which will help further the understanding of the pathogenesis and development of ALS therapeutics.
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Affiliation(s)
- Hyun-Jun Choi
- Soonchunhyang Institute of Medi-bio Science, Soonchunhyang University, Cheonan, 31151, Korea
- Department of Integrated Biomedical Sciences, Soonchunhyang University, Cheonan, 31151, Korea
| | - Ji Young Lee
- Department of Medical Biotechnology, Soonchunhyang University, Asan, 31538, Korea
- Department of Medical Science, Soonchunhyang University, Asan, 31538, Korea
| | - Kiyoung Kim
- Department of Medical Science, Soonchunhyang University, Asan, 31538, Korea.
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17
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Voss PG, Wang JL. Liquid-liquid phase separation: Galectin-3 in nuclear speckles and ribonucleoprotein complexes. Exp Cell Res 2023; 427:113571. [PMID: 37003559 DOI: 10.1016/j.yexcr.2023.113571] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2023] [Revised: 03/19/2023] [Accepted: 03/24/2023] [Indexed: 04/03/2023]
Abstract
Nuclear speckles are subcellular structures originally characterized by punctate immunofluorescence staining of the monoclonal antibody SC35, which recognizes an epitope on SRRM2 (serine/arginine repetitive matrix protein 2) and Sfrs2, a member of the SR (serine/arginine-rich) family of splicing factors. Galectin-3 co-localizes with SC35 in nuclear speckles, which represent one group of nuclear bodies that include the nucleolus, Cajal bodies and gems, paraspeckles, etc. Although they appear to have well-delineated physical boundaries, these nuclear bodies are not membrane-bound structures but represent macromolecular assemblies arising from a phenomenon called liquid-liquid phase separation. There has been much recent interest in liquid phase condensation as a newly recognized mechanism by which a cell can organize and compartmentalize subcellular structures with distinct composition. The punctate/speckled staining of galectin-3 with SC3 demonstrates their co-localization in a phase-separated body in vivo, under conditions endogenous to the cell. The purpose of the present review is to summarize the studies that document three key features of galectin-3 for its localization in liquid phase condensates: (a) an intrinsically disordered domain; (b) oligomer formation for multivalent binding; and (c) association with RNA and ribonucleoprotein complexes.
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Affiliation(s)
- Patricia G Voss
- Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, 48824, USA
| | - John L Wang
- Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, 48824, USA.
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18
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Che X, Wu J, Liu H, Su J, Chen X. Cellular liquid-liquid phase separation: Concept, functions, regulations, and detections. J Cell Physiol 2023; 238:847-865. [PMID: 36870067 DOI: 10.1002/jcp.30980] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2022] [Revised: 10/05/2022] [Accepted: 02/08/2023] [Indexed: 03/06/2023]
Abstract
Liquid-liquid phase separation is a multicomponent system separated into phases with different compositions and structures. It has been identified and explored in organisms after being introduced from the thermodynamic field. Condensate, the product of phase separation, exists in different scales of cellular structures, such as nucleolus, stress granules, and other organelles in nuclei or cytoplasm. And also play critical roles in different cellular behaviors. Here, we review the concept, thermodynamical and biochemical principles of phase separation. We summarized the main functions including the adjustment of biochemical reaction rates, the regulation of macromolecule folding state, subcellular structural support, the mediation of subcellular location, and intimately linked to different kinds of diseases, such as cancer and neurodegeneration. Advanced detection methods to investigate phase separation are collected and analyzed. We conclude with the discussion of anxiety of phase separation, and thought about how progress can be made to develop precise detection methods and disclose the potential application of condensates.
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Affiliation(s)
- Xuanlin Che
- Department of Dermatology, Xiangya Hospital, Central South University, Changsha, Hunan, China.,Hunan Key Laboratory of Skin Cancer and Psoriasis, Xiangya Hospital, Changsha, Hunan, China.,Hunan Engineering Research Center of Skin Health and Disease, Xiangya Hospital, Changsha, Hunan, China.,Xiangya Clinical Research Center for Cancer Immunotherapy, Central South University, Changsha, Hunan, China
| | - Jiajun Wu
- Department of Orthopaedics, Xiangya Hospital, Central South University, Changsha, China
| | - Hua Liu
- Department of Orthopaedics, Xiangya Hospital, Central South University, Changsha, China
| | - Juan Su
- Department of Dermatology, Xiangya Hospital, Central South University, Changsha, Hunan, China.,Hunan Key Laboratory of Skin Cancer and Psoriasis, Xiangya Hospital, Changsha, Hunan, China.,Hunan Engineering Research Center of Skin Health and Disease, Xiangya Hospital, Changsha, Hunan, China.,Xiangya Clinical Research Center for Cancer Immunotherapy, Central South University, Changsha, Hunan, China
| | - Xiang Chen
- Department of Dermatology, Xiangya Hospital, Central South University, Changsha, Hunan, China.,Hunan Key Laboratory of Skin Cancer and Psoriasis, Xiangya Hospital, Changsha, Hunan, China.,Hunan Engineering Research Center of Skin Health and Disease, Xiangya Hospital, Changsha, Hunan, China.,Xiangya Clinical Research Center for Cancer Immunotherapy, Central South University, Changsha, Hunan, China
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19
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Bcl10 phosphorylation-dependent droplet-like condensation positively regulates DNA virus-induced innate immune signaling. SCIENCE CHINA. LIFE SCIENCES 2023; 66:283-297. [PMID: 36115893 DOI: 10.1007/s11427-022-2169-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/27/2022] [Accepted: 07/20/2022] [Indexed: 10/14/2022]
Abstract
B-cell lymphoma 10 (Bcl10) is a scaffolding protein that functions as an upstream regulator of NF-κB signaling by forming a complex with Mucosa-associated lymphoid tissue lymphoma translocation protein 1 (Malt1) and CARD-coiled coil protein family. This study showed that Bcl10 was involved in type I interferon (IFN) expression in response to DNA virus infection and that Bcl10-deficient mice were more susceptible to Herpes simplex virus 1 (HSV-1) infection than control mice. Mechanistically, DNA virus infection can trigger Bcl10 recruitment to the STING-TBK1 complex, leading to Bcl10 phosphorylation by TBK1. The phosphorylated Bcl10 undergoes droplet-like condensation and forms oligomers, which induce TBK1 phosphorylation and translocation to the perinuclear region. The activated TBK1 phosphorylates IRF3, which induces the expression of type I IFNs. This study elucidates that Bcl10 induces an innate immune response by undergoing droplet-like condensation and participating in signalosome formation downstream of the cGAS-STING pathway.
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20
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Wang H, Ming X, Zhang S, Chen J, Liu X, Wu X, Zhang S, Zhang Y, Cui W, Li W, Liu Y. Knockdown of Toe1 causes developmental arrest during the morula-to-blastocyst transition in mice. Theriogenology 2022; 194:154-161. [PMID: 36257135 DOI: 10.1016/j.theriogenology.2022.10.011] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2022] [Revised: 10/06/2022] [Accepted: 10/07/2022] [Indexed: 11/04/2022]
Abstract
The target of EGR1 protein 1 (TOE1) is evolutionarily conserved from Caenorhabditis elegans to mammals, which plays a critical role in the maturation of a variety of small nuclear RNAs. Mutation in human TOE1 has been reported to cause pontocerebellar hypoplasia type 7, a severe neurodegenerative syndrome. However, the role of TOE1 in early embryonic development remains unclear. Herein, we found that Toe1 mRNA and protein were expressed in mouse preimplantation embryos. Silencing Toe1 by siRNA led to morula-to-blastocyst transition failure. This developmental arrest can be rescued by Toe1 mRNA microinjection. EdU incorporation assay showed a defect in blastomere proliferation within developmentally arrested embryos. Further studies revealed that Toe1 knockdown caused increased signals for γH2AX and micronuclei, indicative of sustained DNA damage. Moreover, mRNA levels of cell cycle inhibitor p21 were significantly upregulated in Toe1 knockdown embryos before developmental arrest. Together, these results suggest that TOE1 is indispensable for mouse early embryo development potentially through maintaining genomic integrity. Our findings provide further insight into the role of TOE1 in mouse preimplantation embryonic development.
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Affiliation(s)
- Hongcheng Wang
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China; Linquan Modern Agricultural Technology Cooperation and Extension Service Center, The Anhui Agricultural University's Comprehensive Experimental Station in the Northwest of Anhui Province, Linquan, Anhui, 236400, China
| | - Xin Ming
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Shengnan Zhang
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Ji Chen
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Xinli Liu
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Xiaoqing Wu
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Shangrong Zhang
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China
| | - Yunhai Zhang
- Linquan Modern Agricultural Technology Cooperation and Extension Service Center, The Anhui Agricultural University's Comprehensive Experimental Station in the Northwest of Anhui Province, Linquan, Anhui, 236400, China
| | - Wei Cui
- Department of Veterinary and Animal Sciences, Animal Models Core Facility, Institute for Applied Life Sciences (IALS), University of Massachusetts, Amherst, MA, 01002, USA
| | - Wenyong Li
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China.
| | - Yong Liu
- Anhui Province Key Laboratory of Embryo Development and Reproductive Regulation, Anhui Province Key Laboratory of Environmental Hormone and Reproduction, Fuyang Normal University, Fuyang City, Anhui Province, 236037, China; Department of Veterinary and Animal Sciences, Animal Models Core Facility, Institute for Applied Life Sciences (IALS), University of Massachusetts, Amherst, MA, 01002, USA.
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21
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A brief guideline for studies of phase-separated biomolecular condensates. Nat Chem Biol 2022; 18:1307-1318. [DOI: 10.1038/s41589-022-01204-2] [Citation(s) in RCA: 88] [Impact Index Per Article: 29.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2022] [Accepted: 10/10/2022] [Indexed: 11/20/2022]
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22
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Ilık İA, Aktaş T. Nuclear speckles: dynamic hubs of gene expression regulation. FEBS J 2022; 289:7234-7245. [PMID: 34245118 DOI: 10.1111/febs.16117] [Citation(s) in RCA: 64] [Impact Index Per Article: 21.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2021] [Revised: 06/13/2021] [Accepted: 07/08/2021] [Indexed: 01/13/2023]
Abstract
Complex, multistep biochemical reactions that routinely take place in our cells require high concentrations of enzymes, substrates, and other structural components to proceed efficiently and typically require chemical environments that can inhibit other reactions in their immediate vicinity. Eukaryotic cells solve these problems by restricting such reactions into diffusion-restricted compartments within the cell called organelles that can be separated from their environment by a lipid membrane, or into membrane-less compartments that form through liquid-liquid phase separation (LLPS). One of the most easily noticeable and the earliest discovered organelle is the nucleus, which harbors the genetic material in cells where transcription by RNA polymerases produces most of the messenger RNAs and a plethora of noncoding RNAs, which in turn are required for translation of mRNAs in the cytoplasm. The interior of the nucleus is not a uniform soup of biomolecules and rather consists of a variety of membrane-less bodies, such as the nucleolus, nuclear speckles (NS), paraspeckles, Cajal bodies, histone locus bodies, and more. In this review, we will focus on NS with an emphasis on recent developments including our own findings about the formation of NS by two large IDR-rich proteins SON and SRRM2.
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Affiliation(s)
| | - Tuğçe Aktaş
- Max Planck Institute for Molecular Genetics, Berlin, Germany
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23
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Kim CG, Hwang DE, Kumar R, Chung M, Eom YG, Kim H, Koo DH, Choi JM. Recent trends in studies of biomolecular phase separation. BMB Rep 2022. [PMID: 35880435 PMCID: PMC9442351 DOI: 10.5483/bmbrep.2022.55.8.101] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Biomolecular phase separation has recently attracted broad in-terest, due to its role in the spatiotemporal compartmentalization of living cells. It governs the formation, regulation, and dissociation of biomolecular condensates, which play multiple roles in vivo, from activating specific biochemical reactions to organizing chromatin. Interestingly, biomolecular phase separation seems to be a mainly passive process, which can be ex-plained by relatively simple physical principles and reproduced in vitro with a minimal set of components. This Mini review focuses on our current understanding of the fundamental principles of biomolecular phase separation and the recent progress in the research on this topic.
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Affiliation(s)
- Chan-Geun Kim
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Da-Eun Hwang
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Rajeev Kumar
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Min Chung
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Yu-Gon Eom
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Hyunji Kim
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Da-Hyun Koo
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Jeong-Mo Choi
- Department of Chemistry, Pusan National University, Busan 46241, Korea
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24
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Yamazaki T, Yamamoto T, Hirose T. Micellization: A new principle in the formation of biomolecular condensates. Front Mol Biosci 2022; 9:974772. [PMID: 36106018 PMCID: PMC9465675 DOI: 10.3389/fmolb.2022.974772] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2022] [Accepted: 07/20/2022] [Indexed: 11/18/2022] Open
Abstract
Phase separation is a fundamental mechanism for compartmentalization in cells and leads to the formation of biomolecular condensates, generally containing various RNA molecules. RNAs are biomolecules that can serve as suitable scaffolds for biomolecular condensates and determine their forms and functions. Many studies have focused on biomolecular condensates formed by liquid-liquid phase separation (LLPS), one type of intracellular phase separation mechanism. We recently identified that paraspeckle nuclear bodies use an intracellular phase separation mechanism called micellization of block copolymers in their formation. The paraspeckles are scaffolded by NEAT1_2 long non-coding RNAs (lncRNAs) and their partner RNA-binding proteins (NEAT1_2 RNA-protein complexes [RNPs]). The NEAT1_2 RNPs act as block copolymers and the paraspeckles assemble through micellization. In LLPS, condensates grow without bound as long as components are available and typically have spherical shapes to minimize surface tension. In contrast, the size, shape, and internal morphology of the condensates are more strictly controlled in micellization. Here, we discuss the potential importance and future perspectives of micellization of block copolymers of RNPs in cells, including the construction of designer condensates with optimal internal organization, shape, and size according to design guidelines of block copolymers.
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Affiliation(s)
- Tomohiro Yamazaki
- Graduate School of Frontier Biosciences, Osaka University, Suita, Japan
| | - Tetsuya Yamamoto
- Institute for Chemical Reaction Design and Discovery, Hokkaido University, Sapporo, Japan
| | - Tetsuro Hirose
- Graduate School of Frontier Biosciences, Osaka University, Suita, Japan
- Institute for Open and Transdisciplinary Research Initiatives (OTRI), Osaka University, Suita, Japan
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25
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Raskó T, Pande A, Radscheit K, Zink A, Singh M, Sommer C, Wachtl G, Kolacsek O, Inak G, Szvetnik A, Petrakis S, Bunse M, Bansal V, Selbach M, Orbán TI, Prigione A, Hurst LD, Izsvák Z. A Novel Gene Controls a New Structure: PiggyBac Transposable Element-Derived 1, Unique to Mammals, Controls Mammal-Specific Neuronal Paraspeckles. Mol Biol Evol 2022; 39:6661922. [PMID: 36205081 PMCID: PMC9538788 DOI: 10.1093/molbev/msac175] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Although new genes can arrive from modes other than duplication, few examples are well characterized. Given high expression in some human brain subregions and a putative link to psychological disorders [e.g., schizophrenia (SCZ)], suggestive of brain functionality, here we characterize piggyBac transposable element-derived 1 (PGBD1). PGBD1 is nonmonotreme mammal-specific and under purifying selection, consistent with functionality. The gene body of human PGBD1 retains much of the original DNA transposon but has additionally captured SCAN and KRAB domains. Despite gene body retention, PGBD1 has lost transposition abilities, thus transposase functionality is absent. PGBD1 no longer recognizes piggyBac transposon-like inverted repeats, nonetheless PGBD1 has DNA binding activity. Genome scale analysis identifies enrichment of binding sites in and around genes involved in neuronal development, with association with both histone activating and repressing marks. We focus on one of the repressed genes, the long noncoding RNA NEAT1, also dysregulated in SCZ, the core structural RNA of paraspeckles. DNA binding assays confirm specific binding of PGBD1 both in the NEAT1 promoter and in the gene body. Depletion of PGBD1 in neuronal progenitor cells (NPCs) results in increased NEAT1/paraspeckles and differentiation. We conclude that PGBD1 has evolved core regulatory functionality for the maintenance of NPCs. As paraspeckles are a mammal-specific structure, the results presented here show a rare example of the evolution of a novel gene coupled to the evolution of a contemporaneous new structure.
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Affiliation(s)
- Tamás Raskó
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | | | | | - Annika Zink
- Department of General Pediatrics, Neonatology and Pediatric Cardiology, Medical Faculty, Heinrich Heine University, Duesseldorf, Germany
| | - Manvendra Singh
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | - Christian Sommer
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | - Gerda Wachtl
- Institute of Enzymology, Research Centre for Natural Sciences, ELKH, Budapest, Hungary,Doctoral School of Biology, Institute of Biology, ELTE Eötvös Loránd University, Budapest, Hungary
| | - Orsolya Kolacsek
- Institute of Enzymology, Research Centre for Natural Sciences, ELKH, Budapest, Hungary
| | - Gizem Inak
- Department of General Pediatrics, Neonatology and Pediatric Cardiology, Medical Faculty, Heinrich Heine University, Duesseldorf, Germany
| | - Attila Szvetnik
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | - Spyros Petrakis
- Institute of Applied Biosciences/Centre for Research and Technology Hellas, 57001 Thessaloniki, Greece
| | - Mario Bunse
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | - Vikas Bansal
- Biomedical Data Science and Machine Learning Group, German Center for Neurodegenerative Diseases, Tübingen 72076, Germany
| | - Matthias Selbach
- Max Delbrück Center for Molecular Medicine in the Helmholtz Society, Berlin, Germany
| | - Tamás I Orbán
- Institute of Enzymology, Research Centre for Natural Sciences, ELKH, Budapest, Hungary
| | - Alessandro Prigione
- Department of General Pediatrics, Neonatology and Pediatric Cardiology, Medical Faculty, Heinrich Heine University, Duesseldorf, Germany
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26
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Muñoz-Díaz E, Sáez-Vásquez J. Nuclear dynamics: Formation of bodies and trafficking in plant nuclei. FRONTIERS IN PLANT SCIENCE 2022; 13:984163. [PMID: 36082296 PMCID: PMC9445803 DOI: 10.3389/fpls.2022.984163] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/01/2022] [Accepted: 08/04/2022] [Indexed: 06/01/2023]
Abstract
The existence of the nucleus distinguishes prokaryotes and eukaryotes. Apart from containing most of the genetic material, the nucleus possesses several nuclear bodies composed of protein and RNA molecules. The nucleus is separated from the cytoplasm by a double membrane, regulating the trafficking of molecules in- and outwards. Here, we investigate the composition and function of the different plant nuclear bodies and molecular clues involved in nuclear trafficking. The behavior of the nucleolus, Cajal bodies, dicing bodies, nuclear speckles, cyclophilin-containing bodies, photobodies and DNA damage foci is analyzed in response to different abiotic stresses. Furthermore, we research the literature to collect the different protein localization signals that rule nucleocytoplasmic trafficking. These signals include the different types of nuclear localization signals (NLSs) for nuclear import, and the nuclear export signals (NESs) for nuclear export. In contrast to these unidirectional-movement signals, the existence of nucleocytoplasmic shuttling signals (NSSs) allows bidirectional movement through the nuclear envelope. Likewise, nucleolar signals are also described, which mainly include the nucleolar localization signals (NoLSs) controlling nucleolar import. In contrast, few examples of nucleolar export signals, called nucleoplasmic localization signals (NpLSs) or nucleolar export signals (NoESs), have been reported. The existence of consensus sequences for these localization signals led to the generation of prediction tools, allowing the detection of these signals from an amino acid sequence. Additionally, the effect of high temperatures as well as different post-translational modifications in nuclear and nucleolar import and export is discussed.
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Affiliation(s)
- Eduardo Muñoz-Díaz
- Centre National de la Recherche Scientifique (CNRS), Laboratoire Génome et Développement des Plantes, UMR 5096, Perpignan, France
- Univ. Perpignan Via Domitia, Laboratoire Génome et Développement des Plantes, UMR 5096, Perpignan, France
| | - Julio Sáez-Vásquez
- Centre National de la Recherche Scientifique (CNRS), Laboratoire Génome et Développement des Plantes, UMR 5096, Perpignan, France
- Univ. Perpignan Via Domitia, Laboratoire Génome et Développement des Plantes, UMR 5096, Perpignan, France
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27
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Yamada A, Toya H, Tanahashi M, Kurihara M, Mito M, Iwasaki S, Kurosaka S, Takumi T, Fox A, Kawamura Y, Miura K, Nakagawa S. Species-specific formation of paraspeckles in intestinal epithelium revealed by characterization of NEAT1 in naked mole-rat. RNA (NEW YORK, N.Y.) 2022; 28:1128-1143. [PMID: 35654483 PMCID: PMC9297846 DOI: 10.1261/rna.079135.122] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/17/2022] [Accepted: 05/30/2022] [Indexed: 06/15/2023]
Abstract
Paraspeckles are mammalian-specific nuclear bodies built on the long noncoding RNA NEAT1_2 The molecular mechanisms of paraspeckle formation have been mainly studied using human or mouse cells, and it is not known if the same molecular components are involved in the formation of paraspeckles in other mammalian species. We thus investigated the expression pattern of NEAT1_2 in naked mole-rats (nNEAT1_2), which exhibit extreme longevity and lower susceptibility to cancer. In the intestine, nNEAT1_2 is widely expressed along the entire intestinal epithelium, which is different from the expression of mNeat1_2 that is restricted to the cells of the distal tip in mice. Notably, the expression of FUS, a FET family RNA binding protein, essential for the formation of paraspeckles both in humans and mice, was absent in the distal part of the intestinal epithelium in naked mole-rats. Instead, mRNAs of other FET family proteins EWSR1 and TAF15 were expressed in the distal region. Exogenous expression of these proteins in Fus-deficient murine embryonic fibroblast cells rescued the formation of paraspeckles. These observations suggest that nNEAT1_2 recruits a different set of RNA binding proteins in a cell type-specific manner during the formation of paraspeckles in different organisms.
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Affiliation(s)
- Akihiro Yamada
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Hikaru Toya
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Mayuko Tanahashi
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Misuzu Kurihara
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Mari Mito
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
| | - Shintaro Iwasaki
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
- Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba 277-8561, Japan
| | | | - Toru Takumi
- RIKEN Brain Science Institute, Saitama 351-0198, Japan
- Department of Physiology and Cell Biology, Kobe University School of Medicine, Kobe 670-0017, Japan
| | - Archa Fox
- School of Human Sciences, University of Western Australia, Crawley, Western Australia 6009, Australia
- Harry Perkins Institute of Medical Research, Nedlands, Western Australia 6009, Australia
- School of Molecular Sciences, University of Western Australia, Crawley, Western Australia 6009, Australia
| | - Yoshimi Kawamura
- Department of Aging and Longevity Research, Faculty of Life Sciences, Kumamoto University, Kumamoto 860-8556, Japan
- Center for Metabolic Regulation of Healthy Aging, Kumamoto University, Kumamoto 860-8556, Japan
| | - Kyoko Miura
- Department of Aging and Longevity Research, Faculty of Life Sciences, Kumamoto University, Kumamoto 860-8556, Japan
- Center for Metabolic Regulation of Healthy Aging, Kumamoto University, Kumamoto 860-8556, Japan
| | - Shinichi Nakagawa
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
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28
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Kim CG, Hwang DE, Kumar R, Chung M, Eom YG, Kim H, Koo DH, Choi JM. Recent trends in studies of biomolecular phase separation. BMB Rep 2022; 55:363-369. [PMID: 35880435 PMCID: PMC9442351] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2022] [Revised: 07/07/2022] [Accepted: 07/20/2022] [Indexed: 03/08/2024] Open
Abstract
Biomolecular phase separation has recently attracted broad interest, due to its role in the spatiotemporal compartmentalization of living cells. It governs the formation, regulation, and dissociation of biomolecular condensates, which play multiple roles in vivo, from activating specific biochemical reactions to organizing chromatin. Interestingly, biomolecular phase separation seems to be a mainly passive process, which can be explained by relatively simple physical principles and reproduced in vitro with a minimal set of components. This Mini review focuses on our current understanding of the fundamental principles of biomolecular phase separation and the recent progress in the research on this topic. [BMB Reports 2022; 55(8): 363-369].
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Affiliation(s)
- Chan-Geun Kim
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Da-Eun Hwang
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Rajeev Kumar
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Min Chung
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Yu-Gon Eom
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Hyunji Kim
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Da-Hyun Koo
- Department of Chemistry, Pusan National University, Busan 46241, Korea
| | - Jeong-Mo Choi
- Department of Chemistry, Pusan National University, Busan 46241, Korea
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29
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Faber GP, Nadav-Eliyahu S, Shav-Tal Y. Nuclear speckles - a driving force in gene expression. J Cell Sci 2022; 135:275909. [PMID: 35788677 DOI: 10.1242/jcs.259594] [Citation(s) in RCA: 42] [Impact Index Per Article: 14.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Nuclear speckles are dynamic membraneless bodies located in the cell nucleus. They harbor RNAs and proteins, many of which are splicing factors, that together display complex biophysical properties dictating nuclear speckle formation and maintenance. Although these nuclear bodies were discovered decades ago, only recently has in-depth genomic analysis begun to unravel their essential functions in modulation of gene activity. Major advancements in genomic mapping techniques combined with microscopy approaches have enabled insights into the roles nuclear speckles may play in enhancing gene expression, and how gene positioning to specific nuclear landmarks can regulate gene expression and RNA processing. Some studies have drawn a link between nuclear speckles and disease. Certain maladies either involve nuclear speckles directly or dictate the localization and reorganization of many nuclear speckle factors. This is most striking during viral infection, as viruses alter the entire nuclear architecture and highjack host machinery. As discussed in this Review, nuclear speckles represent a fascinating target of study not only to reveal the links between gene positioning, genome subcompartments and gene activity, but also as a potential target for therapeutics.
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Affiliation(s)
- Gabriel P Faber
- The Mina and Everard Goodman Faculty of Life Sciences , Bar-Ilan University, Ramat Gan 5290002, Israel.,Institute of Nanotechnology , Bar-Ilan University, Ramat Gan 5290002, Israel
| | - Shani Nadav-Eliyahu
- The Mina and Everard Goodman Faculty of Life Sciences , Bar-Ilan University, Ramat Gan 5290002, Israel.,Institute of Nanotechnology , Bar-Ilan University, Ramat Gan 5290002, Israel
| | - Yaron Shav-Tal
- The Mina and Everard Goodman Faculty of Life Sciences , Bar-Ilan University, Ramat Gan 5290002, Israel.,Institute of Nanotechnology , Bar-Ilan University, Ramat Gan 5290002, Israel
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30
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Shan H, Liu T, Gan H, He S, Deng J, Hu X, Li L, Cai L, He J, Long H, Cai J, Li H, Zhang Q, Wang L, Chen F, Chen Y, Zhang H, Li J, Yang L, Liu Y, Yang J, Kuang DM, Pang P, He H. RNA helicase DDX24 stabilizes LAMB1 to promote hepatocellular carcinoma progression. Cancer Res 2022; 82:3074-3087. [PMID: 35763670 DOI: 10.1158/0008-5472.can-21-3748] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2021] [Revised: 04/22/2022] [Accepted: 06/23/2022] [Indexed: 11/16/2022]
Abstract
Hepatocellular carcinoma (HCC) is one of the most aggressive malignancies. Elucidating the underlying mechanisms of this disease could provide new therapeutic strategies for treating HCC. Here, we identified a novel role of DEAD-box helicase 24 (DDX24), a member of the DEAD-box protein family, in promoting HCC progression. DDX24 levels were significantly elevated in HCC tissues and were associated with poor prognosis of HCC. Overexpression of DDX24 promoted HCC migration and proliferation in vitro and in vivo, whereas suppression of DDX24 inhibited both functions. Mechanistically, DDX24 bound the mRNA618-624nt of laminin subunit beta 1 (LAMB1) and increased its stability in a manner dependent upon the interaction between nucleolin (NCL) and the C-terminal region of DDX24. Moreover, RFX8 was identified as a DDX24 promoter-binding protein that transcriptionally upregulated DDX24 expression. Collectively, these findings demonstrate that the RFX8/DDX24/LAMB1 axis promotes HCC progression, providing potential therapeutic targets for HCC.
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Affiliation(s)
- Hong Shan
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, Guangdong, China
| | - Tianze Liu
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Hairun Gan
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Simeng He
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Jia Deng
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Xinyan Hu
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Luting Li
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Li Cai
- MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory for Biocontrol, School of Life Sciences, Sun Yat-sen University, China
| | - JianZhong He
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Haoyu Long
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Jianxun Cai
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Hanjie Li
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Qianqian Zhang
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Lijie Wang
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Fangbin Chen
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Yuming Chen
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Haopei Zhang
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Jian Li
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Lukun Yang
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Ye Liu
- Sun Yat-sen University 5th Hospital, Zhuhai, Guangdong province, China
| | | | - Dong-Ming Kuang
- Sun Yat-sen University, Guangzhou, Outside the United States or C, China
| | - Pengfei Pang
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
| | - Huanhuan He
- Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China
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31
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Expression and functions of long non-coding RNA NEAT1 and isoforms in breast cancer. Br J Cancer 2022; 126:551-561. [PMID: 34671127 PMCID: PMC8854383 DOI: 10.1038/s41416-021-01588-3] [Citation(s) in RCA: 40] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2021] [Revised: 09/08/2021] [Accepted: 10/05/2021] [Indexed: 02/07/2023] Open
Abstract
NEAT1 is a highly abundant nuclear architectural long non-coding RNA. There are two overlapping NEAT1 isoforms, NEAT1_1 and NEAT1_2, of which the latter is an essential scaffold for the assembly of a class of nuclear ribonucleoprotein bodies called paraspeckles. Paraspeckle formation is elevated by a wide variety of cellular stressors and in certain developmental processes, either through transcriptional upregulation of the NEAT1 gene or through a switch from NEAT1_1 to NEAT1_2 isoform production. In such conditions, paraspeckles modulate cellular processes by sequestering proteins or RNA molecules. NEAT1 is abnormally expressed in many cancers and a growing body of evidence suggests that, in many cases, high NEAT1 levels are associated with therapy resistance and poor clinical outcome. Here we review the current knowledge of NEAT1 expression and functions in breast cancer, highlighting its established role in postnatal mammary gland development. We will discuss possible isoform-specific roles of NEAT1_1 and NEAT1_2 in different breast cancer subtypes, which critically needs to be considered when studying NEAT1 and breast cancer.
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Zheng J, Wu Z, Qiu Y, Wang X, Jiang X. An integrative multi-omics analysis based on liquid–liquid phase separation delineates distinct subtypes of lower-grade glioma and identifies a prognostic signature. J Transl Med 2022; 20:55. [PMID: 35093128 PMCID: PMC8800244 DOI: 10.1186/s12967-022-03266-1] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2021] [Accepted: 01/17/2022] [Indexed: 12/11/2022] Open
Abstract
Abstract
Background
Emerging evidences have indicated that the aberrant liquid–liquid phase separation (LLPS) leads to the dysfunction of biomolecular condensates, thereby contributing to the tumorigenesis and progression. Nevertheless, it remains unclear whether or how the LLPS of specific molecules affects the prognosis and tumor immune microenvironment (TIME) of patients with lower-grade glioma (LGG).
Methods
We integrated the transcriptome information of 3585 LLPS-related genes to comprehensively evaluate the LLPS patterns of 423 patients with LGG in The Cancer Genome Atlas (TCGA) cohort. Then, we systematically demonstrated the differences among four LLPS subtypes based on multi-omics analyses. In addition, we constructed the LLPS-related prognostic risk score (LPRS) for individualized integrative assessment.
Results
Based on the expression profiles of 85 scaffolds, 355 regulators, and 3145 clients in LGG, we identified four LLPS subtypes, namely LS1, LS2, LS3 and LS4.
We confirmed that there were significant differences in prognosis, clinicopathological features, cancer hallmarks, genomic alterations, TIME patterns and immunotherapeutic responses among four LLPS subtypes. In addition, a prognostic signature called LPRS was constructed for individualized integrative assessment. LPRS exhibited a robust predictive capacity for prognosis of LGG patients in multiple cohorts. Moreover, LPRS was found to be correlated with clinicopathological features, cancer hallmarks, genomic alterations and TIME patterns of LGG patients. The predictive power of LPRS in response to immune checkpoint inhibitor (ICI) therapy was also prominent.
Conclusions
This study provided a novel classification of LGG patients based on LLPS. The constructed LPRS might facilitate individualized prognosis prediction and better immunotherapy options for LGG patients.
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Rota IA, Handel AE, Maio S, Klein F, Dhalla F, Deadman ME, Cheuk S, Newman JA, Michaels YS, Zuklys S, Prevot N, Hublitz P, Charles PD, Gkazi AS, Adamopoulou E, Qasim W, Davies EG, Hanson I, Pagnamenta AT, Camps C, Dreau HM, White A, James K, Fischer R, Gileadi O, Taylor JC, Fulga T, Lagerholm BC, Anderson G, Sezgin E, Holländer GA. FOXN1 forms higher-order nuclear condensates displaced by mutations causing immunodeficiency. SCIENCE ADVANCES 2021; 7:eabj9247. [PMID: 34860543 PMCID: PMC8641933 DOI: 10.1126/sciadv.abj9247] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/16/2021] [Accepted: 10/15/2021] [Indexed: 05/04/2023]
Abstract
The transcription factor FOXN1 is a master regulator of thymic epithelial cell (TEC) development and function. Here, we demonstrate that FOXN1 expression is differentially regulated during organogenesis and participates in multimolecular nuclear condensates essential for the factor’s transcriptional activity. FOXN1’s C-terminal sequence regulates the diffusion velocity within these aggregates and modulates the binding to proximal gene regulatory regions. These dynamics are altered in a patient with a mutant FOXN1 that is modified in its C-terminal sequence. This mutant is transcriptionally inactive and acts as a dominant negative factor displacing wild-type FOXN1 from condensates and causing athymia and severe lymphopenia in heterozygotes. Expression of the mutated mouse ortholog selectively impairs mouse TEC differentiation, revealing a gene dose dependency for individual TEC subtypes. We have therefore identified the cause for a primary immunodeficiency disease and determined the mechanism by which this FOXN1 gain-of-function mutant mediates its dominant negative effect.
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Affiliation(s)
- Ioanna A. Rota
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Adam E. Handel
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
- Nuffield Department of Clinical Neurosciences, University of Oxford, Oxford, UK
| | - Stefano Maio
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Fabian Klein
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Fatima Dhalla
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Mary E. Deadman
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Stanley Cheuk
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Joseph A. Newman
- Structural Genomics Consortium, University of Oxford, ORCRB, Roosevelt Drive, Oxford, UK
| | - Yale S. Michaels
- Genome Engineering and Synthetic Biology Unit, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Saulius Zuklys
- Paediatric Immunology, Department of Biomedicine, University of Basel and University Children’s Hospital Basel, Basel, Switzerland
| | - Nicolas Prevot
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Philip Hublitz
- MRC Weatherall Institute of Molecular Medicine, Genome engineering services, Radcliffe Department of Medicine, University of Oxford, Oxford, UK
| | - Philip D. Charles
- Target Discovery Institute, University of Oxford, Oxford OX3 7FZ, UK
| | - Athina Soragia Gkazi
- Great Ormond Street Hospital and Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
| | - Eleni Adamopoulou
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Waseem Qasim
- Great Ormond Street Hospital and Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
| | - Edward Graham Davies
- Great Ormond Street Hospital and Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
| | - Imelda Hanson
- Department of Pediatrics, Section of Pediatric Immunology, Allergy, and Retrovirology, Baylor College of Medicine, Houston, TX, USA
| | - Alistair T. Pagnamenta
- National Institute for Health Research Biomedical Research Centre, Oxford, UK
- Wellcome Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK
| | - Carme Camps
- National Institute for Health Research Biomedical Research Centre, Oxford, UK
- Wellcome Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK
| | - Helene M. Dreau
- Department of Oncology, University of Oxford, Oxford OX3 7DQ, UK
| | - Andrea White
- Institute for Immunology and Immunotherapy, Medical School, University of Birmingham, Birmingham B15 2TT, UK
| | - Kieran James
- Institute for Immunology and Immunotherapy, Medical School, University of Birmingham, Birmingham B15 2TT, UK
| | - Roman Fischer
- Target Discovery Institute, University of Oxford, Oxford OX3 7FZ, UK
| | - Opher Gileadi
- Structural Genomics Consortium, University of Oxford, ORCRB, Roosevelt Drive, Oxford, UK
| | - Jenny C. Taylor
- National Institute for Health Research Biomedical Research Centre, Oxford, UK
- Wellcome Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK
| | - Tudor Fulga
- Genome Engineering and Synthetic Biology Unit, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - B. Christoffer Lagerholm
- Wolfson Imaging Centre Oxford, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Headley Way, Oxford OX3 9DS, UK
| | - Graham Anderson
- Institute for Immunology and Immunotherapy, Medical School, University of Birmingham, Birmingham B15 2TT, UK
| | - Erdinc Sezgin
- Paediatric Immunology, Department of Biomedicine, University of Basel and University Children’s Hospital Basel, Basel, Switzerland
- MRC Human Immunology Unit, MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
| | - Georg A. Holländer
- Department of Paediatrics and the MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK
- Paediatric Immunology, Department of Biomedicine, University of Basel and University Children’s Hospital Basel, Basel, Switzerland
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
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34
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Kachaev ZM, Ivashchenko SD, Kozlov EN, Lebedeva LA, Shidlovskii YV. Localization and Functional Roles of Components of the Translation Apparatus in the Eukaryotic Cell Nucleus. Cells 2021; 10:3239. [PMID: 34831461 PMCID: PMC8623629 DOI: 10.3390/cells10113239] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2021] [Revised: 11/11/2021] [Accepted: 11/16/2021] [Indexed: 12/15/2022] Open
Abstract
Components of the translation apparatus, including ribosomal proteins, have been found in cell nuclei in various organisms. Components of the translation apparatus are involved in various nuclear processes, particularly those associated with genome integrity control and the nuclear stages of gene expression, such as transcription, mRNA processing, and mRNA export. Components of the translation apparatus control intranuclear trafficking; the nuclear import and export of RNA and proteins; and regulate the activity, stability, and functional recruitment of nuclear proteins. The nuclear translocation of these components is often involved in the cell response to stimulation and stress, in addition to playing critical roles in oncogenesis and viral infection. Many components of the translation apparatus are moonlighting proteins, involved in integral cell stress response and coupling of gene expression subprocesses. Thus, this phenomenon represents a significant interest for both basic and applied molecular biology. Here, we provide an overview of the current data regarding the molecular functions of translation factors and ribosomal proteins in the cell nucleus.
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Affiliation(s)
- Zaur M. Kachaev
- Department of Gene Expression Regulation in Development, Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia; (Z.M.K.); (S.D.I.); (E.N.K.); (L.A.L.)
- Center for Genetics and Life Science, Sirius University of Science and Technology, 354340 Sochi, Russia
| | - Sergey D. Ivashchenko
- Department of Gene Expression Regulation in Development, Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia; (Z.M.K.); (S.D.I.); (E.N.K.); (L.A.L.)
| | - Eugene N. Kozlov
- Department of Gene Expression Regulation in Development, Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia; (Z.M.K.); (S.D.I.); (E.N.K.); (L.A.L.)
| | - Lyubov A. Lebedeva
- Department of Gene Expression Regulation in Development, Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia; (Z.M.K.); (S.D.I.); (E.N.K.); (L.A.L.)
| | - Yulii V. Shidlovskii
- Department of Gene Expression Regulation in Development, Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia; (Z.M.K.); (S.D.I.); (E.N.K.); (L.A.L.)
- Center for Genetics and Life Science, Sirius University of Science and Technology, 354340 Sochi, Russia
- Department of Biology and General Genetics, Sechenov First Moscow State Medical University (Sechenov University), 119992 Moscow, Russia
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35
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Nanjappa DP, Babu N, Khanna-Gupta A, O'Donohue MF, Sips P, Chakraborty A. Poly (A)-specific ribonuclease (PARN): More than just "mRNA stock clearing". Life Sci 2021; 285:119953. [PMID: 34520768 DOI: 10.1016/j.lfs.2021.119953] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2021] [Revised: 09/07/2021] [Accepted: 09/08/2021] [Indexed: 11/24/2022]
Abstract
In eukaryotic cells, the balance between the synthesis and the degradation decides the steady-state levels of messenger RNAs (mRNA). The removal of adenosine residues from the poly(A) tail, called deadenylation, is the first and the most crucial step in the process of mRNA degradation. Poly (A)-specific ribonuclease (PARN) is one such enzyme that catalyses the process of deadenylation. Although PARN has been primarily known as the regulator of the mRNA stability, recent evidence clearly suggests several other functions of PARN, including a role in embryogenesis, oocyte maturation, cell-cycle progression, telomere biology, non-coding RNA maturation and ribosome biogenesis. Also, deregulated PARN activity is shown to be a hallmark of specific disease conditions. Pathogenic variants in the PARN gene have been observed in various cancers and inherited bone marrow failure syndromes. The focus in this review is to highlight the emerging functions of PARN, particularly in the context of human diseases.
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Affiliation(s)
- Dechamma Pandyanda Nanjappa
- Division of Molecular Genetics and Cancer, Nitte University Centre for Science Education and Research (NUCSER), NITTE (Deemed to be University), Deralakate, Mangaluru 575018, India
| | - Nishith Babu
- Division of Molecular Genetics and Cancer, Nitte University Centre for Science Education and Research (NUCSER), NITTE (Deemed to be University), Deralakate, Mangaluru 575018, India
| | - Arati Khanna-Gupta
- Consortium of Rare Genetic and Bone Marrow Disorders, India network@NitteDU, NITTE (Deemed to be University, Deralakatte, Mangaluru, India
| | - Marie-Françoise O'Donohue
- Laboratoire de Biologie Moléculaire Eucaryote, Centre de Biologie Intégrative CBI, Université de Toulouse- CNRS- UPS- Toulouse-, Dynamics and Disorders of Ribosome Synthesis, Toulouse, France
| | - Patrick Sips
- Department of Biomolecular Medicine, Ghent University, Ghent, Belgium
| | - Anirban Chakraborty
- Division of Molecular Genetics and Cancer, Nitte University Centre for Science Education and Research (NUCSER), NITTE (Deemed to be University), Deralakate, Mangaluru 575018, India.
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36
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Orti F, Navarro AM, Rabinovich A, Wodak SJ, Marino-Buslje C. Insight into membraneless organelles and their associated proteins: Drivers, Clients and Regulators. Comput Struct Biotechnol J 2021; 19:3964-3977. [PMID: 34377363 PMCID: PMC8318826 DOI: 10.1016/j.csbj.2021.06.042] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2021] [Revised: 06/26/2021] [Accepted: 06/27/2021] [Indexed: 02/06/2023] Open
Abstract
In recent years, attention has been devoted to proteins forming immiscible liquid phases within the liquid intracellular medium, commonly referred to as membraneless organelles (MLO). These organelles enable the spatiotemporal associations of cellular components that exchange dynamically with the cellular milieu. The dysregulation of these liquid-liquid phase separation processes (LLPS) may cause various diseases including neurodegenerative pathologies and cancer, among others. Until very recently, databases containing information on proteins forming MLOs, as well as tools and resources facilitating their analysis, were missing. This has recently changed with the publication of 4 databases that focus on different types of experiments, sets of proteins, inclusion criteria, and levels of annotation or curation. In this study we integrate and analyze the information across these databases, complement their records, and produce a consolidated set of proteins that enables the investigation of the LLPS phenomenon. To gain insight into the features that characterize different types of MLOs and the roles of their associated proteins, they were grouped into categories: High Confidence MLO associated (including Drivers and reviewed proteins), Potential Clients and Regulators, according to their annotated functions. We show that none of the databases taken alone covers the data sufficiently to enable meaningful analysis, validating our integration effort as essential for gaining better understanding of phase separation and laying the foundations for the discovery of new proteins potentially involved in this important cellular process. Lastly, we developed a server, enabling customized selections of different sets of proteins based on MLO location, database, disorder content, among other attributes (https://forti.shinyapps.io/mlos/).
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Affiliation(s)
- Fernando Orti
- Bioinformatics Unit, Fundación Instituto Leloir. Avda. Patricias Argentinas 435, Buenos Aires B1405WE, Argentina
| | - Alvaro M. Navarro
- Bioinformatics Unit, Fundación Instituto Leloir. Avda. Patricias Argentinas 435, Buenos Aires B1405WE, Argentina
| | - Andres Rabinovich
- Bioinformatics Unit, Fundación Instituto Leloir. Avda. Patricias Argentinas 435, Buenos Aires B1405WE, Argentina
| | - Shoshana J. Wodak
- VIB-VUB Center for Structural Biology, Flemish Institute for Biotechnology, Brussels, Belgium
- Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium
| | - Cristina Marino-Buslje
- Bioinformatics Unit, Fundación Instituto Leloir. Avda. Patricias Argentinas 435, Buenos Aires B1405WE, Argentina
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Norppa AJ, Frilander MJ. The integrity of the U12 snRNA 3' stem-loop is necessary for its overall stability. Nucleic Acids Res 2021; 49:2835-2847. [PMID: 33577674 PMCID: PMC7968993 DOI: 10.1093/nar/gkab048] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2020] [Revised: 01/14/2021] [Accepted: 02/07/2021] [Indexed: 12/20/2022] Open
Abstract
Disruption of minor spliceosome functions underlies several genetic diseases with mutations in the minor spliceosome-specific small nuclear RNAs (snRNAs) and proteins. Here, we define the molecular outcome of the U12 snRNA mutation (84C>U) resulting in an early-onset form of cerebellar ataxia. To understand the molecular consequences of the U12 snRNA mutation, we created cell lines harboring the 84C>T mutation in the U12 snRNA gene (RNU12). We show that the 84C>U mutation leads to accelerated decay of the snRNA, resulting in significantly reduced steady-state U12 snRNA levels. Additionally, the mutation leads to accumulation of 3′-truncated forms of U12 snRNA, which have undergone the cytoplasmic steps of snRNP biogenesis. Our data suggests that the 84C>U-mutant snRNA is targeted for decay following reimport into the nucleus, and that the U12 snRNA fragments are decay intermediates that result from the stalling of a 3′-to-5′ exonuclease. Finally, we show that several other single-nucleotide variants in the 3′ stem-loop of U12 snRNA that are segregating in the human population are also highly destabilizing. This suggests that the 3′ stem-loop is important for the overall stability of the U12 snRNA and that additional disease-causing mutations are likely to exist in this region.
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Affiliation(s)
- Antto J Norppa
- Institute of Biotechnology, P.O. Box 56, Viikinkaari 5, University of Helsinki, FI-00014 Helsinki, Finland
| | - Mikko J Frilander
- Institute of Biotechnology, P.O. Box 56, Viikinkaari 5, University of Helsinki, FI-00014 Helsinki, Finland
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Abstract
The HIV-1 Rev protein is a nuclear export factor for unspliced and incompletely spliced HIV-1 RNAs. Without Rev, these intron-retaining RNAs are trapped in the nucleus. A genome-wide screen identified nine proteins of the spliceosome, which all enhanced expression from the HIV-1 unspliced RNA after CRISPR/Cas knockdown. Depletion of DHX38, WDR70, and four proteins of the Prp19-associated complex (ISY1, BUD31, XAB2, and CRNKL1) resulted in a more than 20-fold enhancement of unspliced HIV-1 RNA levels in the cytoplasm. Targeting of CRNKL1, DHX38, and BUD31 affected nuclear export efficiencies of the HIV-1 unspliced RNA to a much larger extent than splicing. Transcriptomic analyses further revealed that CRNKL1 also suppresses cytoplasmic levels of a subset of cellular mRNAs, including some with selectively retained introns. Thus, CRNKL1-dependent nuclear retention is a novel cellular mechanism for the regulation of cytoplasmic levels of intron-retaining HIV-1 mRNAs, which HIV-1 may have harnessed to direct its complex splicing pattern.IMPORTANCE To regulate its complex splicing pattern, HIV-1 uses the adaptor protein Rev to shuttle unspliced or partially spliced mRNA from the nucleus to the cytoplasm. In the absence of Rev, these RNAs are retained in the nucleus, but it is unclear why. Here we identify cellular proteins whose depletion enhances cytoplasmic levels of the HIV-1 unspliced RNA. Depletion of one of them, CRNKL1, also increases cytoplasmic levels of a subset of intron-retaining cellular mRNA, suggesting that CRNKL1-dependent nuclear retention may be a basic cellular mechanism exploited by HIV-1.
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Christy ATR, Kusumaatmaja H, Miller MA. Control of Superselectivity by Crowding in Three-Dimensional Hosts. PHYSICAL REVIEW LETTERS 2021; 126:028002. [PMID: 33512214 DOI: 10.1103/physrevlett.126.028002] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/06/2020] [Accepted: 12/09/2020] [Indexed: 06/12/2023]
Abstract
Motivated by the fine compositional control observed in membraneless droplet organelles in cells, we investigate how a sharp binding-unbinding transition can occur between multivalent client molecules and receptors embedded in a porous three-dimensional structure. In contrast to similar superselective binding previously observed at surfaces, we have identified that a key effect in a three-dimensional environment is that the presence of inert crowding agents can significantly enhance or even introduce superselectivity. In essence, molecular crowding initially suppresses binding via an entropic penalty, but the clients can then more easily form many bonds simultaneously. We demonstrate the robustness of the superselective behavior with respect to client valency, linker length, and binding interactions in Monte Carlo simulations of an archetypal lattice polymer model.
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Affiliation(s)
- Andrew T R Christy
- Department of Chemistry, Durham University, South Road, Durham DH1 3LE, United Kingdom
| | - Halim Kusumaatmaja
- Department of Physics, Durham University, South Road, Durham DH1 3LE, United Kingdom
| | - Mark A Miller
- Department of Chemistry, Durham University, South Road, Durham DH1 3LE, United Kingdom
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40
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Hardenberg M, Horvath A, Ambrus V, Fuxreiter M, Vendruscolo M. Widespread occurrence of the droplet state of proteins in the human proteome. Proc Natl Acad Sci U S A 2020; 117:33254-33262. [PMID: 33318217 PMCID: PMC7777240 DOI: 10.1073/pnas.2007670117] [Citation(s) in RCA: 208] [Impact Index Per Article: 41.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
A wide range of proteins have been reported to condensate into a dense liquid phase, forming a reversible droplet state. Failure in the control of the droplet state can lead to the formation of the more stable amyloid state, which is often disease-related. These observations prompt the question of how many proteins can undergo liquid-liquid phase separation. Here, in order to address this problem, we discuss the biophysical principles underlying the droplet state of proteins by analyzing current evidence for droplet-driver and droplet-client proteins. Based on the concept that the droplet state is stabilized by the large conformational entropy associated with nonspecific side-chain interactions, we develop the FuzDrop method to predict droplet-promoting regions and proteins, which can spontaneously phase separate. We use this approach to carry out a proteome-level study to rank proteins according to their propensity to form the droplet state, spontaneously or via partner interactions. Our results lead to the conclusion that the droplet state could be, at least transiently, accessible to most proteins under conditions found in the cellular environment.
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Affiliation(s)
- Maarten Hardenberg
- Centre for Misfolding Diseases, Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom
| | - Attila Horvath
- The John Curtin School of Medical Research, The Australian National University, Canberra, ACT 2601, Australia
| | - Viktor Ambrus
- Laboratory of Protein Dynamics, Department of Biochemistry and Molecular Biology, University of Debrecen, H-4010 Debrecen, Hungary
| | - Monika Fuxreiter
- Laboratory of Protein Dynamics, Department of Biochemistry and Molecular Biology, University of Debrecen, H-4010 Debrecen, Hungary;
- Department of Biomedical Sciences, University of Padova, 35131 Padova, Italy
| | - Michele Vendruscolo
- Centre for Misfolding Diseases, Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom;
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41
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Organization and function of paraspeckles. Essays Biochem 2020; 64:875-882. [DOI: 10.1042/ebc20200010] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2020] [Revised: 07/11/2020] [Accepted: 07/14/2020] [Indexed: 12/11/2022]
Abstract
Abstract
Paraspeckles are a type of subnuclear bodies built on the long noncoding RNA NEAT1 (nuclear paraspeckle assembly transcript 1, also known as MEN-ε/β or VINC-1). Paraspeckles are involved in many physiological processes including cellular stress responses, cell differentiation, corpus luteum formation and cancer progression. Recently, ultra-resolution microscopy coupled with multicolor-labeling of paraspeckle components (the NEAT1 RNA and paraspeckle proteins) revealed the exquisite details of paraspeckle structure and function. NEAT1 transcripts are radially arranged to form a core–shell spheroidal structure, while paraspeckle proteins (PSPs) localize within different layers. Functional dissection of NEAT1 shows that the subdomains of NEAT1_2 are important for RNA stability, isoform switching and paraspeckle assembly via a liquid–liquid phase separation (LLPS) mechanism. We review recent progress on structure and organization of paraspeckles as well as how paraspeckles spatiotemporally control gene regulation through sequestration of diverse proteins and RNAs in cells.
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42
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Yu C, Shen B, You K, Huang Q, Shi M, Wu C, Chen Y, Zhang C, Li T. Proteome-scale analysis of phase-separated proteins in immunofluorescence images. Brief Bioinform 2020; 22:5900570. [PMID: 34020549 DOI: 10.1093/bib/bbaa187] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2020] [Revised: 07/16/2020] [Accepted: 07/21/2020] [Indexed: 12/25/2022] Open
Abstract
Phase separation is an important mechanism that mediates the spatial distribution of proteins in different cellular compartments. While phase-separated proteins share certain sequence characteristics, including intrinsically disordered regions (IDRs) and prion-like domains, such characteristics are insufficient for making accurate predictions; thus, a proteome-wide understanding of phase separation is currently lacking. Here, we define phase-separated proteomes based on the systematic analysis of immunofluorescence images of 12 073 proteins in the Human Protein Atlas. The analysis of these proteins reveals that phase-separated candidate proteins exhibit higher IDR contents, higher mean net charge and lower hydropathy and prefer to bind to RNA. Kinases and transcription factors are also enriched among these candidate proteins. Strikingly, both phase-separated kinases and phase-separated transcription factors display significantly reduced substrate specificity. Our work provides the first global view of the phase-separated proteome and suggests that the spatial proximity resulting from phase separation reduces the requirement for motif specificity and expands the repertoire of substrates. The source code and data are available at https://github.com/cheneyyu/deepphase.
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Affiliation(s)
- Chunyu Yu
- Department of Biomedical Informatics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Boyan Shen
- Department of Biomedical Informatics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Kaiqiang You
- Department of Biomedical Informatics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Qi Huang
- Department of Biomedical Informatics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Minglei Shi
- MOE Key Laboratory of Bioinformatics; Bioinformatics Division and Center for Synthetic & Systems Biology, TNLIST; School of Medicine, Tsinghua University, Beijing, China
| | - Congying Wu
- Institute of Systems Biomedicine, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
| | - Yang Chen
- MOE Key Laboratory of Bioinformatics; Bioinformatics Division and Center for Synthetic & Systems Biology, TNLIST; School of Medicine, Tsinghua University, Beijing, China
| | - Chaolin Zhang
- Department of Systems Biology, Department of Biochemistry and Molecular Biophysics, Center for Motor Neuron Biology and Disease, Columbia University, New York, USA
| | - Tingting Li
- Department of Biomedical Informatics, School of Basic Medical Sciences, Peking University Health Science Center, Beijing 100191, China
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43
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Lardelli RM, Lykke-Andersen J. Competition between maturation and degradation drives human snRNA 3' end quality control. Genes Dev 2020; 34:989-1001. [PMID: 32499401 PMCID: PMC7328512 DOI: 10.1101/gad.336891.120] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2020] [Accepted: 04/29/2020] [Indexed: 12/11/2022]
Abstract
Polymerases and exonucleases act on 3' ends of nascent RNAs to promote their maturation or degradation but how the balance between these activities is controlled to dictate the fates of cellular RNAs remains poorly understood. Here, we identify a central role for the human DEDD deadenylase TOE1 in distinguishing the fates of small nuclear (sn)RNAs of the spliceosome from unstable genome-encoded snRNA variants. We found that TOE1 promotes maturation of all regular RNA polymerase II transcribed snRNAs of the major and minor spliceosomes by removing posttranscriptional oligo(A) tails, trimming 3' ends, and preventing nuclear exosome targeting. In contrast, TOE1 promotes little to no maturation of tested U1 variant snRNAs, which are instead targeted by the nuclear exosome. These observations suggest that TOE1 is positioned at the center of a 3' end quality control pathway that selectively promotes maturation and stability of regular snRNAs while leaving snRNA variants unprocessed and exposed to degradation in what could be a widespread mechanism of RNA quality control given the large number of noncoding RNAs processed by DEDD deadenylases.
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Affiliation(s)
- Rea M Lardelli
- Division of Biological Sciences, University of California San Diego, La Jolla, California 92093, USA
| | - Jens Lykke-Andersen
- Division of Biological Sciences, University of California San Diego, La Jolla, California 92093, USA
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Sapaly D, Delers P, Coridon J, Salman B, Letourneur F, Dumont F, Lefebvre S. The Small-Molecule Flunarizine in Spinal Muscular Atrophy Patient Fibroblasts Impacts on the Gemin Components of the SMN Complex and TDP43, an RNA-Binding Protein Relevant to Motor Neuron Diseases. Front Mol Biosci 2020; 7:55. [PMID: 32363199 PMCID: PMC7181958 DOI: 10.3389/fmolb.2020.00055] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2019] [Accepted: 03/18/2020] [Indexed: 01/01/2023] Open
Abstract
The motor neurodegenerative disease spinal muscular atrophy (SMA) is caused by alterations of the survival motor neuron 1 (SMN1) gene involved in RNA metabolism. Although the disease mechanisms are not completely elucidated, SMN protein deficiency leads to abnormal small nuclear ribonucleoproteins (snRNPs) assembly responsible for widespread splicing defects. SMN protein localizes in nuclear bodies that are lost in SMA and adult onset amyotrophic lateral sclerosis (ALS) patient cells harboring TDP-43 or FUS/TLS mutations. We previously reported that flunarizine recruits SMN into nuclear bodies and improves the phenotype of an SMA mouse model. However, the precise mode of action remains elusive. Here, a marked reduction of the integral components of the SMN complex is observed in severe SMA patient fibroblast cells. We show that flunarizine increases the protein levels of a subset of components of the SMN-Gemins complex, Gemins2-4, and markedly reduces the RNA and protein levels of the pro-oxydant thioredoxin-interacting protein (TXNIP) encoded by an mRNA target of Gemin5. We further show that SMN deficiency causes a dissociation of the localization of the SMN complex components from the same nuclear bodies. The accumulation of TDP-43 in SMN-positive nuclear bodies is also perturbed in SMA cells. Notably, TDP-43 is found to co-localize with SMN in nuclear bodies of flunarizine-treated SMA cells. Our findings indicate that flunarizine reverses cellular changes caused by SMN deficiency in SMA cells and further support the view of a common pathway in RNA metabolism underlying infantile and adult motor neuron diseases.
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Affiliation(s)
- Delphine Sapaly
- INSERM UMR-S 1124, Toxicité Environnementale, Cibles Thérapeutiques, Signalisation Cellulaire et Biomarqueurs, Campus Saint-Germain-des-Prés, Université de Paris, Paris, France
| | - Perrine Delers
- INSERM UMR-S 1124, Toxicité Environnementale, Cibles Thérapeutiques, Signalisation Cellulaire et Biomarqueurs, Campus Saint-Germain-des-Prés, Université de Paris, Paris, France
| | - Jennifer Coridon
- BioMedTech Facilities INSERM US36 - CNRS UMS 2009, Campus Saint-Germain-des-Prés, Université de Paris, Paris, France
| | - Badih Salman
- INSERM UMR-S 1124, Toxicité Environnementale, Cibles Thérapeutiques, Signalisation Cellulaire et Biomarqueurs, Campus Saint-Germain-des-Prés, Université de Paris, Paris, France
| | | | - Florent Dumont
- Genom'ic Platform, INSERM U1016, Institut Cochin, Paris, France
| | - Suzie Lefebvre
- INSERM UMR-S 1124, Toxicité Environnementale, Cibles Thérapeutiques, Signalisation Cellulaire et Biomarqueurs, Campus Saint-Germain-des-Prés, Université de Paris, Paris, France
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45
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Isobe M, Toya H, Mito M, Chiba T, Asahara H, Hirose T, Nakagawa S. Forced isoform switching of Neat1_1 to Neat1_2 leads to the loss of Neat1_1 and the hyperformation of paraspeckles but does not affect the development and growth of mice. RNA (NEW YORK, N.Y.) 2020; 26:251-264. [PMID: 31822595 PMCID: PMC7025509 DOI: 10.1261/rna.072587.119] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/10/2019] [Accepted: 12/04/2019] [Indexed: 05/27/2023]
Abstract
Neat1 is a long noncoding RNA (lncRNA) that serves as an architectural component of the nuclear bodies known as paraspeckles. Two isoforms of Neat1, the short isoform Neat1_1 and the long isoform Neat1_2, are generated from the same gene locus by alternative 3' processing. Neat1_1 is the most abundant and the best conserved isoform expressed in various cell types, whereas Neat1_2 is expressed in a small population of particular cell types, including the tip cells of the intestinal epithelium. To investigate the physiological significance of isoform switching, we created mutant mice that solely expressed Neat1_2 by deleting the upstream polyadenylation (poly-A) signal (PAS) required for the production of Neat1_1. We observed the loss of Neat1_1 and strong up-regulation of Neat1_2 in various tissues and cells and the subsequent hyperformation of paraspeckles, especially in cells that normally express Neat1_2. However, the mutant mice were born at the expected Mendelian ratios and did not exhibit obvious external and histological abnormalities. These observations suggested that the hyperformation of paraspeckles does not interfere with the development and growth of these animals under normal laboratory conditions.
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Affiliation(s)
- Momo Isobe
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Hikaru Toya
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Mari Mito
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
| | - Tomoki Chiba
- Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo 113-8510, Japan
| | - Hiroshi Asahara
- Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo 113-8510, Japan
| | - Tetsuro Hirose
- Institute for Genetic Medicine, Hokkaido University, Sapporo 060-0815, Japan
| | - Shinichi Nakagawa
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
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46
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Isobe M, Toya H, Mito M, Chiba T, Asahara H, Hirose T, Nakagawa S. Forced isoform switching of Neat1_1 to Neat1_2 leads to the loss of Neat1_1 and the hyperformation of paraspeckles but does not affect the development and growth of mice. RNA (NEW YORK, N.Y.) 2020. [PMID: 31822595 DOI: 10.1101/698068] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/16/2023]
Abstract
Neat1 is a long noncoding RNA (lncRNA) that serves as an architectural component of the nuclear bodies known as paraspeckles. Two isoforms of Neat1, the short isoform Neat1_1 and the long isoform Neat1_2, are generated from the same gene locus by alternative 3' processing. Neat1_1 is the most abundant and the best conserved isoform expressed in various cell types, whereas Neat1_2 is expressed in a small population of particular cell types, including the tip cells of the intestinal epithelium. To investigate the physiological significance of isoform switching, we created mutant mice that solely expressed Neat1_2 by deleting the upstream polyadenylation (poly-A) signal (PAS) required for the production of Neat1_1. We observed the loss of Neat1_1 and strong up-regulation of Neat1_2 in various tissues and cells and the subsequent hyperformation of paraspeckles, especially in cells that normally express Neat1_2. However, the mutant mice were born at the expected Mendelian ratios and did not exhibit obvious external and histological abnormalities. These observations suggested that the hyperformation of paraspeckles does not interfere with the development and growth of these animals under normal laboratory conditions.
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Affiliation(s)
- Momo Isobe
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Hikaru Toya
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
| | - Mari Mito
- RNA Systems Biochemistry Laboratory, RIKEN Cluster for Pioneering Research, Saitama 351-0198, Japan
| | - Tomoki Chiba
- Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo 113-8510, Japan
| | - Hiroshi Asahara
- Department of Systems BioMedicine, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo 113-8510, Japan
| | - Tetsuro Hirose
- Institute for Genetic Medicine, Hokkaido University, Sapporo 060-0815, Japan
| | - Shinichi Nakagawa
- RNA Biology Laboratory, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan
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Yamazaki T, Nakagawa S, Hirose T. Architectural RNAs for Membraneless Nuclear Body Formation. COLD SPRING HARBOR SYMPOSIA ON QUANTITATIVE BIOLOGY 2020; 84:227-237. [PMID: 32019862 DOI: 10.1101/sqb.2019.84.039404] [Citation(s) in RCA: 25] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/10/2023]
Abstract
Long noncoding RNAs (lncRNAs) are fundamental regulators of various cellular processes. A subset of lncRNAs, termed architectural RNAs (arcRNAs), function in the formation and maintenance of phase-separated membraneless organelles in multiple eukaryotic species. These membraneless organelles represent an important type of compartmentalization in the crowded cellular environment and have several distinct features. The NEAT1_2 lncRNA is a well-characterized arcRNA that functions as an essential scaffold of paraspeckle nuclear bodies. Here, we describe the biogenesis of paraspeckles on arcRNAs through phase separation, focusing on the specific functions of multiple NEAT1_2 RNA domains and their partner RNA-binding proteins. Finally, we present an updated model of paraspeckle formation and discuss future perspectives of research into arcRNA-instructed architectures of phase-separated nuclear bodies.
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Affiliation(s)
- Tomohiro Yamazaki
- Institute for Genetic Medicine, Hokkaido University, Sapporo, 060-0815 Japan
| | - Shinichi Nakagawa
- Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, 060-0812 Japan
| | - Tetsuro Hirose
- Institute for Genetic Medicine, Hokkaido University, Sapporo, 060-0815 Japan
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48
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C9-ALS/FTD-linked proline-arginine dipeptide repeat protein associates with paraspeckle components and increases paraspeckle formation. Cell Death Dis 2019; 10:746. [PMID: 31582731 PMCID: PMC6776546 DOI: 10.1038/s41419-019-1983-5] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2019] [Revised: 08/29/2019] [Accepted: 09/16/2019] [Indexed: 12/12/2022]
Abstract
A GGGGCC hexanucleotide repeat expansion in the C9ORF72 gene has been identified as the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. The repeat expansion undergoes unconventional translation to produce five dipeptide repeat proteins (DPRs). Although DPRs are thought to be neurotoxic, the molecular mechanism underlying the DPR-caused neurotoxicity has not been fully elucidated. The current study shows that poly-proline-arginine (poly-PR), the most toxic DPR in vitro, binds to and up-regulates nuclear paraspeckle assembly transcript 1 (NEAT1) that plays an essential role as a scaffold non-coding RNA during the paraspeckle formation. The CRISPR-assisted up-regulation of endogenous NEAT1 causes neurotoxicity. We also show that the poly-PR modulates the function of several paraspeckle-localizing heterogeneous nuclear ribonucleoproteins. Furthermore, dysregulated expression of TAR DNA-binding protein 43 (TDP-43) up-regulates NEAT1 expression and induces neurotoxicity. These results suggest that the increase in the paraspeckle formation may be involved in the poly-PR- and TDP-43-mediated neurotoxicity.
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49
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Simchovitz A, Hanan M, Niederhoffer N, Madrer N, Yayon N, Bennett ER, Greenberg DS, Kadener S, Soreq H. NEAT1 is overexpressed in Parkinson's disease substantia nigra and confers drug-inducible neuroprotection from oxidative stress. FASEB J 2019; 33:11223-11234. [PMID: 31311324 PMCID: PMC6766647 DOI: 10.1096/fj.201900830r] [Citation(s) in RCA: 69] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2019] [Accepted: 06/17/2019] [Indexed: 12/17/2022]
Abstract
Recent reports attribute numerous regulatory functions to the nuclear paraspeckle-forming long noncoding RNA, nuclear enriched assembly transcript 1 (NEAT1), but the implications of its involvement in Parkinson's disease (PD) remain controversial. To address this issue, we assessed NEAT1 expression levels and cell type patterns in the substantia nigra (SN) from 53 donors with and without PD, as well as in interference tissue culture tests followed by multiple in-house and web-available models of PD. PCR quantification identified elevated levels of NEAT1 expression in the PD SN compared with control brains, an elevation that was reproducible across a multitude of disease models. In situ RNA hybridization supported neuron-specific formation of NEAT1-based paraspeckles at the SN and demonstrated coincreases of NEAT1 and paraspeckles in cultured cells under paraquat (PQ)-induced oxidative stress. Furthermore, neuroprotective agents, including fenofibrate and simvastatin, induced NEAT1 up-regulation, whereas RNA interference-mediated depletion of NEAT1 exacerbated death of PQ-exposed cells in a leucine-rich repeat kinase 2-mediated manner. Our findings highlight a novel protective role for NEAT1 in PD and suggest a previously unknown mechanism for the neuroprotective traits of widely used preventive therapeutics.-Simchovitz, A., Hanan, M., Niederhoffer, N., Madrer, N., Yayon, N., Bennett, E. R., Greenberg, D. S., Kadener, S., Soreq, H. NEAT1 is overexpressed in Parkinson's disease substantia nigra and confers drug-inducible neuroprotection from oxidative stress.
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Affiliation(s)
- Alon Simchovitz
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Mor Hanan
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Naomi Niederhoffer
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Nimrod Madrer
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Nadav Yayon
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Estelle R. Bennett
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - David S. Greenberg
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
| | - Sebastian Kadener
- Biology Department, Brandeis University, Waltham, Massachusetts, USA
| | - Hermona Soreq
- Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
- Edmond and Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel
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50
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Son A, Park JE, Kim VN. PARN and TOE1 Constitute a 3' End Maturation Module for Nuclear Non-coding RNAs. Cell Rep 2019; 23:888-898. [PMID: 29669292 DOI: 10.1016/j.celrep.2018.03.089] [Citation(s) in RCA: 56] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2017] [Revised: 12/27/2017] [Accepted: 03/20/2018] [Indexed: 10/17/2022] Open
Abstract
Poly(A)-specific ribonuclease (PARN) and target of EGR1 protein 1 (TOE1) are nuclear granule-associated deadenylases, whose mutations are linked to multiple human diseases. Here, we applied mTAIL-seq and RNA sequencing (RNA-seq) to systematically identify the substrates of PARN and TOE1 and elucidate their molecular functions. We found that PARN and TOE1 do not modulate the length of mRNA poly(A) tails. Rather, they promote the maturation of nuclear small non-coding RNAs (ncRNAs). PARN and TOE1 act redundantly on some ncRNAs, most prominently small Cajal body-specific RNAs (scaRNAs). scaRNAs are strongly downregulated when PARN and TOE1 are compromised together, leading to defects in small nuclear RNA (snRNA) pseudouridylation. They also function redundantly in the biogenesis of telomerase RNA component (TERC), which shares sequence motifs found in H/ACA box scaRNAs. Our findings extend the knowledge of nuclear ncRNA biogenesis, and they provide insights into the pathology of PARN/TOE1-associated genetic disorders whose therapeutic treatments are currently unavailable.
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Affiliation(s)
- Ahyeon Son
- Center for RNA Research, Institute for Basic Science, Seoul 08826, Korea; School of Biological Sciences, Seoul National University, Seoul 08826, Korea
| | - Jong-Eun Park
- Center for RNA Research, Institute for Basic Science, Seoul 08826, Korea; School of Biological Sciences, Seoul National University, Seoul 08826, Korea; Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK
| | - V Narry Kim
- Center for RNA Research, Institute for Basic Science, Seoul 08826, Korea; School of Biological Sciences, Seoul National University, Seoul 08826, Korea.
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