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Liu YB, Liu X, Li XF, Qiao L, Wang HL, Dong YF, Zhang F, Liu Y, Liu HY, Ji ML, Li L, Jiang Q, Lu J. Multifunctional piezoelectric hydrogels under ultrasound stimulation boost chondrogenesis by recruiting autologous stem cells and activating the Ca 2+/CaM/CaN signaling pathway. Bioact Mater 2025; 50:344-363. [PMID: 40297641 PMCID: PMC12036080 DOI: 10.1016/j.bioactmat.2025.04.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2025] [Revised: 03/25/2025] [Accepted: 04/10/2025] [Indexed: 04/30/2025] Open
Abstract
Articular cartilage, owing to the lack of undifferentiated stem cells after injury, faces significant challenges in reconstruction and repair, making it a major clinical challenge. Therefore, there is an urgent need to design a multifunctional hydrogels capable of recruiting autologous stem cells to achieve in situ cartilage regeneration. Here, our study investigated the potential of a piezoelectric hydrogel (Hyd6) for enhancing cartilage regeneration through ultrasound (US) stimulation. Hyd6 has multiple properties including injectability, self-healing capabilities, and piezoelectric characteristics. These properties synergistically promote stem cell chondrogenesis. The fabrication and characterization of Hyd6 revealed its excellent biocompatibility, biodegradability, and electromechanical conversion capabilities. In vitro and in vivo experiments revealed that Hyd6, when combined with US stimulation, significantly promotes the recruitment of autologous stem cells and enhances chondrogenesis by generating electrical signals that promote the influx of Ca2+, activating downstream CaM/CaN signaling pathways and accelerating cartilage formation. An in vivo study in a rabbit model of chondral defects revealed that Hyd6 combined with US treatment significantly improved cartilage regeneration, as evidenced by better integration of the regenerated tissue with the surrounding cartilage, greater collagen type II expression, and improved mechanical properties. The results highlight the potential of Hyd6 as a novel therapeutic approach for treating cartilage injuries, offering a self-powered, noninvasive, and effective strategy for tissue engineering and regenerative medicine.
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Affiliation(s)
- Yu-Bao Liu
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
| | - Xu Liu
- Department of Orthopedics, The Yangzhou Clinical Medical College of Xuzhou Medical University, Yangzhou, 225009, China
- Orthopedics Department, Nanjing Drum Tower Hospital & Group's Suqian Hospital, Affiliated Hospital of Medical School, Nanjing University, Suqian, 223800, China
| | - Xiao-Fei Li
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
| | - Liang Qiao
- Division of Sports Medicine and Adult Reconstructive Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing, 210008, China
| | - Hao-Liang Wang
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
| | - Yue-Fu Dong
- Department of Joint Surgery, The First People's Hospital of Lianyungang City, Lianyungang, 222000, China
| | - Feng Zhang
- Orthopedics Department, Xuyi County People's Hospital, Huai'an, 211700, China
| | - Yang Liu
- Orthopedics Department, Dan Yang Third People's Hospital, Zhenjiang, 212300, China
| | - Hao-Yang Liu
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
| | - Ming-Liang Ji
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
| | - Lan Li
- Division of Sports Medicine and Adult Reconstructive Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing, 210008, China
- Institute of Medical 3D Printing, Nanjing University, Nanjing, 210093, China
| | - Qing Jiang
- Division of Sports Medicine and Adult Reconstructive Surgery, Department of Orthopedic Surgery, Nanjing Drum Tower Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing, 210008, China
- Institute of Medical 3D Printing, Nanjing University, Nanjing, 210093, China
| | - Jun Lu
- The Center of Joint and Sports Medicine, Orthopedics Department, Zhongda Hospital, School of Medicine, Southeast University, Nanjing, 210009, China
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Hao YX, Chen YY, Han X, Wang X, Wu FP, Wen CL, Chen TQ, Tan S, Zheng DD, Hong Y, Shen XY. Genkwanin blocks the interaction between phosphorylated JNK and NFATc1 to promote osteogenic differentiation and collagen Ⅰ α1 production. Eur J Pharmacol 2025; 999:177687. [PMID: 40311832 DOI: 10.1016/j.ejphar.2025.177687] [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: 01/23/2025] [Revised: 04/24/2025] [Accepted: 04/25/2025] [Indexed: 05/03/2025]
Abstract
Genkwanin (GWA), a flavonoid compound found abundantly in various traditional Chinese medicines, has demonstrated pharmacological effectiveness against a range of diseases. However, its role in bone formation and the underlying mechanisms remain to be elucidated. The aim of this study is to investigate the effects of GWA on osteogenic differentiation and to uncover the possible mechanisms. Primary bone marrow-derived mesenchymal stem cells (BMSCs) and MC3T3-E1 cells induced for osteogenic differentiation were used as in vitro cell models. An ovariectomy (OVX) mouse model to induce secondary osteoporosis was used to evaluate the in vivo pharmacologic efficacy of GWA. Our studies revealed that GWA facilitated osteogenic differentiation and bio-mineralization of BMSCs and MC3T3-E1 cells in vitro, and could effectively prevent OVX-induced systematic bone loss and collagen Ⅰ α1 (COL1A1) reduction in vivo. Mechanism studies indicated that GWA could directly bind to phosphorylated c-Jun N-terminal kinase (pJNK) to prevent the phosphorylation of nuclear factor of activated T cells 1 (NFATc1) by JNK, thereby promote osteogenic differentiation and COL1A1 production via increasing the nuclear localization of NFATc1. This is distinct from the previously recognized function of the JNK/AP-1/NFATc1 signaling pathway in activating osteoclast differentiation. More importantly, GWA had no significant effects on estrogen-related signaling pathways, indicating a unique advantage in lowering the risk of gynecological cancer. In conclusion, our data suggest that GWA may be a promising candidate for the therapy of diseases associated with bone loss. Targeting pJNK-NFATc1 interaction may represent a new strategy to stimulate osteogenesis.
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Affiliation(s)
- Yu-Xuan Hao
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Yong-Yan Chen
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Xu Han
- Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, Shanghai, China; Shanghai Clinical Research Center for Aging and Medicine, Shanghai, China; Center of Community-Based Health Research, Fudan University, Shanghai, China
| | - Xu Wang
- Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, Shanghai, China; Shanghai Clinical Research Center for Aging and Medicine, Shanghai, China; Center of Community-Based Health Research, Fudan University, Shanghai, China; Shanghai Key Laboratory of Complex Prescription, and Ministry of Education (MOE) Key Laboratory for Standardization of Chinese Medicines and the State Administration of Traditional Chinese Medicine (SATCM) Key Laboratory for New Resources and Quality Evaluation of Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China
| | - Fu-Peng Wu
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China; Department of Emergency, Minhang Hospital, Fudan University, Shanghai, China
| | - Cai-Ling Wen
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Tong-Qing Chen
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Sheng Tan
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Dan-Dan Zheng
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China
| | - Yang Hong
- Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, Shanghai, China; Shanghai Clinical Research Center for Aging and Medicine, Shanghai, China; Center of Community-Based Health Research, Fudan University, Shanghai, China.
| | - Xiao-Yan Shen
- Department of Pharmacology & Quzhou Fudan Institute, School of Pharmacy, Fudan University, Shanghai, China; Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, Shanghai, China; Shanghai Clinical Research Center for Aging and Medicine, Shanghai, China; Center of Community-Based Health Research, Fudan University, Shanghai, China; The Key Laboratory of Smart Drug Delivery, Ministry of Education, School of Pharmacy, Fudan University, Shanghai, China; MOE Innovative Center for New Drug Development of Immune Inflammatory Diseases, Fudan University, Shanghai, China.
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3
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Shreya S, Dagar N, Gaikwad AB. Unlocking the therapeutic potential of the NFAT pathway in kidney diseases. NAUNYN-SCHMIEDEBERG'S ARCHIVES OF PHARMACOLOGY 2025:10.1007/s00210-025-04033-x. [PMID: 40088333 DOI: 10.1007/s00210-025-04033-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/09/2024] [Accepted: 03/06/2025] [Indexed: 03/17/2025]
Abstract
The nuclear factor of activated T cells (NFAT) is a novel renoprotective transcription factor in an inactive form in the cytoplasm and an active form in the nucleus. NFAT is expressed in T cells, heart, kidney and lymphocytes. NFAT plays an essential role in inducing apoptosis of renal tubular epithelial cells. NFAT levels have been observed to increase significantly during kidney diseases. Further, downregulation or silencing of endogenous NFAT mitigates kidney diseases. NFAT regulation depends upon the intricate interplay between calcium ions and calcineurin (CaN), thus orchestrating the NFAT/calcineurin signalling pathway. When CaN is activated, it induces dephosphorylation of NFAT and localises the active NFAT into the nucleus, which ultimately leads to inflammation, fibrosis and apoptosis of kidney cells. Further, the global incidence (> 800 million) due to kidney disease imposes a significant economic burden on the healthcare system. Therefore, it is crucial to comprehend the pathways involved in the pathophysiology of kidney diseases to develop targeted interventions. Ongoing studies indicate potential therapies, including anandamide, 11R-VIVIT and maxacalcitol to regulate NFAT levels in kidney disease. The present review discusses the role and regulation of NFAT in the pathogenesis of kidney diseases. This is focused on various preclinical studies that have shown NFAT downregulation as a potential therapeutic strategy against kidney disease setting the foundation for future clinical investigations.
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Affiliation(s)
- Shruti Shreya
- Department of Pharmacy, Birla Institute of Technology and Science, Pilani Campus, Pilani, Rajasthan, 333031, India
| | - Neha Dagar
- Department of Pharmacy, Birla Institute of Technology and Science, Pilani Campus, Pilani, Rajasthan, 333031, India
| | - Anil Bhanudas Gaikwad
- Department of Pharmacy, Birla Institute of Technology and Science, Pilani Campus, Pilani, Rajasthan, 333031, India.
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Udemgba C, Pillay B, Shafer S, Alberstadt A, Abers M, Gilliaux O, Chen K, Rae W, Hanitsch L, Von Bernuth H, Neves JF, Raje N, Moens L, van Hagen PM, Bergerson J, Hartog N, Niehues T, Dückers G, Falcone E, Keller M, Hsu A, Meyts I, Holland SM. IRF2BP2 deficiency: An important form of common variable immunodeficiency with inflammation. J Allergy Clin Immunol 2025:S0091-6749(25)00275-1. [PMID: 40090425 DOI: 10.1016/j.jaci.2025.02.038] [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: 11/30/2023] [Revised: 02/05/2025] [Accepted: 02/06/2025] [Indexed: 03/18/2025]
Abstract
BACKGROUND IRF2BP2 is a transcription factor that plays an important role in regulating immune pathways, angiogenesis, apoptosis, and cell differentiation. Defects in this gene have been implicated in immunodeficiency. OBJECTIVES To deepen the understanding of the clinical implications of IRF2BP2 variants, we sought to clinically characterize and functionally test 34 individuals with IRF2BP2 variants. METHODS We collected 34 subjects across 18 families with mutations in IRF2BP2. Records were abstracted for clinical phenotypes. Functional testing was performed on PBMCs. NFAT luciferase gene reporter constructs and quantitative cDNA determinations were used to evaluate repressor activity associated with ectopic expression of various IRF2BP2 mutant constructs in Jurkat cells. RESULTS Most subjects had immunodeficiency (91%, n = 30 of 33) with variable gastrointestinal (65%, n = 20 of 31) and inflammatory or autoimmune features (57%, n = 17 of 30), including chronic abdominal pain, colitis, diarrhea, constipation, vitiligo, alopecia, and migratory rashes. There was a reduced frequency of memory B cells with poor immunoglobulin production and reduced calcium flux in response to B-cell receptor stimuli. PBMCs had increased apoptosis in vitro compared to healthy controls. Impaired IRF2BP2 repression of NFAT activation was observed using patient-derived mutant IRF2BP2 constructs compared to wild-type constructs. Similarly, TNF-α transcript levels were higher using patient-derived mutations compared to wild-type IRF2BP2 constructs. CONCLUSIONS IRF2BP2 deficiency causes a complex immunodeficiency including gastrointestinal and inflammatory disorders as well as impaired B-cell maturation. Impaired repression of the NFAT pathway appears to enhance proinflammatory signaling through proinflammatory cytokine expression.
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Affiliation(s)
- Chioma Udemgba
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Bethany Pillay
- Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium; Laboratory for Inborn Errors of Immunity, Department of Microbiology, Immunology, and Transplantation, Katholieke Universiteit Leuven, Leuven, Belgium
| | - Samantha Shafer
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Angelika Alberstadt
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Michael Abers
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Olivier Gilliaux
- Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium; Laboratory for Inborn Errors of Immunity, Department of Microbiology, Immunology, and Transplantation, Katholieke Universiteit Leuven, Leuven, Belgium
| | - Karin Chen
- Division of Immunology, Department of Pediatrics, University of Washington and Seattle Children's Research Institute, Seattle, Wash
| | - William Rae
- Cambridge Institute of Therapeutic Immunology and Infectious Disease, Jeffrey Cheah Biomedical Centre, University of Cambridge, Cambridge, United Kingdom
| | - Leif Hanitsch
- Berlin-Brandenburg Center for Regenerative Therapies, Berlin Institute of Health at Charité, Berlin, Germany
| | - Horst Von Bernuth
- Department of Pediatric Respiratory Medicine, Immunology and Critical Care Medicine, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin and Berlin Institute of Health, Berlin, Germany
| | - Joao Farela Neves
- Primary Immunodeficiencies Unit, Hospital Dona Estefânia, Centro Hospitalar Universitário de Lisboa Central, Lisbon, Portugal
| | - Nikita Raje
- Division of Allergy, Immunology, Pulmonology, and Sleep Medicine, Department of Pediatrics, Children's Mercy Kansas City, University of Missouri Kansas City, Washington, DC
| | - Leen Moens
- Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium; Laboratory for Inborn Errors of Immunity, Department of Microbiology, Immunology, and Transplantation, Katholieke Universiteit Leuven, Leuven, Belgium
| | - P Martin van Hagen
- Division of Clinical Immunology, Department of Internal Medicine, Rotterdam, The Netherlands; Department of Immunology, Erasmus Medical Center, Rotterdam, The Netherlands
| | - Jenna Bergerson
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Nicholas Hartog
- Michigan State University College of Human Medicine Corewell Health and Helen DeVos Children's Hospital, Grand Rapids, Mich
| | - Tim Niehues
- Centre for Child and Adolescent Health, Helios Klinikum Krefeld, affiliated with Rheinisch-Westfälische Technische Hochschule University Aachen, Aachen, Germany
| | - Gregor Dückers
- Centre for Child and Adolescent Health, Helios Klinikum Krefeld, affiliated with Rheinisch-Westfälische Technische Hochschule University Aachen, Aachen, Germany
| | - Emilia Falcone
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Michael Keller
- Division of Allergy and Immunology, Children's National Hospital, Washington, DC
| | - Amy Hsu
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md
| | - Isabelle Meyts
- Department of Pediatrics, University Hospitals Leuven, Leuven, Belgium; Laboratory for Inborn Errors of Immunity, Department of Microbiology, Immunology, and Transplantation, Katholieke Universiteit Leuven, Leuven, Belgium
| | - Steven M Holland
- National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Md.
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Hopl V, Tiffner A, Wutscher A, Sallinger M, Grabmayr H, Prantl M, Fröhlich M, Söllner J, Weiß S, Najjar H, Nazarenko Y, Harant S, Kriško N, Fahrner M, Humer C, Höglinger C, Krobath H, Bonhenry D, Derler I. Water in peripheral TM-interfaces of Orai1-channels triggers pore opening. Commun Biol 2024; 7:1522. [PMID: 39550503 PMCID: PMC11569263 DOI: 10.1038/s42003-024-07174-6] [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: 12/21/2023] [Accepted: 10/30/2024] [Indexed: 11/18/2024] Open
Abstract
The activation of the Ca2+-channel Orai1 via the physiological activator stromal interaction molecule 1 (STIM1) requires structural rearrangements within the entire channel complex involving a series of gating checkpoints. Focusing on the gating mechanism operating along the peripheral transmembrane domain (TM) 3/TM4-interface, we report here that some charged substitutions close to the center of TM3 or TM4 lead to constitutively active Orai1 variants triggering nuclear factor of activated T-cell (NFAT) translocation into the nucleus. Molecular dynamics simulations unveil that this gain-of-function correlates with enhanced hydration at peripheral TM-interfaces, leading to increased local structural flexibility of the channel periphery and global conformational changes permitting pore opening. Our findings indicate that efficient dehydration of the peripheral TM-interfaces driven by the hydrophobic effect is critical for maintaining the closed state of Orai1. We conclude that a charge close to the center of TM3 or TM4 facilitates concomitant hydration and widening of peripheral TM interfaces to trigger constitutive Orai1 pore opening to a level comparable to or exceeding that of native activated Orai1.
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Affiliation(s)
- Valentina Hopl
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Adéla Tiffner
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
- Institute for Physiology and Pathophysiology, Johannes Kepler University Linz, Linz, Austria
| | - Armin Wutscher
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Matthias Sallinger
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Herwig Grabmayr
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Magdalena Prantl
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Maximilian Fröhlich
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Julia Söllner
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Sarah Weiß
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Hadil Najjar
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Yuliia Nazarenko
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Selina Harant
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Natalia Kriško
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Marc Fahrner
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Christina Humer
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Carmen Höglinger
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria
| | - Heinrich Krobath
- Institute of Theoretical Physics, Johannes Kepler University Linz, Linz, Austria
| | - Daniel Bonhenry
- Department of Physics and Materials Science, University of Luxembourg, Luxembourg City, Luxembourg.
| | - Isabella Derler
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz, Austria.
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6
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Hamar J, Cnaani A, Kültz D. Transcriptional upregulation of the myo-inositol biosynthesis pathway is enhanced by NFAT5 in hyperosmotically stressed tilapia cells. Am J Physiol Cell Physiol 2024; 327:C545-C556. [PMID: 38946247 DOI: 10.1152/ajpcell.00187.2024] [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: 03/25/2024] [Revised: 06/13/2024] [Accepted: 06/16/2024] [Indexed: 07/02/2024]
Abstract
Euryhaline fish experience variable osmotic environments requiring physiological adjustments to tolerate elevated salinity. Mozambique tilapia (Oreochromis mossambicus) possess one of the highest salinity tolerance limits of any fish. In tilapia and other euryhaline fish species, the myo-inositol biosynthesis (MIB) pathway enzymes, myo-inositol phosphate synthase (MIPS) and inositol monophosphatase 1 (IMPA1.1), are among the most upregulated mRNAs and proteins indicating the high importance of this pathway for hyperosmotic (HO) stress tolerance. These abundance changes must be precluded by HO perception and signaling mechanism activation to regulate the expression of MIPS and IMPA1.1 genes. In previous work using a O. mossambicus cell line (OmB), a reoccurring osmosensitive enhancer element (OSRE1) in both MIPS and IMPA1.1 was shown to transcriptionally upregulate these enzymes in response to HO stress. The OSRE1 core consensus (5'-GGAAA-3') matches the core binding sequence of the predominant mammalian HO response transcription factor, nuclear factor of activated T-cells (NFAT5). HO-challenged OmB cells showed an increase in NFAT5 mRNA suggesting NFAT5 may contribute to MIB pathway regulation in euryhaline fish. Ectopic expression of wild-type NFAT5 induced an IMPA1.1 promoter-driven reporter by 5.1-fold (P < 0.01). Moreover, expression of dominant negative NFAT5 in HO media resulted in a 47% suppression of the reporter signal (P < 0.005). Furthermore, reductions of IMPA1.1 (37-49%) and MIPS (6-37%) mRNA abundance were observed in HO-challenged NFAT5 knockout cells relative to control cells. Collectively, these multiple lines of experimental evidence establish NFAT5 as a tilapia transcription factor contributing to HO-induced activation of the MIB pathway.NEW & NOTEWORTHY In our study, we use a multi-pronged synthetic biology approach to demonstrate that the fish homolog of the predominant mammalian osmotic stress transcription factor nuclear factor of activated T-cells (NFAT5) also contributes to the activation of hyperosmolality inducible genes in cells of extremely euryhaline fish. However, in addition to NFAT5 the presence of other strong osmotically inducible signaling mechanisms is required for full activation of osmoregulated tilapia genes.
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Affiliation(s)
- Jens Hamar
- Department of Animal Sciences and Genome Center, University of California Davis, Davis, California, United States
| | - Avner Cnaani
- Department of Poultry and Aquaculture, Institute of Animal Sciences, Agricultural Research Organization, Rishon LeZion, Israel
| | - Dietmar Kültz
- Department of Animal Sciences and Genome Center, University of California Davis, Davis, California, United States
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7
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Gourisankar S, Krokhotin A, Wenderski W, Crabtree GR. Context-specific functions of chromatin remodellers in development and disease. Nat Rev Genet 2024; 25:340-361. [PMID: 38001317 PMCID: PMC11867214 DOI: 10.1038/s41576-023-00666-x] [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] [Accepted: 09/27/2023] [Indexed: 11/26/2023]
Abstract
Chromatin remodellers were once thought to be highly redundant and nonspecific in their actions. However, recent human genetic studies demonstrate remarkable biological specificity and dosage sensitivity of the thirty-two adenosine triphosphate (ATP)-dependent chromatin remodellers encoded in the human genome. Mutations in remodellers produce many human developmental disorders and cancers, motivating efforts to investigate their distinct functions in biologically relevant settings. Exquisitely specific biological functions seem to be an emergent property in mammals, and in many cases are based on the combinatorial assembly of subunits and the generation of stable, composite surfaces. Critical interactions between remodelling complex subunits, the nucleosome and other transcriptional regulators are now being defined from structural and biochemical studies. In addition, in vivo analyses of remodellers at relevant genetic loci have provided minute-by-minute insights into their dynamics. These studies are proposing new models for the determinants of remodeller localization and function on chromatin.
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Affiliation(s)
- Sai Gourisankar
- Department of Pathology, Stanford University, Stanford, CA, USA
- Department of Chemical Engineering, Stanford University, Stanford, CA, USA
| | - Andrey Krokhotin
- Department of Pathology, Stanford University, Stanford, CA, USA
- Department of Developmental Biology, Stanford University, Stanford, CA, USA
| | - Wendy Wenderski
- Department of Pathology, Stanford University, Stanford, CA, USA
- Department of Developmental Biology, Stanford University, Stanford, CA, USA
| | - Gerald R Crabtree
- Department of Pathology, Stanford University, Stanford, CA, USA.
- Department of Developmental Biology, Stanford University, Stanford, CA, USA.
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8
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Dashti NK, Perret R, Balzer B, Naous R, Michal M, Dermawan JK, Antonescu CR. Vascular Neoplasms With NFATC1/C2 Gene Alterations : Expanding the Clinicopathologic and Molecular Characteristics of a Distinct Entity. Am J Surg Pathol 2024; 48:487-496. [PMID: 38189436 PMCID: PMC11591551 DOI: 10.1097/pas.0000000000002175] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2024]
Abstract
Despite significant advances in their molecular pathogenesis, skeletal vascular tumors remain diagnostically challenging due to their aggressive radiologic appearance and significant morphologic overlap. Within the epithelioid category and at the benign end of the spectrum, recurrent FOS/FOSB fusions have defined most epithelioid hemangiomas, distinguishing them from epithelioid hemangioendothelioma and angiosarcoma. More recently, the presence of EWSR1/FUS :: NFATC1/2 fusions emerged as the genetic hallmark of a novel group of unusual vascular proliferations, often displaying epithelioid morphology, with alternating vasoformative and solid growth, variable atypia, reminiscent of composite hemangioendothelioma. In this study, we further our understanding and morphologic spectrum of NFATC -fusion positive vascular neoplasms by describing 9 new cases, including soft tissue locations and novel fusion partners. Combining with the initial cohort of 5 cases, a total of 14 patients were analyzed, showing slight female predilection and an age range of 10 to 66 (mean 42 y). Twelve patients had solitary lesions, while 2 had multifocal polyostotic (pelvic bones) disease. Overall, 12 lesions were intra-osseous and 2 in soft tissue. By targeted RNA Fusion panels or FISH, there were 6 cases of EWSR1::NFATC1 , 4 EWSR1::NFATC2 , 2 FUS::NFATC2 , 1 EWSR1 rearrangement, and 1 with a novel FABP4::NFATC2 fusion. Follow-up was available in 4 patients. One patient experienced 2 local recurrences, 11 and 15 years postdiagnosis, and one patient experienced progressive disease despite multimodality treatment (curettings, embolization, radiation) over 3 years. In summary, our extended investigation confirms that NFATC -related fusions define a distinct group of vascular neoplasms with variable architecture, epithelioid phenotype, and cytologic atypia, commonly located in the bone, occasionally multifocal and with potential for local recurrence and aggressive behavior but no metastatic potential. Molecular analysis is recommended in diagnostically challenging cases with atypical histology to exclude malignancy.
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Affiliation(s)
- Nooshin K. Dashti
- Department of Pathology and Laboratory Medicine Dartmouth Health, Lebanon, NH, USA
- Geisel School of Medicine at Dartmouth, Lebanon, NH, USA
| | - Raul Perret
- Department of Biopathology, Institut Bergonié, Comprehensive Cancer Center, Bordeaux, France
- Bordeaux Institute of Oncology, BRIC, INSERM, Bordeaux University, Bergonié Institute, Bordeaux, France
| | | | - Rana Naous
- University of Pittsburgh Medical Center, Shadyside, Pittsburgh, PA, USA
| | - Michael Michal
- Biopticka Laboratory, Pilsen, Czech Republic
- Department of Pathology, Charles University, Faculty of Medicine in Plzen, Czech Republic
| | - Josephine K. Dermawan
- Robert J. Tomsich Pathology and Laboratory Medicine Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Cristina R. Antonescu
- Department of Pathology and Lab Medicine, Memorial Sloan-Kettering Cancer Center, New York, NY, USA
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9
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Lin Y, Song Y, Zhang Y, Shi M, Hou A, Han S. NFAT signaling dysregulation in cancer: Emerging roles in cancer stem cells. Biomed Pharmacother 2023; 165:115167. [PMID: 37454598 DOI: 10.1016/j.biopha.2023.115167] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2023] [Revised: 07/08/2023] [Accepted: 07/11/2023] [Indexed: 07/18/2023] Open
Abstract
The nuclear factor of activated T cells (NFAT) was first identified as a transcriptional regulator of activated T cells. The NFAT family is involved in the development of tumors. Furthermore, recent evidence reveals that NFAT proteins regulate the development of inflammatory and immune responses. New discoveries have also been made about the mechanisms by which NFAT regulates cancer progression through cancer stem cells (CSC). Here, we discuss the role of the NFAT family in the immune system and various cancer types.
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Affiliation(s)
- Yibin Lin
- Department of Neurosurgery, The First Hospital of China Medical University, Shenyang 110001, China
| | - Yifu Song
- Department of Neurosurgery, The First Hospital of China Medical University, Shenyang 110001, China
| | - Yaochuan Zhang
- Department of Neurosurgery, The First Hospital of China Medical University, Shenyang 110001, China
| | - Mengwu Shi
- Department of Neurosurgery, The First Hospital of China Medical University, Shenyang 110001, China
| | - Ana Hou
- Department of Pediatrics, Shengjing Hospital of China Medical University, Shenyang 110001, China.
| | - Sheng Han
- Department of Neurosurgery, The First Hospital of China Medical University, Shenyang 110001, China.
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10
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Alfaro GF, Palombo V, D’Andrea M, Cao W, Zhang Y, Beever J, Muntifering RB, Pacheco WJ, Rodning SP, Wang X, Moisá SJ. Hepatic transcript profiling in beef cattle: Effects of rumen-protected niacin supplementation. PLoS One 2023; 18:e0289409. [PMID: 37535643 PMCID: PMC10399858 DOI: 10.1371/journal.pone.0289409] [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: 12/01/2022] [Accepted: 07/18/2023] [Indexed: 08/05/2023] Open
Abstract
The objective of our study was to assess the effect of rumen-protected niacin supplementation on the transcriptome of liver tissue in growing Angus × Simmental steers and heifers through RNA-seq analysis. Consequently, we wanted to assess the known role of niacin in the physiological processes of vasodilation, detoxification, and immune function in beef hepatic tissue. Normal weaned calves (~8 months old) were provided either a control diet or a diet supplemented with rumen-protected niacin (6 g/hd/d) for a 30-day period, followed by a liver biopsy. We observed a significant list of changes at the transcriptome level due to rumen-protected niacin supplementation. Several metabolic pathways revealed potential positive effects to the animal's liver metabolism due to administration of rumen-protected niacin; for example, a decrease in lipolysis, apoptosis, inflammatory responses, atherosclerosis, oxidative stress, fibrosis, and vasodilation-related pathways. Therefore, results from our study showed that the liver transcriptional machinery switched several metabolic pathways to a condition that could potentially benefit the health status of animals supplemented with rumen-protected niacin. In conclusion, based on the results of our study, we can suggest the utilization of rumen-protected niacin supplementation as a nutritional strategy could improve the health status of growing beef cattle in different beef production stages, such as backgrounding operations or new arrivals to a feedlot.
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Affiliation(s)
- Gastón F. Alfaro
- Department of Animal Sciences, Auburn University, Auburn, AL, United States of America
| | - Valentino Palombo
- Department of Agricultural, Environmental and Food Sciences, Università degli Studi del Molise, Campobasso, Italy
| | - Mariasilvia D’Andrea
- Department of Agricultural, Environmental and Food Sciences, Università degli Studi del Molise, Campobasso, Italy
| | - Wenqi Cao
- Department of Pathobiology, College of Veterinary Medicine, Auburn University, Auburn, AL, United States of America
| | - Yue Zhang
- Department of Pathobiology, College of Veterinary Medicine, Auburn University, Auburn, AL, United States of America
| | - Jonathan Beever
- Department of Animal Sciences, University of Tennessee, Knoxville, TN, United States of America
| | - Russell B. Muntifering
- Department of Animal Sciences, Auburn University, Auburn, AL, United States of America
- Cooperative Extension Service, University of Kentucky, Kentucky, Lexington, United States of America
| | - Wilmer J. Pacheco
- Department of Poultry Sciences, Auburn University, Auburn, AL, United States of America
| | - Soren P. Rodning
- Department of Animal Sciences, Auburn University, Auburn, AL, United States of America
| | - Xu Wang
- Department of Pathobiology, College of Veterinary Medicine, Auburn University, Auburn, AL, United States of America
- HudsonAlpha Institute for Biotechnology, Huntsville, AL, United States of America
| | - Sonia J. Moisá
- Department of Animal Sciences, University of Tennessee, Knoxville, TN, United States of America
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11
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Gamboa M, Kitamura N, Miura K, Noda S, Kaminuma O. Evolutionary mechanisms underlying the diversification of nuclear factor of activated T cells across vertebrates. Sci Rep 2023; 13:6468. [PMID: 37156933 PMCID: PMC10167247 DOI: 10.1038/s41598-023-33751-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2022] [Accepted: 04/18/2023] [Indexed: 05/10/2023] Open
Abstract
The mechanisms of immunity linked to biological evolution are crucial for understanding animal morphogenesis, organogenesis, and biodiversity. The nuclear factor of activated T cells (NFAT) family consists of five members (NFATc1-c4, 5) with different functions in the immune system. However, the evolutionary dynamics of NFATs in vertebrates has not been explored. Herein, we investigated the origin and mechanisms underlying the diversification of NFATs by comparing the gene, transcript and protein sequences, and chromosome information. We defined an ancestral origin of NFATs during the bilaterian development, dated approximately 650 million years ago, where NFAT5 and NFATc1-c4 were derived independently. The conserved parallel evolution of NFATs in multiple species was probably attributed to their innate nature. Conversely, frequent gene duplications and chromosomal rearrangements in the recently evolved taxa have suggested their roles in the adaptive immune evolution. A significant correlation was observed between the chromosome rearrangements with gene duplications and the structural fixation changes in vertebrate NFATs, suggesting their role in NFAT diversification. Remarkably, a conserved gene structure around NFAT genes with vertebrate evolutionary-related breaking points indicated the inheritance of NFATs with their neighboring genes as a unit. The close relationship between NFAT diversification and vertebrate immune evolution was suggested.
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Affiliation(s)
- Maribet Gamboa
- Department of Disease Model, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, 734-8553, Japan.
- Department of Ecology, Faculty of Sciences, Universidad Católica de la Santísima Concepción, 4090541, Concepción, Chile.
| | - Noriko Kitamura
- Neurovirology Project, Tokyo Metropolitan Institute of Medical Science, Tokyo, 156-8506, Japan
| | - Kento Miura
- Department of Disease Model, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, 734-8553, Japan
| | - Satoko Noda
- Graduate School of Science and Engineering, Ibaraki University, Ibaraki, 310-8512, Japan
| | - Osamu Kaminuma
- Department of Disease Model, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, 734-8553, Japan.
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12
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Giampaolo S, Chiarolla CM, Knöpper K, Vaeth M, Klein M, Muhammad A, Bopp T, Berberich-Siebelt F, Patra AK, Serfling E, Klein-Hessling S. NFATc1 induction by an intronic enhancer restricts NKT γδ cell formation. iScience 2023; 26:106234. [PMID: 36926655 PMCID: PMC10011748 DOI: 10.1016/j.isci.2023.106234] [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: 04/19/2022] [Revised: 06/08/2022] [Accepted: 02/13/2023] [Indexed: 02/22/2023] Open
Abstract
In thymus, the ablation of T cell receptor (TCR)-activated transcription factor NFATc1 or its inducible isoforms during the double-negative (DN) stages of thymocyte development leads to a marked increase in γδ thymocytes whereas the development of αβ thymocytes remains mostly unaffected. These γδ thymocytes are characterized by the upregulation of the promyelocytic leukemia zinc-finger factor (PLZF), the "master regulator" of natural killer T (NKT) cell development, and the acquisition of an NKT γδ cell phenotype with higher cell survival rates. The suppressive function of NFATc1 in NKT γδ cell formation critically depends on the remote enhancer E2, which is essential for the inducible expression of NFATc1 directed by its distal promoter P1. Thus, the enhancer deciphers a strong γδ TCR signal into the expression of inducible NFATc1 isoforms resulting in high levels of NFATc1 protein that are essential to control the numbers of NKT γδ cells.
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Affiliation(s)
- Sabrina Giampaolo
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany
| | - Cristina M Chiarolla
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany
| | - Konrad Knöpper
- Würzburg Institute of Systems Immunology, Max Planck Research Group at the Julius-Maximilians-University Würzburg, Versbacher Strasse 9, 97078 Würzburg, Germany
| | - Martin Vaeth
- Würzburg Institute of Systems Immunology, Max Planck Research Group at the Julius-Maximilians-University Würzburg, Versbacher Strasse 9, 97078 Würzburg, Germany
| | - Matthias Klein
- Institute for Immunology, University Medical Center, University of Mainz, Langenbeckstraße 1, 55131 Mainz, Germany
| | - Azeem Muhammad
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany
| | - Tobias Bopp
- Institute for Immunology, University Medical Center, University of Mainz, Langenbeckstraße 1, 55131 Mainz, Germany
| | - Friederike Berberich-Siebelt
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany
| | - Amiya K Patra
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany.,Peninsula Medical School, University of Plymouth, The John Bull Building, Plymouth Science Park, Research Way, Plymouth PL6 8BU, UK
| | - Edgar Serfling
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany.,Comprehensive Cancer Center Mainfranken, Würzburg, Germany
| | - Stefan Klein-Hessling
- Institute of Pathology, Julius Maximilians University Würzburg, Josef-Schneider-Strasse 2, 97080 Würzburg, Germany
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13
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Huang H, Dong J, Jiang J, Yang F, Zheng Y, Wang S, Wang N, Ma J, Hou M, Ding Y, Meng L, Zhuo W, Yang D, Qian W, Chen Q, You G, Qian G, Gu L, Lv H. The role of FOXO4/NFAT2 signaling pathway in dysfunction of human coronary endothelial cells and inflammatory infiltration of vasculitis in Kawasaki disease. Front Immunol 2023; 13:1090056. [PMID: 36700213 PMCID: PMC9869249 DOI: 10.3389/fimmu.2022.1090056] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2022] [Accepted: 12/19/2022] [Indexed: 01/12/2023] Open
Abstract
Aims The Ca+/NFAT (Nuclear factor of activated T cells) signaling pathway activation is implicated in the pathogenesis of Kawasaki disease (KD); however, we lack detailed information regarding the regulatory network involved in the human coronary endothelial cell dysfunction and cardiovascular lesion development. Herein, we aimed to use mouse and endothelial cell models of KD vasculitis in vivo and in vitro to characterize the regulatory network of NFAT pathway in KD. Methods and Results Among the NFAT gene family, NFAT2 showed the strongest transcriptional activity in peripheral blood mononuclear cells (PBMCs) from patients with KD. Then, NFAT2 overexpression and knockdown experiments in Human coronary artery endothelial cells (HCAECs) indicated that NFAT2 overexpression disrupted endothelial cell homeostasis by regulation of adherens junctions, whereas its knockdown protected HCAECs from such dysfunction. Combined analysis using RNA-sequencing and transcription factor (TF) binding site analysis in the NFAT2 promoter region predicted regulation by Forkhead box O4 (FOXO4). Western blotting, chromatin immunoprecipitation, and luciferase assays validated that FOXO4 binds to the promoter and transcriptionally represses NFAT2. Moreover, Foxo4 knockout increased the extent of inflamed vascular tissues in a mouse model of KD vasculitis. Functional experiments showed that inhibition NFAT2 relieved Foxo4 knockout exaggerated vasculitis in vivo. Conclusions Our findings revealed the FOXO4/NFAT2 axis as a vital pathway in the progression of KD that is associated with endothelial cell homeostasis and cardiovascular inflammation development.
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Affiliation(s)
- Hongbiao Huang
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China,Department of Pediatrics, Fujian Provincial Hospital, Fujian Provincial Clinical College of Fujian Medical University, Fuzhou, Fujian, China,Epigenetics Laboratory, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany
| | - Jinfeng Dong
- Department of Hematology, the First Affiliated Hospital of Fujian Medical University, Fuzhou, Fujian, China
| | - Jiaqi Jiang
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Fang Yang
- Department of Pediatrics, Fujian Provincial Hospital, Fujian Provincial Clinical College of Fujian Medical University, Fuzhou, Fujian, China
| | - Yiming Zheng
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Shuhui Wang
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Nana Wang
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Jin Ma
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Miao Hou
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Yueyue Ding
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Lijun Meng
- Department of Hematology, Children’s Hospital of Soochow University, Suzhou, China
| | - Wenyu Zhuo
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Daoping Yang
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Weiguo Qian
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Qiaobin Chen
- Department of Pediatrics, Fujian Provincial Hospital, Fujian Provincial Clinical College of Fujian Medical University, Fuzhou, Fujian, China
| | - Guoping You
- Department of Emergency, Fujian Provincial Hospital, Fujian Provincial Clinical College of Fujian Medical University, Fuzhou, Fujian, China
| | - Guanghui Qian
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China
| | - Lei Gu
- Epigenetics Laboratory, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany,Cardiopulmonary Institute (CPI), Bad Nauheim, Germany,*Correspondence: Haitao Lv, ; Lei Gu,
| | - Haitao Lv
- Department of Pediatrics, Institute of Pediatric Research, Children’s Hospital of Soochow University, Suzhou, Jiangsu, China,*Correspondence: Haitao Lv, ; Lei Gu,
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14
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Increased Expression of the RBPMS Splice Variants Inhibits Cell Proliferation in Ovarian Cancer Cells. Int J Mol Sci 2022; 23:ijms232314742. [PMID: 36499073 PMCID: PMC9738375 DOI: 10.3390/ijms232314742] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2022] [Revised: 11/22/2022] [Accepted: 11/23/2022] [Indexed: 11/29/2022] Open
Abstract
RNA-Binding Protein with Multiple Splicing (RBPMS) is a member of family proteins that bind to nascent RNA transcripts and regulate their splicing, localization, and stability. Evidence indicates that RBPMS controls the activity of transcription factors associated with cell growth and proliferation, including AP-1 and Smads. Three major RBPMS protein splice variants (RBPMSA, RBPMSB, and RBPMSC) have been described in the literature. We previously reported that reduced RBPMS levels decreased the sensitivity of ovarian cancer cells to cisplatin treatment. However, little is known about the biological role of the RBPMS splice variants in ovarian cancer cells. We performed RT-PCR and Western blots and observed that both RBPMSA and RBPMSC are reduced at the mRNA and protein levels in cisplatin resistant as compared with cisplatin sensitive ovarian cancer cells. The mRNA and protein levels of RBPMSB were not detectable in any of the ovarian cancer cells tested. To better understand the biological role of each RBPMSA and RBPMSC, we transfected these two splice variants in the A2780CP20 and OVCAR3CIS cisplatin resistant ovarian cancer cells and performed cell proliferation, cell migration, and invasion assays. Compared with control clones, a significant reduction in the number of colonies, colony size, cell migration, and invasion was observed with RBPMSA and RBPMSC overexpressed cells. Moreover, A2780CP20-RBPMSA and A2780CP20-RBPMSC clones showed reduced senescence-associated β-galactosidase (β-Gal)-levels when compared with control clones. A2780CP20-RBPMSA clones were more sensitive to cisplatin treatment as compared with A2780CP20-RBPMSC clones. The A2780CP20-RBPMSA and A2780CP20-RBPMSC clones subcutaneously injected into athymic nude mice formed smaller tumors as compared with A2780CP20-EV control group. Additionally, immunohistochemical analysis showed lower proliferation (Ki67) and angiogenesis (CD31) staining in tissue sections of A2780CP20-RBPMSA and A2780CP20-RBPMSC tumors compared with controls. RNAseq studies revealed many common RNA transcripts altered in A2780CP20-RBPMSA and A2780CP20-RBPMSC clones. Unique RNA transcripts deregulated by each RBPMS variant were also observed. Kaplan-Meier (KM) plotter database information identified clinically relevant RBPMSA and RBPMSC downstream effectors. These studies suggest that increased levels of RBPMSA and RBPMSC reduce cell proliferation in ovarian cancer cells. However, only RBPMSA expression levels were associated with the sensitivity of ovarian cancer cells to cisplatin treatment.
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15
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Gallo C, Manzo E, Barra G, Fioretto L, Ziaco M, Nuzzo G, d'Ippolito G, Ferrera F, Contini P, Castiglia D, Angelini C, De Palma R, Fontana A. Sulfavant A as the first synthetic TREM2 ligand discloses a homeostatic response of dendritic cells after receptor engagement. Cell Mol Life Sci 2022; 79:369. [PMID: 35723745 PMCID: PMC9207826 DOI: 10.1007/s00018-022-04297-z] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2021] [Revised: 03/18/2022] [Accepted: 04/08/2022] [Indexed: 11/30/2022]
Abstract
OBJECTIVE The immune response arises from a fine balance of mechanisms that provide for surveillance, tolerance, and elimination of dangers. Sulfavant A (SULF A) is a sulfolipid with a promising adjuvant activity. Here we studied the mechanism of action of SULF A and addressed the identification of its molecular target in human dendritic cells (hDCs). METHODS Adjuvant effect and immunological response to SULF A were assessed on DCs derived from human donors. In addition to testing various reporter cells, target identification and downstream signalling was supported by a reverse pharmacology approach based on antibody blocking and gene silencing, crosstalk with TLR pathways, use of human allogeneic mixed lymphocyte reaction. RESULTS SULF A binds to the Triggering Receptor Expressed on Myeloid cells-2 (TREM2) and initiates an unconventional maturation of hDCs leading to enhanced migration activity and up-regulation of MHC and co-stimulatory molecules without release of conventional cytokines. This response involves the SYK-NFAT axis and is compromised by blockade or gene silencing of TREM2. Activation by SULF A preserved the DC functions to excite the allogeneic T cell response, and increased interleukin-10 release after lipopolysaccharide stimulation. CONCLUSION SULF A is the first synthetic small molecule that binds to TREM2. The receptor engagement drives differentiation of an unprecedented DC phenotype (homeDCs) that contributes to immune homeostasis without compromising lymphocyte activation and immunogenic response. This mechanism fully supports the adjuvant and immunoregulatory activity of SULF A. We also propose that the biological properties of SULF A can be of interest in various physiopathological mechanisms and therapies involving TREM2.
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Affiliation(s)
- Carmela Gallo
- Consiglio Nazionale delle Ricerche, Istituto di Chimica Biomolecolare, Via Campi Flegrei 34, 80078, Pozzuoli, Napoli, Italy.
| | - Emiliano Manzo
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy
| | - Giusi Barra
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy
| | - Laura Fioretto
- Consorzio Italbiotec, Via Fantoli, 16/15, 20138, Milan, Italy
| | - Marcello Ziaco
- BioSearch Srl., Villa Comunale c/o Stazione Zoologica "A. Dohrn", 80121, Naples, Italy
| | - Genoveffa Nuzzo
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy
| | - Giuliana d'Ippolito
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy
| | - Francesca Ferrera
- Department of Internal Medicine, University of Genova, Viale Benedetto XV 6, 16100, Genoa, Italy
| | - Paola Contini
- Department of Internal Medicine, University of Genova, Viale Benedetto XV 6, 16100, Genoa, Italy
| | - Daniela Castiglia
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy
| | - Claudia Angelini
- Institute for Applied Mathematics "Mauro Picone", National Research Council, Via Pietro Castellino 111, 80131, Naples, Italy
| | - Raffaele De Palma
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy.,Department of Internal Medicine, University of Genova, Viale Benedetto XV 6, 16100, Genoa, Italy
| | - Angelo Fontana
- Bio-Organic Chemistry Unit, Institute of Bio-Molecular Chemistry, National Research Council, Via Campi Flegrei 34, 80078, Pozzuoli, Italy. .,Department of Biology, University of Napoli "Federico II", Via Cupa Nuova Cinthia 21, 80126, Naples, Italy.
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16
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Zhang N, Liu Y, Shi X, Zhang Y, Li W, Yang Y, Chen L, Yin Y, Tong L, Yang J, Luo J. Microscale thermophoresis and fluorescence polarization assays of calcineurin-peptide interactions. Anal Biochem 2022; 646:114626. [PMID: 35218735 DOI: 10.1016/j.ab.2022.114626] [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: 11/13/2021] [Revised: 02/20/2022] [Accepted: 02/21/2022] [Indexed: 11/28/2022]
Abstract
Calcineurin is a Ca2+/calmodulin-dependent phosphatase. It is very important to study the affinity between calcineurin and its substrate or other interacting proteins. Two conserved motifs have been reported on the interactive proteins of calcineurin, namely, the PxIxIT motif and the LxVP motif. Here, we used 5(6)-carboxyfluorescein to fluorescently label the N-terminus of the short peptides derived from the two motifs and then determined the affinity between the protein and polypeptides. Microscale thermophoresis (MST) is very suitable for determining calcineurin with peptides containing the LxVP motif. The Kd values of the binding of calcineurin with NFATc1-YLAVP, NHE1-YLTVP, and A238L-FLCVK peptides were 6.72 ± 0.19 μM, 17.14 ± 0.35 μM, and 15.57 ± 0.10 μM, respectively. The GST pull-down results further confirmed the binding trend of the three peptides to calcineurin. However, fluorescently labeled PxIxIT polypeptides are not suitable for MST due to their own aggregation. We determined the binding affinity of the RCAN1-PSVVVH polypeptide to calcineurin by the fluorescence polarization (FP) method. MST and FP assays are fast and accurate in determining the affinity between protein-peptide interactions. Our research laid the foundation for screening the molecules that affect the binding between calcineurin and its substrates in the future.
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Affiliation(s)
- Nan Zhang
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Yueyang Liu
- Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang, 111016, China
| | - Xiaoyu Shi
- College of Life Sciences, Langfang Normal University, Hebei, 065000, China
| | - Yuchen Zhang
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Wenying Li
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Yumeng Yang
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Limin Chen
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Yanxia Yin
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Li Tong
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China
| | - Jingyu Yang
- Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang, 111016, China.
| | - Jing Luo
- Department of Biochemistry and Molecular Biology, Gene Engineering and Biotechnology of Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.
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17
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Yassouf MY, Zhang X, Huang Z, Zhai D, Sekiya R, Kawabata T, Li TS. Biphasic effect of mechanical stress on lymphocyte activation. J Cell Physiol 2022; 237:1521-1531. [PMID: 34724217 DOI: 10.1002/jcp.30623] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2021] [Revised: 10/19/2021] [Accepted: 10/22/2021] [Indexed: 01/08/2023]
Abstract
Mechanical forces can modulate the immune response, mostly described as promoting the activation of immune cells, but the role and mechanism of pathological levels of mechanical stress in lymphocyte activation have not been focused on before. By an ex vivo experimental approach, we observed that mechanical stressing of murine spleen lymphocytes with 50 mmHg for 3 h induced the nuclear localization of NFAT1, increased C-Jun, and increased the expression of early activation marker CD69 in resting CD8+ cells. Interestingly, 50 mmHg mechanical stressing induced the nuclear localization of NFAT1; but conversely decreased C-Jun and inhibited the expression of CD69 in lymphocytes under lipopolysaccharide or phorbol 12-myristate 13-acetate/ionomycin stimulation. Additionally, we observed similar changes trends when comparing RNA-seq data of hypertensive and normotensive COVID-19 patients. Our results indicate a biphasic effect of mechanical stress on lymphocyte activation, which provides insight into the variety of immune responses in pathologies involving elevated mechanical stress.
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Affiliation(s)
- Mhd Yousuf Yassouf
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Xu Zhang
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Zisheng Huang
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Da Zhai
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Reiko Sekiya
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Tsuyoshi Kawabata
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
| | - Tao-Sheng Li
- Department of Stem Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
- Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University, Nagasaki, Japan
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18
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Current Promising Therapeutic Targets for Aspergillosis Treatment. JOURNAL OF PURE AND APPLIED MICROBIOLOGY 2021. [DOI: 10.22207/jpam.15.2.09] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Aspergillosis is a fungal disease caused by different species of Aspergillus. They live in soil,dust and decomposed material. Number of Aspergillus species found till now is about 300 and more are still to be identified. Only few Aspergillus species can cause human disease and the most common species for human infection is Aspergillus fumigatus, which is a ubiquitous airborne saprophytic fungus. Severity of the disease ranges from an allergic response to life-threatening generalized infection. They grow optimally at 37°C and can grow upto 50°C. The fungal conidia are being constantly inhaled by humans and animals everyday normally gets eliminated by innate immune mechanism. Due to increasing number of immunocompromised patients, severe and fatal Aspergillosis cases have augmented. Currently, available antifungal drug for the treatment of Aspergillosis act on these three molecular target are 14 alpha demethylase for Azoles, ergosterol for Polyene and β-1,3-glucan synthase for Echinocandin. These antifungal drug show high resistance problem and toxicity. So, it is high time to develop new drugs for treatment with reduced toxicity and drug resistant problem. Synthesis of essential amino acid is absent in human as they obtain it from their diet but fungi synthesis these amino acid. Thus, enzymes in this pathway acts as novel drug target. This article summarizes promising drug targets presents in different metabolic pathway of Aspergillus genome and discusses their molecular functions in detail. This review also list down the inhibitors of these novel target. We present a comprehensive review that will pave way for discovery and development of novel antifungals against these drug targets for Aspergillosis treatment.
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19
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Grimm J, Heckl D, Klusmann JH. Molecular Mechanisms of the Genetic Predisposition to Acute Megakaryoblastic Leukemia in Infants With Down Syndrome. Front Oncol 2021; 11:636633. [PMID: 33777792 PMCID: PMC7992977 DOI: 10.3389/fonc.2021.636633] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2020] [Accepted: 01/12/2021] [Indexed: 01/28/2023] Open
Abstract
Individuals with Down syndrome are genetically predisposed to developing acute megakaryoblastic leukemia. This myeloid leukemia associated with Down syndrome (ML–DS) demonstrates a model of step-wise leukemogenesis with perturbed hematopoiesis already presenting in utero, facilitating the acquisition of additional driver mutations such as truncating GATA1 variants, which are pathognomonic to the disease. Consequently, the affected individuals suffer from a transient abnormal myelopoiesis (TAM)—a pre-leukemic state preceding the progression to ML–DS. In our review, we focus on the molecular mechanisms of the different steps of clonal evolution in Down syndrome leukemogenesis, and aim to provide a comprehensive view on the complex interplay between gene dosage imbalances, GATA1 mutations and somatic mutations affecting JAK-STAT signaling, the cohesin complex and epigenetic regulators.
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Affiliation(s)
- Juliane Grimm
- Pediatric Hematology and Oncology, Martin Luther University Halle-Wittenberg, Halle, Germany.,Department of Internal Medicine IV, Oncology/Hematology, Martin Luther University Halle-Wittenberg, Halle, Germany
| | - Dirk Heckl
- Pediatric Hematology and Oncology, Martin Luther University Halle-Wittenberg, Halle, Germany
| | - Jan-Henning Klusmann
- Pediatric Hematology and Oncology, Martin Luther University Halle-Wittenberg, Halle, Germany
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20
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Loo Yau H, Bell E, Ettayebi I, de Almeida FC, Boukhaled GM, Shen SY, Allard D, Morancho B, Marhon SA, Ishak CA, Gonzaga IM, da Silva Medina T, Singhania R, Chakravarthy A, Chen R, Mehdipour P, Pommey S, Klein C, Amarante-Mendes GP, Roulois D, Arribas J, Stagg J, Brooks DG, De Carvalho DD. DNA hypomethylating agents increase activation and cytolytic activity of CD8 + T cells. Mol Cell 2021; 81:1469-1483.e8. [PMID: 33609448 DOI: 10.1016/j.molcel.2021.01.038] [Citation(s) in RCA: 60] [Impact Index Per Article: 15.0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2020] [Revised: 12/16/2020] [Accepted: 01/27/2021] [Indexed: 12/15/2022]
Abstract
We demonstrate that DNA hypomethylating agent (HMA) treatment can directly modulate the anti-tumor response and effector function of CD8+ T cells. In vivo HMA treatment promotes CD8+ T cell tumor infiltration and suppresses tumor growth via CD8+ T cell-dependent activity. Ex vivo, HMAs enhance primary human CD8+ T cell activation markers, effector cytokine production, and anti-tumor cytolytic activity. Epigenomic and transcriptomic profiling shows that HMAs vastly regulate T cell activation-related transcriptional networks, culminating with over-activation of NFATc1 short isoforms. Mechanistically, demethylation of an intragenic CpG island immediately downstream to the 3' UTR of the short isoform was associated with antisense transcription and alternative polyadenylation of NFATc1 short isoforms. High-dimensional single-cell mass cytometry analyses reveal a selective effect of HMAs on a subset of human CD8+ T cell subpopulations, increasing both the number and abundance of a granzyme Bhigh, perforinhigh effector subpopulation. Overall, our findings support the use of HMAs as a therapeutic strategy to boost anti-tumor immune response.
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Affiliation(s)
- Helen Loo Yau
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada
| | - Emma Bell
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Ilias Ettayebi
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada
| | - Felipe Campos de Almeida
- Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo 05508-000, Brazil; Instituto de Investigação em Imunologia, Institutos Nacionais de Ciência e Tecnologia (INCT-iii), São Paulo 05403-900, Brazil
| | - Giselle M Boukhaled
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada
| | - Shu Yi Shen
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - David Allard
- Centre de recherche du Centre Hospitalier de l'Université de Montréal et Institut du Cancer de Montréal, Montréal, QC H2X 0A9, Canada; Faculté de Pharmacie, Université de Montréal, Montréal, QC H3T 1J4, Canada
| | - Beatriz Morancho
- Preclinical Research Program, Vall d'Hebron Institute of Oncology (VHIO) and CIBERONC, 08035 Barcelona, Spain
| | - Sajid A Marhon
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Charles A Ishak
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Isabela M Gonzaga
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Tiago da Silva Medina
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Translational Immuno-oncology Laboratory, A.C. Camargo Cancer Center, São Paulo 01509-001, Brazil
| | - Rajat Singhania
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Ankur Chakravarthy
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Raymond Chen
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada
| | - Parinaz Mehdipour
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada
| | - Sandra Pommey
- Centre de recherche du Centre Hospitalier de l'Université de Montréal et Institut du Cancer de Montréal, Montréal, QC H2X 0A9, Canada
| | - Christian Klein
- Roche Pharma Research and Early Development, Roche Innovation Center Zurich, Wagistrasse 10, 8952 Schlieren, Switzerland
| | - Gustavo P Amarante-Mendes
- Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo 05508-000, Brazil; Instituto de Investigação em Imunologia, Institutos Nacionais de Ciência e Tecnologia (INCT-iii), São Paulo 05403-900, Brazil
| | - David Roulois
- UMR U1236, INSERM, Université de Rennes 1, EFS, 35000 Rennes, France
| | - Joaquín Arribas
- Preclinical Research Program, Vall d'Hebron Institute of Oncology (VHIO) and CIBERONC, 08035 Barcelona, Spain; Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona, Spain; Cancer Research Program, IMIM (Hospital del Mar Medical Research Institute), 08003 Barcelona, Spain
| | - John Stagg
- Centre de recherche du Centre Hospitalier de l'Université de Montréal et Institut du Cancer de Montréal, Montréal, QC H2X 0A9, Canada; Faculté de Pharmacie, Université de Montréal, Montréal, QC H3T 1J4, Canada
| | - David G Brooks
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada
| | - Daniel D De Carvalho
- Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2C1, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada.
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21
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Tiffner A, Derler I. Molecular Choreography and Structure of Ca 2+ Release-Activated Ca 2+ (CRAC) and K Ca2+ Channels and Their Relevance in Disease with Special Focus on Cancer. MEMBRANES 2020; 10:E425. [PMID: 33333945 PMCID: PMC7765462 DOI: 10.3390/membranes10120425] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/04/2020] [Revised: 12/04/2020] [Accepted: 12/07/2020] [Indexed: 12/16/2022]
Abstract
Ca2+ ions play a variety of roles in the human body as well as within a single cell. Cellular Ca2+ signal transduction processes are governed by Ca2+ sensing and Ca2+ transporting proteins. In this review, we discuss the Ca2+ and the Ca2+-sensing ion channels with particular focus on the structure-function relationship of the Ca2+ release-activated Ca2+ (CRAC) ion channel, the Ca2+-activated K+ (KCa2+) ion channels, and their modulation via other cellular components. Moreover, we highlight their roles in healthy signaling processes as well as in disease with a special focus on cancer. As KCa2+ channels are activated via elevations of intracellular Ca2+ levels, we summarize the current knowledge on the action mechanisms of the interplay of CRAC and KCa2+ ion channels and their role in cancer cell development.
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Affiliation(s)
| | - Isabella Derler
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, A-4020 Linz, Austria;
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22
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Mastrogiovanni M, Juzans M, Alcover A, Di Bartolo V. Coordinating Cytoskeleton and Molecular Traffic in T Cell Migration, Activation, and Effector Functions. Front Cell Dev Biol 2020; 8:591348. [PMID: 33195256 PMCID: PMC7609836 DOI: 10.3389/fcell.2020.591348] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2020] [Accepted: 09/24/2020] [Indexed: 12/28/2022] Open
Abstract
Dynamic localization of receptors and signaling molecules at the plasma membrane and within intracellular vesicular compartments is crucial for T lymphocyte sensing environmental cues, triggering membrane receptors, recruiting signaling molecules, and fine-tuning of intracellular signals. The orchestrated action of actin and microtubule cytoskeleton and intracellular vesicle traffic plays a key role in all these events that together ensure important steps in T cell physiology. These include extravasation and migration through lymphoid and peripheral tissues, T cell interactions with antigen-presenting cells, T cell receptor (TCR) triggering by cognate antigen-major histocompatibility complex (MHC) complexes, immunological synapse formation, cell activation, and effector functions. Cytoskeletal and vesicle traffic dynamics and their interplay are coordinated by a variety of regulatory molecules. Among them, polarity regulators and membrane-cytoskeleton linkers are master controllers of this interplay. Here, we review the various ways the T cell plasma membrane, receptors, and their signaling machinery interplay with the actin and microtubule cytoskeleton and with intracellular vesicular compartments. We highlight the importance of this fine-tuned crosstalk in three key stages of T cell biology involving cell polarization: T cell migration in response to chemokines, immunological synapse formation in response to antigen cues, and effector functions. Finally, we discuss two examples of perturbation of this interplay in pathological settings, such as HIV-1 infection and mutation of the polarity regulator and tumor suppressor adenomatous polyposis coli (Apc) that leads to familial polyposis and colorectal cancer.
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Affiliation(s)
- Marta Mastrogiovanni
- Ligue Nationale Contre le Cancer – Equipe Labellisée LIGUE 2018, Lymphocyte Cell Biology Unit, INSERM-U1221, Department of Immunology, Institut Pasteur, Paris, France
- Collège Doctoral, Sorbonne Université, Paris, France
| | - Marie Juzans
- Ligue Nationale Contre le Cancer – Equipe Labellisée LIGUE 2018, Lymphocyte Cell Biology Unit, INSERM-U1221, Department of Immunology, Institut Pasteur, Paris, France
| | - Andrés Alcover
- Ligue Nationale Contre le Cancer – Equipe Labellisée LIGUE 2018, Lymphocyte Cell Biology Unit, INSERM-U1221, Department of Immunology, Institut Pasteur, Paris, France
| | - Vincenzo Di Bartolo
- Ligue Nationale Contre le Cancer – Equipe Labellisée LIGUE 2018, Lymphocyte Cell Biology Unit, INSERM-U1221, Department of Immunology, Institut Pasteur, Paris, France
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23
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Kongsomboonvech AK, Rodriguez F, Diep AL, Justice BM, Castallanos BE, Camejo A, Mukhopadhyay D, Taylor GA, Yamamoto M, Saeij JPJ, Reese ML, Jensen KDC. Naïve CD8 T cell IFNγ responses to a vacuolar antigen are regulated by an inflammasome-independent NLRP3 pathway and Toxoplasma gondii ROP5. PLoS Pathog 2020; 16:e1008327. [PMID: 32853276 PMCID: PMC7480859 DOI: 10.1371/journal.ppat.1008327] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2020] [Revised: 09/09/2020] [Accepted: 07/05/2020] [Indexed: 12/31/2022] Open
Abstract
Host resistance to Toxoplasma gondii relies on CD8 T cell IFNγ responses, which if modulated by the host or parasite could influence chronic infection and parasite transmission between hosts. Since host-parasite interactions that govern this response are not fully elucidated, we investigated requirements for eliciting naïve CD8 T cell IFNγ responses to a vacuolar resident antigen of T. gondii, TGD057. Naïve TGD057 antigen-specific CD8 T cells (T57) were isolated from transnuclear mice and responded to parasite-infected bone marrow-derived macrophages (BMDMs) in an antigen-dependent manner, first by producing IL-2 and then IFNγ. T57 IFNγ responses to TGD057 were independent of the parasite’s protein export machinery ASP5 and MYR1. Instead, host immunity pathways downstream of the regulatory Immunity-Related GTPases (IRG), including partial dependence on Guanylate-Binding Proteins, are required. Multiple T. gondii ROP5 isoforms and allele types, including ‘avirulent’ ROP5A from clade A and D parasite strains, were able to suppress CD8 T cell IFNγ responses to parasite-infected BMDMs. Phenotypic variance between clades B, C, D, F, and A strains suggest T57 IFNγ differentiation occurs independently of parasite virulence or any known IRG-ROP5 interaction. Consistent with this, removal of ROP5 is not enough to elicit maximal CD8 T cell IFNγ production to parasite-infected cells. Instead, macrophage expression of the pathogen sensors, NLRP3 and to a large extent NLRP1, were absolute requirements. Other members of the conventional inflammasome cascade are only partially required, as revealed by decreased but not abrogated T57 IFNγ responses to parasite-infected ASC, caspase-1/11, and gasdermin D deficient cells. Moreover, IFNγ production was only partially reduced in the absence of IL-12, IL-18 or IL-1R signaling. In summary, T. gondii effectors and host machinery that modulate parasitophorous vacuolar membranes, as well as NLR-dependent but inflammasome-independent pathways, determine the full commitment of CD8 T cells IFNγ responses to a vacuolar antigen. Parasites are excellent “students” of our immune system as they can deflect, antagonize and confuse the immune response making it difficult to vaccinate against these pathogens. In this report, we analyzed how a widespread parasite of mammals, Toxoplasma gondii, manipulates an immune cell needed for immunity to many intracellular pathogens, the CD8 T cell. Host pathways that govern CD8 T cell production of the immune protective cytokine, IFNγ, were also explored. We hypothesized the secreted T. gondii virulence factor, ROP5, work to inhibit the MHC 1 antigen presentation pathway therefore making it difficult for CD8 T cells to see T. gondii antigens sequestered inside a parasitophorous vacuole. However, manipulation through T. gondii ROP5 does not fully explain how CD8 T cells commit to making IFNγ in response to infection. Importantly, CD8 T cell IFNγ responses to T. gondii require the pathogen sensor NLRP3 to be expressed in the infected cell. Other proteins associated with NLRP3 activation, including members of the conventional inflammasome activation cascade pathway, are only partially involved. Our results identify a novel pathway by which NLRP3 regulates T cell function and underscore the need for NLRP3-activating adjuvants in vaccines aimed at inducing CD8 T cell IFNγ responses to parasites.
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Affiliation(s)
- Angel K. Kongsomboonvech
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
| | - Felipe Rodriguez
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
| | - Anh L. Diep
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
| | - Brandon M. Justice
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
| | - Brayan E. Castallanos
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
| | - Ana Camejo
- Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America
| | - Debanjan Mukhopadhyay
- Department of Pathology, Microbiology and Immunology, School of Veterinary Medicine, University of California, Davis, Davis, California, United States of America
| | - Gregory A. Taylor
- Departments of Medicine; Molecular Genetics and Microbiology; and Immunology; and Center for the Study of Aging and Human Development, Duke University Medical Center, Durham, North Carolina, United States of America
- Geriatric Research, Education, and Clinical Center, Durham VA Health Care System, Durham, North Carolina, United States of America
| | - Masahiro Yamamoto
- Department of Immunoparasitology, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
| | - Jeroen P. J. Saeij
- Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America
- Department of Pathology, Microbiology and Immunology, School of Veterinary Medicine, University of California, Davis, Davis, California, United States of America
| | - Michael L. Reese
- Department of Pharmacology, University of Texas, Southwestern Medical Center, Dallas, Texas, United States of America
| | - Kirk D. C. Jensen
- Department of Molecular and Cell Biology, University of California, Merced, Merced, California, United States of America
- Health Sciences Research Institute, University of California, Merced, Merced, California, United States of America
- * E-mail:
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24
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Cole AJ, Iyengar M, Panesso-Gómez S, O'Hayer P, Chan D, Delgoffe GM, Aird KM, Yoon E, Bai S, Buckanovich RJ. NFATC4 promotes quiescence and chemotherapy resistance in ovarian cancer. JCI Insight 2020; 5:131486. [PMID: 32182216 DOI: 10.1172/jci.insight.131486] [Citation(s) in RCA: 30] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2019] [Accepted: 03/11/2020] [Indexed: 12/30/2022] Open
Abstract
Development of chemotherapy resistance is a major problem in ovarian cancer. One understudied mechanism of chemoresistance is the induction of quiescence, a reversible nonproliferative state. Unfortunately, little is known about regulators of quiescence. Here, we identify the master transcription factor nuclear factor of activated T cells cytoplasmic 4 (NFATC4) as a regulator of quiescence in ovarian cancer. NFATC4 is enriched in ovarian cancer stem-like cells and correlates with decreased proliferation and poor prognosis. Treatment of cancer cells with cisplatin resulted in NFATC4 nuclear translocation and activation of the NFATC4 pathway, while inhibition of the pathway increased chemotherapy response. Induction of NFATC4 activity resulted in a marked decrease in proliferation, G0 cell cycle arrest, and chemotherapy resistance, both in vitro and in vivo. Finally, NFATC4 drove a quiescent phenotype in part via downregulation of MYC. Together, these data identify NFATC4 as a driver of quiescence and a potential new target to combat chemoresistance in ovarian cancer.
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Affiliation(s)
- Alexander J Cole
- Department of Internal Medicine and Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Mangala Iyengar
- Department of Cellular and Molecular Biology, University of Michigan, Ann Arbor, Michigan, USA
| | - Santiago Panesso-Gómez
- Department of Internal Medicine and Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Patrick O'Hayer
- Department of Cellular and Molecular Biology, University of Michigan, Ann Arbor, Michigan, USA
| | - Daniel Chan
- Department of Internal Medicine and Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Greg M Delgoffe
- Tumor Microenvironment Center, UPMC Hillman Cancer Center; and Department of Immunology, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Katherine M Aird
- Department of Cellular & Molecular Physiology, Penn State College of Medicine, Hershey, Pennsylvania, USA
| | - Euisik Yoon
- Department of Electrical Engineering and Computer Science, University of Michigan, Ann Arbor, Michigan, USA
| | - Shoumei Bai
- Department of Internal Medicine and Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Ronald J Buckanovich
- Department of Internal Medicine and Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.,Division of Gynecologic Oncology, Department of Obstetrics, Gynecology, and Reproductive Sciences, UPMC Hillman Cancer Center, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
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25
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Park YJ, Yoo SA, Kim M, Kim WU. The Role of Calcium-Calcineurin-NFAT Signaling Pathway in Health and Autoimmune Diseases. Front Immunol 2020; 11:195. [PMID: 32210952 PMCID: PMC7075805 DOI: 10.3389/fimmu.2020.00195] [Citation(s) in RCA: 213] [Impact Index Per Article: 42.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2019] [Accepted: 01/24/2020] [Indexed: 01/05/2023] Open
Abstract
Calcium (Ca2+) is an essential signaling molecule that controls a wide range of biological functions. In the immune system, calcium signals play a central role in a variety of cellular functions such as proliferation, differentiation, apoptosis, and numerous gene transcriptions. During an immune response, the engagement of T-cell and B-cell antigen receptors induces a decrease in the intracellular Ca2+ store and then activates store-operated Ca2+ entry (SOCE) to raise the intracellular Ca2+ concentration, which is mediated by the Ca2+ release-activated Ca2+ (CRAC) channels. Recently, identification of the two critical regulators of the CRAC channel, stromal interaction molecule (STIM) and Orai1, has broadened our understanding of the regulatory mechanisms of Ca2+ signaling in lymphocytes. Repetitive or prolonged increase in intracellular Ca2+ is required for the calcineurin-mediated dephosphorylation of the nuclear factor of an activated T cell (NFAT). Recent data indicate that Ca2+-calcineurin-NFAT1 to 4 pathways are dysregulated in autoimmune diseases. Therefore, calcineurin inhibitors, cyclosporine and tacrolimus, have been used for the treatment of such autoimmune diseases as systemic lupus erythematosus and rheumatoid arthritis. Here, we review the role of the Ca2+-calcineurin–NFAT signaling pathway in health and diseases, focusing on the STIM and Orai1, and discuss the deregulated calcium-mediated calcineurin-NFAT pathway in autoimmune diseases.
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Affiliation(s)
- Yune-Jung Park
- POSTEC-CATHOLIC Biomedical Engineering Institute, The Catholic University of Korea, Seoul, South Korea.,Division of Rheumatology, Department of Internal Medicine, St. Vincent's Hospital, The Catholic University of Korea, Suwon, South Korea
| | - Seung-Ah Yoo
- POSTEC-CATHOLIC Biomedical Engineering Institute, The Catholic University of Korea, Seoul, South Korea.,Department of Biomedicine & Health Science, College of Medicine, The Catholic University of Korea, Seoul, South Korea
| | - Mingyo Kim
- Division of Rheumatology, Department of Internal Medicine, Gyeonsang National University Hospital, Jinju, South Korea
| | - Wan-Uk Kim
- POSTEC-CATHOLIC Biomedical Engineering Institute, The Catholic University of Korea, Seoul, South Korea.,Department of Biomedicine & Health Science, College of Medicine, The Catholic University of Korea, Seoul, South Korea.,Division of Rheumatology, Department of Internal Medicine, The Catholic University of Korea, Seoul, South Korea
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26
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Solovey M, Wang Y, Michel C, Metzeler KH, Herold T, Göthert JR, Ellenrieder V, Hessmann E, Gattenlöhner S, Neubauer A, Pavlinic D, Benes V, Rupp O, Burchert A. Nuclear factor of activated T-cells, NFATC1, governs FLT3 ITD-driven hematopoietic stem cell transformation and a poor prognosis in AML. J Hematol Oncol 2019; 12:72. [PMID: 31286998 PMCID: PMC6615262 DOI: 10.1186/s13045-019-0765-y] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2019] [Accepted: 06/30/2019] [Indexed: 12/29/2022] Open
Abstract
Background Acute myeloid leukemia (AML) patients with a high allelic burden of an internal tandem duplication (ITD)-mutated FMS-like Tyrosine Kinase-3 (FLT3) have a dismal outcome. FLT3ITD triggers the proliferation of the quiescent hematopoietic stem cell (HSC) pool but fails to directly transform HSCs. While the inflammatory transcription factor nuclear factor of activated T-cells 2 (NFAT2, NFATC1) is overexpressed in AML, it is unknown whether it plays a role in FLT3ITD-induced HSC transformation. Methods We generated a triple transgenic mouse model, in which tamoxifen-inducible Cre-recombinase targets expression of a constitutively nuclear transcription factor NFATC1 to FLT3ITD positive HSC. Emerging genotypes were phenotypically, biochemically, and also transcriptionally characterized using RNA sequencing. We also retrospectively analyzed the overall survival of AML patients with different NFATC1 expression status. Results We find that NFATC1 governs FLT3ITD-driven precursor cell expansion and transformation, causing a fully penetrant lethal AML. FLT3ITD/NFATC1-AML is re-transplantable in secondary recipients and shows primary resistance to the FLT3ITD-kinase inhibitor quizartinib. Mechanistically, NFATC1 rewires FLT3ITD-dependent signaling output in HSC, involving augmented K-RAS signaling and a selective de novo recruitment of key HSC-transforming signaling pathways such as the Hedgehog- and WNT/B-Catenin signaling pathways. In human AML, NFATC1 overexpression is associated with poor overall survival. Conclusions NFATC1 expression causes FLT3ITD-induced transcriptome changes, which are associated with HSC transformation, quizartinib resistance, and a poor prognosis in AML. Electronic supplementary material The online version of this article (10.1186/s13045-019-0765-y) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Maria Solovey
- Department of Hematology, Oncology and Immunology, University Hospital Giessen and Marburg, Campus Marburg, Philipps University Marburg, Marburg, Germany
| | - Ying Wang
- Department of Hematology, Oncology and Immunology, University Hospital Giessen and Marburg, Campus Marburg, Philipps University Marburg, Marburg, Germany
| | - Christian Michel
- Department of Hematology, Oncology and Immunology, University Hospital Giessen and Marburg, Campus Marburg, Philipps University Marburg, Marburg, Germany
| | - Klaus H Metzeler
- Laboratory for Leukemia Diagnostics, Department of Medicine III, University Hospital, LMU Munich, Munich, Germany
| | - Tobias Herold
- Laboratory for Leukemia Diagnostics, Department of Medicine III, University Hospital, LMU Munich, Munich, Germany
| | | | - Volker Ellenrieder
- Department of Gastroenterology, University Hospital Goettingen, Goettingen, Germany
| | - Elisabeth Hessmann
- Department of Gastroenterology, University Hospital Goettingen, Goettingen, Germany
| | | | - Andreas Neubauer
- Department of Hematology, Oncology and Immunology, University Hospital Giessen and Marburg, Campus Marburg, Philipps University Marburg, Marburg, Germany
| | - Dinko Pavlinic
- Genomics Core Facility, EMBL Heidelberg , Heidelberg, Germany
| | - Vladimir Benes
- Genomics Core Facility, EMBL Heidelberg , Heidelberg, Germany
| | - Oliver Rupp
- Department of Bioinformatics and Systems Biology, University Giessen, Giessen, Germany
| | - Andreas Burchert
- Department of Hematology, Oncology and Immunology, University Hospital Giessen and Marburg, Campus Marburg, Philipps University Marburg, Marburg, Germany.
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Effect of recombinant human vascular endothelial growth factor on testis tissue xenotransplants from prepubertal boys: a three-case study. Reprod Biomed Online 2019; 39:119-133. [DOI: 10.1016/j.rbmo.2019.02.012] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2018] [Revised: 02/15/2019] [Accepted: 02/19/2019] [Indexed: 11/23/2022]
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Lee EH, Kim S, Choi MS, Park SM, Moon KS, Yoon S, Oh JH. Inhibition of PPARα target genes during cyclosporine A-induced nephrotoxicity and hepatotoxicity. Mol Cell Toxicol 2019. [DOI: 10.1007/s13273-019-0022-z] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
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Duck IL-2 promoter cloning and the effects of methylation status on mRNA levels in immune tissues. Cent Eur J Immunol 2019; 43:389-398. [PMID: 30799986 PMCID: PMC6384428 DOI: 10.5114/ceji.2018.81350] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2016] [Accepted: 01/10/2017] [Indexed: 12/23/2022] Open
Abstract
Interleukin 2 (IL-2), a cytokine, plays an important role in animal immune systems. To investigate the influences of epigenetic modifications on transcription of the duck IL-2 gene, the promoter region of the duck IL-2 gene was cloned. Then, the DNA methylation status of the IL-2 gene promoter (-1337 bp/-924 bp) in immune tissues of ducks was determined using the Sequenom Mass Array methylation technique, and their corresponding expression levels were determined using real-time PCR. The results showed that 2850 bp of the duck IL-2 gene promoter region were obtained. There was one CpG island (-1231 bp/-902 bp) in which 11 CpG sites were distributed. The CpG1 and CpG2 sites are located between the binding sites of NFAT and AP-1, and they had higher homology methylation patterns in different individuals and tissues. The methylation frequencies of 28.5% CpG sites showed negative correlations with the expression levels of the IL-2 mRNA, whereas 71.5% showed positive correlations. These results indicate that the transcription of duck IL-2 may be distinct from that of mammals. CpG1 (-1284 bp) and CpG2 (-1264 bp) in the duck IL-2 promoter showed a higher homology of methylation patterns, indicating a similar regulatory effect on their gene expression, and these CpG sites may be essential for the regulation of transcription of duck IL-2. The methylation pattern of the IL-2 gene promoter in duck was tissue specific.
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Yamaguchi K. Tacrolimus treatment for infertility related to maternal-fetal immune interactions. Am J Reprod Immunol 2019; 81:e13097. [PMID: 30689243 DOI: 10.1111/aji.13097] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2018] [Revised: 01/15/2019] [Accepted: 01/16/2019] [Indexed: 12/29/2022] Open
Abstract
Many approaches have been used to achieve successful pregnancies in patients with infertility, though existing treatments remain unsatisfactory in patients with infertility caused by abnormal maternal-fetal immunity. However, our understanding of the immunological aspects of infertility has steadily progressed, aided by recent research into organ transplantation and cancer. The results of these recent analyses have led to the development and evaluation of several candidate immunological treatments, but the use of immunological treatments remains a novel approach. The current paper presents the hypothesis that tacrolimus may have potential as a candidate agent for the treatment of maternal-fetal immunity-related infertility.
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Affiliation(s)
- Koushi Yamaguchi
- Center of Maternal-Fetal, Neonatal and Reproductive Medicine, National Center for Child Health and Development, Tokyo, Japan
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31
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Schober R, Waldherr L, Schmidt T, Graziani A, Stilianu C, Legat L, Groschner K, Schindl R. STIM1 and Orai1 regulate Ca 2+ microdomains for activation of transcription. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2018; 1866:1079-1091. [PMID: 30408546 DOI: 10.1016/j.bbamcr.2018.11.001] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/30/2018] [Revised: 10/31/2018] [Accepted: 11/01/2018] [Indexed: 02/07/2023]
Abstract
Since calcium (Ca2+) regulates a large variety of cellular signaling processes in a cell's life, precise control of Ca2+ concentrations within the cell is essential. This enables the transduction of information via Ca2+ changes in a time-dependent and spatially defined manner. Here, we review molecular and functional aspects of how the store-operated Ca2+ channel Orai1 creates spatiotemporal Ca2+ microdomains. The architecture of this channel is unique, with a long helical pore and a six-fold symmetry. Energetic barriers within the Ca2+ channel pathway limit permeation to allow an extensive local Ca2+ increase in close proximity to the channel. The precise timing of the Orai1 channel function is controlled by direct binding to STIM proteins upon Ca2+ depletion in the endoplasmic reticulum. These induced Ca2+ microdomains are tailored to, and sufficient for, triggering long-term activation processes, such as transcription factor activation and subsequent gene regulation. We describe the principles of spatiotemporal activation of the transcription factor NFAT and compare its signaling characteristics to those of the autophagy regulating transcription factors, MITF and TFEB.
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Affiliation(s)
- Romana Schober
- Institute for Biophysics, Johannes Kepler University Linz, A-4040 Linz, Austria.
| | - Linda Waldherr
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Tony Schmidt
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Annarita Graziani
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Clemens Stilianu
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Lorenz Legat
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Klaus Groschner
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria
| | - Rainer Schindl
- Gottfried Schatz Research Center, Medical University of Graz, A-8010 Graz, Austria.
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32
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Wang Q, Gregg JR, Gu J, Ye Y, Chang DW, Davis JW, Thompson TC, Kim J, Logothetis CJ, Wu X. Genetic associations of T cell cancer immune response with tumor aggressiveness in localized prostate cancer patients and disease reclassification in an active surveillance cohort. Oncoimmunology 2018; 8:e1483303. [PMID: 30546938 DOI: 10.1080/2162402x.2018.1483303] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2018] [Revised: 05/21/2018] [Accepted: 05/26/2018] [Indexed: 12/23/2022] Open
Abstract
Determining prostate cancer (PCa) aggressiveness and reclassification are critical events during the treatment of localized disease and for patients undergoing active surveillance (AS). Since T cells play major roles in cancer surveillance and elimination, we aimed to identify genetic biomarkers related to T cell cancer immune response which are predictive of aggressiveness and reclassification risks in localized PCa. The genotypes of 3,586 single nucleotide polymorphisms (SNPs) from T cell cancer immune response pathways were analyzed in 1762 patients with localized disease and 393 who elected AS. The aggressiveness of PCa was defined according to pathological Gleason score (GS) and D'Amico criteria. PCa reclassification was defined according to changes in GS or tumor characteristics during subsequent surveillance biopsies. Functional characterization and analysis of immune phenotypes were also performed. In the localized PCa cohort, seven SNPs were significantly associated with the risk of aggressive disease. In the AS cohort, another eight SNPs were identified as predictors for aggressiveness and reclassification. Rs1687016 of PSMB8 was the most significant predictor of reclassification. Cumulative analysis showed that a genetic score based on the identified SNPs could significantly predict risk of D'Amico high risk disease (P-trend = 2.4E-09), GS4 + 3 disease (P-trend = 1.3E-04), biochemical recurrence (P-trend = 0.01) and reclassification (P-trend = 0.01). In addition, the rs34309 variant was associated with functional somatic mutations in the PI3K/PTEN/AKT/MTOR pathway and tumor lymphocyte infiltration. Our study provides plausible evidence that genetic variations in T cell cancer immune response can influence risks of aggressiveness and reclassification in localized PCa, which may lead to additional biological insight into these outcomes. Abbreviations: PCa, prostate cancer; AS, active surveillance; GS, Gleason score; PSA, prostate specific antigen; TCGA, The Cancer Genome Atlas; SNP, single nucleotide polymorphisms; UFG, unfavorable genotype.
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Affiliation(s)
- Qinchuan Wang
- Departments of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.,Department of Surgical Oncology, Affiliated Sir Run Run Shaw Hospital, Zhejiang University, Hangzhou, China
| | - Justin R Gregg
- Departments of Urology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Jian Gu
- Departments of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Yuanqing Ye
- Departments of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - David W Chang
- Departments of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - John W Davis
- Departments of Urology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Timothy C Thompson
- Genitourinary Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Jeri Kim
- Departments of Urology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Christopher J Logothetis
- Genitourinary Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Xifeng Wu
- Departments of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
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Chen L, Wang H, Gao F, Zhang J, Zhang Y, Ma R, Pang S, Cui Y, Yang J, Yan B. Functional genetic variants in the SIRT5 gene promoter in acute myocardial infarction. Gene 2018; 675:233-239. [DOI: 10.1016/j.gene.2018.07.010] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2018] [Revised: 06/14/2018] [Accepted: 07/03/2018] [Indexed: 01/03/2023]
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Hellesen A, Bratland E. The potential role for infections in the pathogenesis of autoimmune Addison's disease. Clin Exp Immunol 2018; 195:52-63. [PMID: 30144040 DOI: 10.1111/cei.13207] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2018] [Revised: 08/02/2018] [Accepted: 08/10/2018] [Indexed: 12/21/2022] Open
Abstract
Autoimmune Addison's disease (AAD), or primary adrenocortical insufficiency, is a classical organ-specific autoimmune disease with 160 years of history. AAD is remarkably homogeneous with one major dominant self-antigen, the cytochrome P450 21-hydroxylase enzyme, which is targeted by both autoantibodies and autoreactive T cells. Like most autoimmune diseases, AAD is thought to be caused by an unfortunate combination of genetic and environmental factors. While the number of genetic associations with AAD is increasing, almost nothing is known about environmental factors. A major environmental factor commonly proposed for autoimmune diseases, based partly on experimental and clinical data and partly on shared pathways between anti-viral immunity and autoimmunity, is viral infections. However, there are few reports associating viral infections to AAD, and it has proved difficult to establish which immunological processes that could link any viral infection with the initiation or progression of AAD. In this review, we will summarize the current knowledge on the underlying mechanisms of AAD and take a closer look on the potential involvement of viruses.
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Affiliation(s)
- A Hellesen
- Department of Clinical Science, University of Bergen, Bergen, Norway.,K.G. Jebsen Senter for Autoimmune Sykdommer, University of Bergen, Bergen, Norway
| | - E Bratland
- Department of Clinical Science, University of Bergen, Bergen, Norway.,K.G. Jebsen Senter for Autoimmune Sykdommer, University of Bergen, Bergen, Norway
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35
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Screening for genes that regulate the differentiation of human megakaryocytic lineage cells. Proc Natl Acad Sci U S A 2018; 115:E9308-E9316. [PMID: 30150396 DOI: 10.1073/pnas.1805434115] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023] Open
Abstract
Different combinations of transcription factors (TFs) function at each stage of hematopoiesis, leading to distinct expression patterns of lineage-specific genes. The identification of such regulators and their functions in hematopoiesis remain largely unresolved. In this study, we utilized screening approaches to study the transcriptional regulators of megakaryocyte progenitor (MkP) generation, a key step before platelet production. Promising candidate genes were generated from a microarray platform gene expression commons and individually manipulated in human hematopoietic stem and progenitor cells (HSPCs). Deletion of some of the candidate genes (the hit genes) by CRISPR/Cas9 led to decreased MkP generation during HSPC differentiation, while more MkPs were produced when some hit genes were overexpressed in HSPCs. We then demonstrated that overexpression of these genes can increase the frequency of mature megakaryocytic colonies by functional colony forming unit-megakaryocyte (CFU-Mk) assay and the release of platelets after in vitro maturation. Finally, we showed that the histone deacetylase inhibitors could also increase MkP differentiation, possibly by regulating some of the newly identified TFs. Therefore, identification of such regulators will advance the understanding of basic mechanisms of HSPC differentiation and conceivably enable the generation and maturation of megakaryocytes and platelets in vitro.
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36
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Moon JS, Lee HJ, Ho CC, Shin JS, Ghosh S, Kim JH, Lee SK. Immuno-suppressive function of nucleus-transducible BAF57-ΔPH in T cell activation via degradation of endogenous BAF57. Int J Hematol 2018; 108:375-383. [PMID: 29978433 DOI: 10.1007/s12185-018-2491-6] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2018] [Revised: 06/26/2018] [Accepted: 06/26/2018] [Indexed: 11/28/2022]
Abstract
The BAF57 subunit, an indispensable member of the BAF complex, is functionally implicated in apoptosis, cell cycle, and T cell development through chromosomal remodeling. However, the precise roles of BAF57 in the T cell receptor (TcR)-mediated signaling pathway have not been elucidated. In this study, a nucleus-transducible form of BAF57, absent the proline-rich and HMG domains (ntBAF57-ΔPH), was generated to interfere with the interaction between BAF57 and its binding protein, BAF155. ntBAF57-ΔPH was effectively delivered into mouse CD4+ T cells in a dose- and time-dependent manner, without cellular toxicity. Inhibition of T cell activation by ntBAF57-ΔPH was mediated by its disruption of the interaction between BAF155 and BAF57, leading to the degradation of endogenous BAF57 and BAF155. This phenomenon led to alterations in gene expression similar to those associated with Ciclosporin A treatment. In vivo administration of ntBAF57-ΔPH enhanced survival rate of sepsis-induced mice and reduced the LPS-induced secretion of pro-inflammatory cytokines and the expression of endogenous BAF57. These results reveal a novel function of BAF57 as an essential regulator of T cell activation. ntBAF57-ΔPH represents a novel immune-suppressive drug candidate with potential uses in the treatment of autoimmunity and graft rejection.
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Affiliation(s)
- Jae-Seung Moon
- Department of Biotechnology, Yonsei University College of Life Science and Biotechnology, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea
| | - Hong-Jai Lee
- Department of Biotechnology, Yonsei University College of Life Science and Biotechnology, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea
| | - Chun-Chang Ho
- Department of Biotechnology, Yonsei University College of Life Science and Biotechnology, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea
| | - Jin-Su Shin
- Department of Biotechnology, Yonsei University College of Life Science and Biotechnology, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea
| | - Sankar Ghosh
- Department of Microbiology and Immunology, Columbia University, College of Physicians and Surgeons, New York, NY, USA
| | - Jung-Ho Kim
- Good T cells, Inc, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea
| | - Sang-Kyou Lee
- Department of Biotechnology, Yonsei University College of Life Science and Biotechnology, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea. .,Good T cells, Inc, 50 Yonsei-ro, Seodaemun-gu, Seoul, Republic of Korea.
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37
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Wu X, Tommasi di Vignano A, Zhou Q, Michel-Dziunycz PJ, Bai F, Mi J, Qin J, Zu T, Hofbauer GFL. The ARE-binding protein Tristetraprolin (TTP) is a novel target and mediator of calcineurin tumor suppressing function in the skin. PLoS Genet 2018; 14:e1007366. [PMID: 29723192 PMCID: PMC5953486 DOI: 10.1371/journal.pgen.1007366] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2017] [Revised: 05/15/2018] [Accepted: 04/16/2018] [Indexed: 01/12/2023] Open
Abstract
An increased incidence of skin inflammatory diseases is frequently observed in organtransplanted patients being treated with calcineurin inhibitor-based immunosuppressive agents. The mechanism of increased skin inflammation in this context has however not yet been clarified. Here we report an increased inflammation following inhibition of calcineurin signaling seen in both chemically induced mouse skin tumors and in tumors grafted from H-rasV12 expressing primary human keratinocytes (HKCs). Following UVB or TPA treatment, we specifically found that deletion of the calcineurin gene in mouse keratinocytes (MKCs) resulted in increased inflammation, and this was accompanied by the enhanced production of pro-inflammatory cytokines, such as TNFα, IL-8 and CXCL1. Furthermore, expression of the RNA-binding protein, tristetraprolin (TTP) was down-regulated in response to calcineurin inhibition, wherein TTP was shown to negatively regulate the production of pro-inflammatory cytokines in keratinocytes. The induction of TTP following TPA or UVB treatment was attenuated by calcineurin inhibition in keratinocytes, and correspondingly, disruption of calcineurin signaling down-regulated the amounts of TTP in both clinical and H-rasV12-transformed keratinocyte tumor models. Our results further demonstrated that calcineurin positively controls the stabilization of TTP in keratinocytes through a proteasome-dependent mechanism. Reducing the expression of TTP functionally promoted tumor growth of H-rasV12 expressing HKCs, while stabilizing TTP expression counteracted the tumor-promoting effects of calcineurin inhibition. Collectively these results suggest that calcineurin signaling, acting through TTP protein level stabilization, suppresses keratinocyte tumors by downregulating skin inflammation.
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Affiliation(s)
- Xunwei Wu
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
- Cutaneous Biology Research Centre, Massachusetts General Hospital, Charlestown, MA, United States of America
- * E-mail:
| | - Alice Tommasi di Vignano
- Cutaneous Biology Research Centre, Massachusetts General Hospital, Charlestown, MA, United States of America
| | - Qian Zhou
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
| | | | - Fuxiang Bai
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
| | - Jun Mi
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
| | - Jing Qin
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
| | - Tingjian Zu
- Laboratory for Tissue Engineering and Regeneration and Shandong Provincial Key Laboratory of Oral Tissue Regeneration, School of Stomatology, Shandong University, Jinan, Shandong, China
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Oxysterol-binding protein-related protein 4L promotes cell proliferation by sustaining intracellular Ca2+ homeostasis in cervical carcinoma cell lines. Oncotarget 2018; 7:65849-65861. [PMID: 27588468 PMCID: PMC5323197 DOI: 10.18632/oncotarget.11671] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2016] [Accepted: 08/09/2016] [Indexed: 12/30/2022] Open
Abstract
Oxsterol binding protein-related protein 4 (ORP4) is essential for cell proliferation, but the underlying mechanism is unclear. ORP4 is expressed as three variants, ORP4L, ORP4M and ORP4S. Here, we reported that silencing of ORP4L with specific small interfering RNA (siRNA) inhibited the proliferation of human cervical cancer cell lines C33A, HeLa and CaSki, the reverse effect being observed in ORP4L overexpressing cells. For molecular insight, we found that ORP4L maintained intracellular Ca2+ homeostasis. Through this mechanism, ORP4L activated nuclear factor of activated T cells (NFAT) activity and thus promoted expression of a gene cluster which supported cell proliferation. Of note, ORP4L sustained inositol-1,4,5-trisphosphate receptor 1 (IP3R1) expression at both mRNA and protein levels via Ca2+-dependent NFAT3 activation, which offered a mechanic explanation for the role of ORP4L intracellular Ca2+ homeostasis. Furthermore, ORP4L knockdown markedly inhibited tumor growth in a C33A cell xenograft mouse model. To conclude, our results reveal that ORP4L promotes cell proliferation through maintaining intracellular Ca2+ homeostasis.
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Abstract
Nuclear factor of activated T cells (NFAT) was first described almost three decades ago as a Ca
2+/calcineurin-regulated transcription factor in T cells. Since then, a large body of research uncovered the regulation and physiological function of different NFAT homologues in the immune system and many other tissues. In this review, we will discuss novel roles of NFAT in T cells, focusing mainly on its function in humoral immune responses, immunological tolerance, and the regulation of immune metabolism.
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Affiliation(s)
- Martin Vaeth
- Department of Pathology, New York University School of Medicine, New York, NY, 10016, USA
| | - Stefan Feske
- Department of Pathology, New York University School of Medicine, New York, NY, 10016, USA
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40
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Cho HJ, Yun HJ, Yang HC, Kim SJ, Kang SK, Che C, Lee SD, Kang MW. Prognostic significance of nuclear factor of activated T-cells 5 expression in non-small cell lung cancer patients who underwent surgical resection. J Surg Res 2018; 226:40-47. [PMID: 29661287 DOI: 10.1016/j.jss.2017.12.036] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2017] [Revised: 11/30/2017] [Accepted: 12/28/2017] [Indexed: 12/25/2022]
Abstract
BACKGROUND Nuclear factor of activated T-cells 5 (NFAT5) is known to be correlated with migration or invasion of tumor cells based on previous in vitro studies. The aim of this study was to analyze the relationship between NFAT5 expression and clinical prognosis in non-small cell lung cancer (NSCLC) patients who underwent surgical resection. MATERIALS AND METHODS A total of 92 NSCLC patients who underwent surgical resection were enrolled. The tissue microarray core was obtained from surgically resected tumor specimens. NFAT5 expression was evaluated by immunohistochemistry. Relationships of NFAT5 expression with disease recurrence, overall survival, and disease-free survival (DFS) were analyzed. RESULTS The mean age of 92 patients was 63.7 y. The median follow-up duration was 63.3 mo. Fifty-one (55%) patients exhibited positive expression of NFAT5. Disease recurrence in the NFAT5-positive group was significantly (P = 0.022) higher than that in the NFAT5-negative group. NFAT5-positive expression (odds ratio: 2.632, 95% confidence interval: 1.071-6.465, P = 0.035) and pathologic N stage (N1-2 versus N0; odds ratio: 3.174, 95% confidence interval: 1.241-8.123, P = 0.016) were independent and significant risk factors for disease recurrence. DFS of the NFAT5-positive group was significantly worse than that of the NFAT5-negative group (89.7 versus 48.7 mo, P = 0.011). A multivariate analysis identified NFAT5 expression (P < 0.029) as a significant independent risk factor for DFS of patients with postoperative pathologic T and N stages (P < 0.001 and P = 0.017, respectively). CONCLUSIONS NFAT5 expression is a useful prognostic biomarker for NSCLC patients who underwent surgical resection.
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Affiliation(s)
- Hyun Jin Cho
- Department of Thoracic and Cardiovascular Surgery, Chungnam National University Hospital, Chungnam National University School of Medicine, Daejeon, South Korea
| | - Hwan-Jung Yun
- Department of Internal Medicine, Chungnam National University School of Medicine, Daejeon, South Korea
| | - Hee Chul Yang
- Department of Thoracic and Cardiovascular Surgery, Center for Lung Cancer, Research Institute and Hospital, National Cancer Center, Goyang, South Korea
| | - Soo Jin Kim
- Department of Physiology, Chungnam National University School of Medicine, Daejeon, South Korea
| | - Shin Kwang Kang
- Department of Thoracic and Cardiovascular Surgery, Chungnam National University Hospital, Chungnam National University School of Medicine, Daejeon, South Korea
| | - Chengri Che
- Department of Thoracic Surgery, Yanbian University Hospital, Yanji, China
| | - Sang Do Lee
- Department of Physiology, Chungnam National University School of Medicine, Daejeon, South Korea.
| | - Min-Woong Kang
- Department of Thoracic and Cardiovascular Surgery, Chungnam National University Hospital, Chungnam National University School of Medicine, Daejeon, South Korea.
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Jung WH, Son YE, Oh SH, Fu C, Kim HS, Kwak JH, Cardenas ME, Heitman J, Park HS. Had1 Is Required for Cell Wall Integrity and Fungal Virulence in Cryptococcus neoformans. G3 (BETHESDA, MD.) 2018; 8:643-652. [PMID: 29233914 PMCID: PMC5919746 DOI: 10.1534/g3.117.300444] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/16/2017] [Accepted: 12/07/2017] [Indexed: 12/16/2022]
Abstract
Calcineurin modulates environmental stress survival and virulence of the human fungal pathogen Cryptococcus neoformans Previously, we identified 44 putative calcineurin substrates, and proposed that the calcineurin pathway is branched to regulate targets including Crz1, Pbp1, and Puf4 in C. neoformans In this study, we characterized Had1, which is one of the putative calcineurin substrates belonging to the ubiquitously conserved haloacid dehalogenase β-phosphoglucomutase protein superfamily. Growth of the had1∆ mutant was found to be compromised at 38° or higher. In addition, the had1∆ mutant exhibited increased sensitivity to cell wall perturbing agents, including Congo Red and Calcofluor White, and to an endoplasmic reticulum stress inducer dithiothreitol. Virulence studies revealed that the had1 mutation results in attenuated virulence compared to the wild-type strain in a murine inhalation infection model. Genetic epistasis analysis revealed that Had1 and the zinc finger transcription factor Crz1 play roles in parallel pathways that orchestrate stress survival and fungal virulence. Overall, our results demonstrate that Had1 is a key regulator of thermotolerance, cell wall integrity, and virulence of C. neoformans.
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Affiliation(s)
- Won-Hee Jung
- School of Food Science and Biotechnology, Institute of Agricultural Science and Technology, Kyungpook National University, Daegu 41566, Republic of Korea
| | - Ye-Eun Son
- School of Food Science and Biotechnology, Institute of Agricultural Science and Technology, Kyungpook National University, Daegu 41566, Republic of Korea
| | - Sang-Hun Oh
- School of Life Science, Handong Global University, Pohang 37554, Republic of Korea
| | - Ci Fu
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710
| | - Hye Shin Kim
- School of Life Science, Handong Global University, Pohang 37554, Republic of Korea
| | - Jin-Hwan Kwak
- School of Life Science, Handong Global University, Pohang 37554, Republic of Korea
| | - Maria E Cardenas
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710
| | - Joseph Heitman
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710
| | - Hee-Soo Park
- School of Food Science and Biotechnology, Institute of Agricultural Science and Technology, Kyungpook National University, Daegu 41566, Republic of Korea
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42
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Patterson AR, Endale M, Lampe K, Aksoylar HI, Flagg A, Woodgett JR, Hildeman D, Jordan MB, Singh H, Kucuk Z, Bleesing J, Hoebe K. Gimap5-dependent inactivation of GSK3β is required for CD4 + T cell homeostasis and prevention of immune pathology. Nat Commun 2018; 9:430. [PMID: 29382851 PMCID: PMC5789891 DOI: 10.1038/s41467-018-02897-7] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2017] [Accepted: 01/08/2018] [Indexed: 12/16/2022] Open
Abstract
GTPase of immunity-associated protein 5 (Gimap5) is linked with lymphocyte survival, autoimmunity, and colitis, but its mechanisms of action are unclear. Here, we show that Gimap5 is essential for the inactivation of glycogen synthase kinase-3β (GSK3β) following T cell activation. In the absence of Gimap5, constitutive GSK3β activity constrains c-Myc induction and NFATc1 nuclear import, thereby limiting productive CD4+ T cell proliferation. Additionally, Gimap5 facilitates Ser389 phosphorylation and nuclear translocation of GSK3β, thereby limiting DNA damage in CD4+ T cells. Importantly, pharmacological inhibition and genetic targeting of GSK3β can override Gimap5 deficiency in CD4+ T cells and ameliorates immunopathology in mice. Finally, we show that a human patient with a GIMAP5 loss-of-function mutation has lymphopenia and impaired T cell proliferation in vitro that can be rescued with GSK3 inhibitors. Given that the expression of Gimap5 is lymphocyte-restricted, we propose that its control of GSK3β is an important checkpoint in lymphocyte proliferation. Loss of function GIMAP5 mutation is associated with lymphopenia, but how it mediates T cell homeostasis is unclear. Here the authors study Gimap5−/− mice and a patient with GIMAP5 deficiency to show how this GTPAse negatively regulates GSK3β activity to prevent DNA damage and cell death.
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Affiliation(s)
- Andrew R Patterson
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA.,Immunology Graduate Program, Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, 231 Albert Sabin Way # E251n, Cincinnati, OH, 45267, USA
| | - Mehari Endale
- Division of Neonatology and Pulmonary Biology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA
| | - Kristin Lampe
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA
| | - Halil I Aksoylar
- Department of Genetics and Complex Diseases, Harvard T.H. Chan School of Public Health, 677 Huntington Avenue, Boston, MA, 02115, USA
| | - Aron Flagg
- Pediatric Hematology/Oncology and Blood & Marrow Transplant, Cleveland Clinic Children's, 9500 Euclid Avenue, Cleveland, OH, 44195, USA
| | - Jim R Woodgett
- The Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, ON, M5G 1X5, Canada
| | - David Hildeman
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA.,Immunology Graduate Program, Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, 231 Albert Sabin Way # E251n, Cincinnati, OH, 45267, USA
| | - Michael B Jordan
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA.,Immunology Graduate Program, Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, 231 Albert Sabin Way # E251n, Cincinnati, OH, 45267, USA
| | - Harinder Singh
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA.,Immunology Graduate Program, Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, 231 Albert Sabin Way # E251n, Cincinnati, OH, 45267, USA
| | - Zeynep Kucuk
- Division of Bone Marrow Transplantation & Immune Deficiency, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA
| | - Jack Bleesing
- Division of Bone Marrow Transplantation & Immune Deficiency, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA
| | - Kasper Hoebe
- Division of Immunobiology, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH, 45229, USA. .,Immunology Graduate Program, Cincinnati Children's Hospital Medical Center and the University of Cincinnati College of Medicine, 231 Albert Sabin Way # E251n, Cincinnati, OH, 45267, USA. .,Department of Pediatrics, University of Cincinnati, College of Medicine, 3230 Eden Avenue, Cincinnati, OH, 45267, USA.
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43
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Angulo C, Alamillo E, Ascencio F, Reyes-Becerril M. Characterization of nuclear factor of activated T-cells-c3 (NFATc3) and gene expression of upstream-downstream signaling molecules in response to immunostimulants in Pacific red snapper cells. DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY 2018; 78:149-159. [PMID: 28986213 DOI: 10.1016/j.dci.2017.10.001] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/21/2017] [Revised: 10/01/2017] [Accepted: 10/01/2017] [Indexed: 06/07/2023]
Abstract
The nuclear factor of activated T cells (NFAT) proteins have crucial roles in the development and function of the immune system since they not only regulate activation of T cells but are also involved in the control of thymocyte development and T-cell differentiation. In this study, NFATc3 was characterized from the Pacific red snapper, Lutjanus peru. LpNFAtc3, which contains an open reading of 3300 bp frame coding for a protein of 1100 aa with a predicted molecular weight of 118.52 kDa. The predicted protein showed a conserved NFAT family structure with signature motifs and domains, sharing high identity (up to 76%) compared to other fish sequences. NFATc3 gene expression was analyzed by real time-PCR in head-kidney cells (leukocytes and lymphocytes) following yeast, zymosan and Vibrio parahaemolyticus stimulation along with the expression of upstream (ILF2, ILF3 and CaN) and downstream (CD3, TCRβ, IL-6 and IL-12) molecules. This study revealed a broad expression of NFATc3 with a relative strong expression in intestine and lymphocytes. The expression of NFATc3 was differentially up-regulated after stimulation with yeast in head-kidney leukocytes and after bacterial infection in lymphocytes at 24 h. Interestingly, the yeast and zymosan were able to activate ILF2, ILF3 and CaN mRNA gene expression in both kinds of cells. On the other hand, NFAT downstream genes such as CD3, TCRβ, IL-6 and IL-12 were significantly up-regulated in leukocytes stimulated with yeast or zymosan at 12 h; however in lymphocytes, this up-regulation was detected when cells reacted to V. parahaemolyticus stimuli at 24 h. Stimulating Pacific red snapper leukocytes with immunostimulants as yeast significantly up-regulated the expression of NFATc3, and up- and down-stream molecular genes and NFATc3 lymphocytes expression were potentially involved in responses to invasion of bacterial pathogens in an early immune response.
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Affiliation(s)
- Carlos Angulo
- Immunology & Vaccinology Group, Centro de Investigaciones Biológicas Del Noroeste (CIBNOR), Instituto Politécnico Nacional 195, Playa Palo de Santa Rita Sur, La Paz B.C.S. 23090, Mexico
| | - Erika Alamillo
- Immunology & Vaccinology Group, Centro de Investigaciones Biológicas Del Noroeste (CIBNOR), Instituto Politécnico Nacional 195, Playa Palo de Santa Rita Sur, La Paz B.C.S. 23090, Mexico
| | - Felipe Ascencio
- Immunology & Vaccinology Group, Centro de Investigaciones Biológicas Del Noroeste (CIBNOR), Instituto Politécnico Nacional 195, Playa Palo de Santa Rita Sur, La Paz B.C.S. 23090, Mexico
| | - Martha Reyes-Becerril
- Immunology & Vaccinology Group, Centro de Investigaciones Biológicas Del Noroeste (CIBNOR), Instituto Politécnico Nacional 195, Playa Palo de Santa Rita Sur, La Paz B.C.S. 23090, Mexico.
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44
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Dissecting the Roles of the Calcineurin Pathway in Unisexual Reproduction, Stress Responses, and Virulence in Cryptococcus deneoformans. Genetics 2017; 208:639-653. [PMID: 29233811 DOI: 10.1534/genetics.117.300422] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2017] [Accepted: 12/07/2017] [Indexed: 01/12/2023] Open
Abstract
The Ca2+/calmodulin-dependent protein phosphatase calcineurin orchestrates sexual reproduction, stress responses, and virulence via branched downstream pathways in the opportunistic human fungal pathogen Cryptococcus neoformans The calcineurin-binding protein Cbp1, the calcineurin temperature suppressor Cts1, the calcineurin-responsive zinc finger transcription factor Crz1, and the calcineurin targets Pbp1, Tif3, and Puf4, all function downstream of calcineurin to orchestrate distinct cellular processes. To elucidate how the calcineurin pathway regulatory network governs unisexual reproduction, stress responses, and virulence, we have analyzed the self-filamentous C. deneoformans strain, XL280α, and generated double mutants of these calcineurin downstream genes. We demonstrated that calcineurin governs unisexual reproduction at different sexual developmental stages, in which the initiation of the yeast-hyphal morphological transition is independent of Crz1, whereas the sporulation process is dependent on Crz1. Calcineurin-dependent unisexual reproduction is independent of the pheromone response pathway. Crz1 synergistically interacts with different calcineurin downstream targets in responding to ER, high-calcium, and cell wall stresses. We observed a widespread synergy suggesting that these proteins function in complex branched pathways downstream of calcineurin with some functional redundancy, which may allow efficient signaling network rewiring within the pathway for prompt adaptation to changing environments. Finally, we showed that deletion of PBP1 or TIF3 in the cna1∆ mutant background conferred a modest level of growth tolerance at 37°, but that the cna1∆ pbp1∆ and cna1∆ tif3∆ double mutants were both avirulent, suggesting that calcineurin may control virulence via mechanisms beyond thermotolerance.
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45
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Frischauf I, Litviňuková M, Schober R, Zayats V, Svobodová B, Bonhenry D, Lunz V, Cappello S, Tociu L, Reha D, Stallinger A, Hochreiter A, Pammer T, Butorac C, Muik M, Groschner K, Bogeski I, Ettrich RH, Romanin C, Schindl R. Transmembrane helix connectivity in Orai1 controls two gates for calcium-dependent transcription. Sci Signal 2017; 10:eaao0358. [PMID: 29184031 PMCID: PMC6433236 DOI: 10.1126/scisignal.aao0358] [Citation(s) in RCA: 65] [Impact Index Per Article: 8.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
The channel Orai1 requires Ca2+ store depletion in the endoplasmic reticulum and an interaction with the Ca2+ sensor STIM1 to mediate Ca2+ signaling. Alterations in Orai1-mediated Ca2+ influx have been linked to several pathological conditions including immunodeficiency, tubular myopathy, and cancer. We screened large-scale cancer genomics data sets for dysfunctional Orai1 mutants. Five of the identified Orai1 mutations resulted in constitutively active gating and transcriptional activation. Our analysis showed that certain Orai1 mutations were clustered in the transmembrane 2 helix surrounding the pore, which is a trigger site for Orai1 channel gating. Analysis of the constitutively open Orai1 mutant channels revealed two fundamental gates that enabled Ca2+ influx: Arginine side chains were displaced so they no longer blocked the pore, and a chain of water molecules formed in the hydrophobic pore region. Together, these results enabled us to identify a cluster of Orai1 mutations that trigger Ca2+ permeation associated with gene transcription and provide a gating mechanism for Orai1.
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Affiliation(s)
- Irene Frischauf
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Monika Litviňuková
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
- Max Delbrück Center for Molecular Medicine in the Helmholtz Association, Cardiovascular and Metabolic Sciences, Berlin D-13125, Germany
| | - Romana Schober
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Vasilina Zayats
- Center for Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Nové Hrady CZ-373 33, Czech Republic
- Center of New Technologies, University of Warsaw, Warsaw 02-097, Poland
| | - Barbora Svobodová
- Institute for Biophysics, Medical University of Graz, Graz A-8010, Austria
| | - Daniel Bonhenry
- Center for Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Nové Hrady CZ-373 33, Czech Republic
| | - Victoria Lunz
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Sabrina Cappello
- Molecular Physiology, Institute of Cardiovascular Physiology, University Medical Center, Georg August University Göttingen, Göttingen, Niedersachsen 37073, Germany
- Department of Biophysics, Center for Integrative Physiology and Molecular Medicine, Medical Faculty, Saarland University, Homburg D-66421, Germany
| | - Laura Tociu
- Center for Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Nové Hrady CZ-373 33, Czech Republic
- University of Chicago, Chicago, IL 60637, USA
| | - David Reha
- Center for Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Nové Hrady CZ-373 33, Czech Republic
- Faculty of Sciences, University of South Bohemia, Nové Hrady CZ-373 33, Czech Republic
| | - Amrutha Stallinger
- Institute for Molecular Biosciences, Karl-Franzens-University Graz, Graz A-8010, Austria
| | - Anna Hochreiter
- Institute for Experimental and Clinical Cell Therapy, Paracelsus Medical University, Salzburg A-5020, Austria
| | - Teresa Pammer
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Carmen Butorac
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Martin Muik
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Klaus Groschner
- Institute for Biophysics, Medical University of Graz, Graz A-8010, Austria
| | - Ivan Bogeski
- Molecular Physiology, Institute of Cardiovascular Physiology, University Medical Center, Georg August University Göttingen, Göttingen, Niedersachsen 37073, Germany
| | - Rüdiger H Ettrich
- Center for Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Nové Hrady CZ-373 33, Czech Republic
- Faculty of Sciences, University of South Bohemia, Nové Hrady CZ-373 33, Czech Republic
| | - Christoph Romanin
- Institute of Biophysics, JKU Life Science Center, Johannes Kepler University Linz, Linz A-4020, Austria
| | - Rainer Schindl
- Institute for Biophysics, Medical University of Graz, Graz A-8010, Austria.
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46
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Kaminuma O, Kitamura N, Nishito Y, Nemoto S, Tatsumi H, Mori A, Hiroi T. Downregulation of NFAT3 Due to Lack of T-Box Transcription Factor TBX5 Is Crucial for Cytokine Expression in T Cells. THE JOURNAL OF IMMUNOLOGY 2017; 200:92-100. [PMID: 29180489 DOI: 10.4049/jimmunol.1602113] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/15/2016] [Accepted: 10/19/2017] [Indexed: 11/19/2022]
Abstract
The NFAT family transcription factors play crucial roles in immunological and other biological activities. NFAT3 is rarely expressed in T cells, and the mechanisms and significance of the specific NFAT3 downregulation in T cells have been unknown. In human CD4+ T cells, overexpression of NFAT1 and NFAT3 enhanced and suppressed IL-2 expression, respectively. NFAT3 downregulation in Jurkat cells using RNA interference technology augmented IL-2 expression, whereas a knockdown of NFAT1, NFAT2, and NFAT4 suppressed it. The promoter/enhancer activity of the NFAT-binding site in the IL-2 gene was upregulated and downregulated by NFAT1 and NFAT3, respectively. A study employing NFAT1/NFAT3 chimeric molecules revealed that the region in NFAT3 responsible for NFAT promoter activity inhibition was located within its N-terminal transactivation domain, Ca2+-regulatory domain, and DNA-binding domain. Downregulation of NFAT3 expression in T cells is mediated by lower chromatin accessibility and enhancer activity in its promoter in comparison with aortic smooth muscle cells expressing endogenous NFAT3. The binding sites of T-box transcription factor TBX5 and NK-2 transcription factor-related locus 5 Nkx2.5, which were expressed at higher levels in aortic smooth muscle cells than in T cells, were located within the -387 to +97 NFAT3 promoter region, exhibiting the maximum enhancer activity. Mutating the binding site of TBX5 but not Nkx2.5 diminished the NFAT3 promoter activity, whereas the overexpression of TBX5 enhanced it. Introduction of TBX5 into CD4+ T cells enhanced the expression of NFAT3 and suppressed that of IL-2. TBX5 deficiency-mediated downregulation of NFAT3 is crucial for the high cytokine-producing activity of T cells.
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Affiliation(s)
- Osamu Kaminuma
- Allergy and Immunology Project, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613, Japan; .,Center for Life Science Research, University of Yamanashi, Yamanashi 409-3898, Japan
| | - Noriko Kitamura
- Allergy and Immunology Project, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613, Japan
| | - Yasumasa Nishito
- Center for Basic Technology Research, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613, Japan
| | - Soichi Nemoto
- Department of Obstetrics and Gynecology, National Hospital Organization, Sagamihara National Hospital, Kanagawa 252-0392, Japan; and
| | - Hideki Tatsumi
- Department of Obstetrics and Gynecology, National Hospital Organization, Sagamihara National Hospital, Kanagawa 252-0392, Japan; and
| | - Akio Mori
- Clinical Research Center for Allergy and Rheumatology, National Hospital Organization, Sagamihara National Hospital, Kanagawa 252-0392, Japan
| | - Takachika Hiroi
- Allergy and Immunology Project, The Tokyo Metropolitan Institute of Medical Science, Tokyo 113-8613, Japan
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47
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Wang Z, Yang H, Si S, Han Z, Tao J, Chen H, Ge Y, Guo M, Wang K, Tan R, Wei JF, Gu M. Polymorphisms of nucleotide factor of activated T cells cytoplasmic 2 and 4 and the risk of acute rejection following kidney transplantation. World J Urol 2017; 36:111-116. [PMID: 29103109 PMCID: PMC5758697 DOI: 10.1007/s00345-017-2117-2] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2017] [Accepted: 10/27/2017] [Indexed: 12/17/2022] Open
Abstract
BACKGROUND Acute rejection (AR) is a common complication of kidney transplantation. Nuclear factors of activated T cells (NFATs) are transcription factors involved in the activation of T lymphocytes, but their association with AR is unclear. METHODS This retrospective, case-control study included 200 renal transplant recipients who were divided into the AR group (n = 69) and stable group (n = 131). Their blood samples were collected, and DNA was extracted from the whole blood. High-throughput next-generation sequencing was used to identify single nucleotide polymorphisms (SNPs) of the NFATC2 and NFATC4 genes. The correlation of these SNPs with AR was determined by logistic analysis. RESULTS Seventy-one SNPs of the NFATC2 and NFATC4 genes were identified by the sequencing and Hardy-Weinberg equilibrium analyses. After adjusting for age, gender and immunosuppressive protocols, 27 SNPs were correlated with AR, of which the SNP rs2426295 of the NFATC2 gene showed a significant correlation with AR in the HET model (AA vs. AC: OR = 0.43, 95% CI = 0.19-0.98, P = 0.045), but no significant NFATC4 SNPs were identified. CONCLUSIONS Our study shows that the rs2426295 variant of the NFATC2 gene is significantly associated with the occurrence of AR following kidney transplantation. And patients with AA genotypes in rs2426295 are inclined to suffer from AR pathogenesis.
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Affiliation(s)
- Zijie Wang
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Haiwei Yang
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Shuhui Si
- Research Division of Clinical Pharmacology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Zhijian Han
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Jun Tao
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Hao Chen
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Yuqiu Ge
- School of Public Health, Nanjing Medical University, Nanjing, 211166, People's Republic of China
| | - Miao Guo
- Research Division of Clinical Pharmacology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Ke Wang
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Ruoyun Tan
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China
| | - Ji-Fu Wei
- Research Division of Clinical Pharmacology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China.
| | - Min Gu
- Department of Urology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, 210029, People's Republic of China.
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48
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Hogan PG. Calcium-NFAT transcriptional signalling in T cell activation and T cell exhaustion. Cell Calcium 2017; 63:66-69. [PMID: 28153342 PMCID: PMC5739523 DOI: 10.1016/j.ceca.2017.01.014] [Citation(s) in RCA: 130] [Impact Index Per Article: 16.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2016] [Accepted: 01/26/2017] [Indexed: 02/02/2023]
Abstract
A cornerstone of the adaptive immune response is the T cell receptor (TcR)-calcium-calcineurin signalling pathway leading to T cell activation. The 'nuclear factor of activated T cells' proteins NFAT1, NFAT2, and NFAT4 are transcription factors that promote expression of a panel of genes required for activation. It has become apparent that these same NFAT transcription factors underlie an alternative transcriptional program in T cells that serves to limit the immune response. This duality in NFAT transcriptional functions raises the possibility that NFAT transcriptional complexes could be targeted therapeutically to alter the relative strength of the effector and alternative transcriptional programs, thereby modulating immune responses.
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Affiliation(s)
- Patrick G Hogan
- La Jolla Institute for Allergy & Immunology, 9420 Athena Circle, La Jolla, CA 92037, United States.
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49
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Chow EWL, Clancey SA, Billmyre RB, Averette AF, Granek JA, Mieczkowski P, Cardenas ME, Heitman J. Elucidation of the calcineurin-Crz1 stress response transcriptional network in the human fungal pathogen Cryptococcus neoformans. PLoS Genet 2017; 13:e1006667. [PMID: 28376087 PMCID: PMC5380312 DOI: 10.1371/journal.pgen.1006667] [Citation(s) in RCA: 78] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2016] [Accepted: 03/01/2017] [Indexed: 12/02/2022] Open
Abstract
Calcineurin is a highly conserved Ca2+/calmodulin-dependent serine/threonine-specific protein phosphatase that orchestrates cellular Ca2+ signaling responses. In Cryptococcus neoformans, calcineurin is activated by multiple stresses including high temperature, and is essential for stress adaptation and virulence. The transcription factor Crz1 is a major calcineurin effector in Saccharomyces cerevisiae and other fungi. Calcineurin dephosphorylates Crz1, thereby enabling Crz1 nuclear translocation and transcription of target genes. Here we show that loss of Crz1 confers phenotypes intermediate between wild-type and calcineurin mutants, and demonstrate that deletion of the calcineurin docking domain results in the inability of Crz1 to translocate into the nucleus under thermal stress. RNA-sequencing revealed 102 genes that are regulated in a calcineurin-Crz1-dependent manner at 37°C. The majority of genes were down-regulated in cna1Δ and crz1Δ mutants, indicating these genes are normally activated by the calcineurin-Crz1 pathway at high temperature. About 58% of calcineurin-Crz1 target genes have unknown functions, while genes with known or predicted functions are involved in cell wall remodeling, calcium transport, and pheromone production. We identified three calcineurin-dependent response element motifs within the promoter regions of calcineurin-Crz1 target genes, and show that Crz1 binding to target gene promoters is increased upon thermal stress in a calcineurin-dependent fashion. Additionally, we found a large set of genes independently regulated by calcineurin, and Crz1 regulates 59 genes independently of calcineurin. Given the intermediate crz1Δ mutant phenotype, and our recent evidence for a calcineurin regulatory network impacting mRNA in P-bodies and stress granules independently of Crz1, calcineurin likely acts on factors beyond Crz1 that govern mRNA expression/stability to operate a branched transcriptional/post-transcriptional stress response network necessary for fungal virulence. Taken together, our findings reveal the core calcineurin-Crz1 stress response cascade is maintained from ascomycetes to a pathogenic basidiomycete fungus, but its output in C. neoformans appears to be adapted to promote fungal virulence. The ubquitiously conserved serine/threonine-specific protein phosphatase calcineurin is crucial for virulence of several opportunistic human fungal pathogens including Candida albicans, Aspergillus fumigatus, and Cryptococcus neoformans. We show that Crz1 acts downstream of calcineurin, to 1) govern expression of genes involved in cell wall integrity, and calcium and small molecule transport, and 2) contribute to stress survival and virulence of C. neoformans. Our studies reveal that calcineurin also controls mRNA expression levels of other genes independently of Crz1. We propose that calcineurin operates in a branched signal transduction cascade controlling targets at both the transcriptional and post-transcriptional levels.
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Affiliation(s)
- Eve W. L. Chow
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Shelly A. Clancey
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - R. Blake Billmyre
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Anna Floyd Averette
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Joshua A. Granek
- Department of Biostatistics and Bioinformatics, Duke University Medical Center, Durham, North Carolina, United States of America
- Duke Center for the Genomics of Microbial Systems, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Piotr Mieczkowski
- High-Throughput Sequencing Facility, School of Medicine, University of North Carolina, Chapel Hill, North Carolina, United States of America
| | - Maria E. Cardenas
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
| | - Joseph Heitman
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
- * E-mail:
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Xia Y, Zhao G, Lin J, Li C, Cong L, Jiang N, Xu Q, Wang Q. 1,25(OH)2D3 and VDR Signaling Pathways Regulate the Inhibition of Dectin-1 Caused by Cyclosporine A in Response to Aspergillus Fumigatus in Human Corneal Epithelial Cells. PLoS One 2016; 11:e0164717. [PMID: 27755569 PMCID: PMC5068748 DOI: 10.1371/journal.pone.0164717] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2016] [Accepted: 09/29/2016] [Indexed: 01/24/2023] Open
Abstract
Background The objective of this study is to observe whether cyclosporine A (CsA) inhibits the expression of dectin-1 in human corneal epithelial cells infected with Aspergillus fumigatus (A. fumigatus) and to investigate the molecular mechanisms of the inhibition. Methods Immortalized human corneal epithelial cells (HCECs) were pretreated with 1,25(OH)2D3 and VDR inhibitor for 1 h, and then they were pretreated with CsA for 12h. After these pretreatments, the HCECs were stimulated with A. fumigatus and curdlan respectively, and the expression of dectin-1 and proinflammatory cytokines (IL-1β and TNF-α) were detected by RT-PCR, western blot and ELISA. Results Dectin-1 mRNA and dectin-1 protein expression increased when HCECs were stimulated with A. fumigatus or curdlan, and CsA inhibited the dectin-1 expression both in mRNA and protein levels specifically. Dectin-1 and proinflammatory cytokine expression levels were higher when HCECs were pretreated with VDR inhibitor and CsA compared to pretreatment with CsA alone, while dectin-1 and proinflammatory cytokine levels were lower when HCECs were pretreated with 1,25(OH)2D3 and CsA compared to pretreatment with CsA alone. Conclusions These data provide evidence that CsA can inhibit the expression of dectin-1 and proinflammatory cytokines through dectin-1 when HCECs are stimulated by A. fumigatus or curdlan. The active form of vitamin D, 1,25(OH)2D3, and VDR signaling pathway regulate the inhibition of CsA. The inhibition is enhanced by 1,25(OH)2D3, and the VDR inhibitor suppresses the inhibition.
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MESH Headings
- Antigens, Bacterial/immunology
- Antigens, Bacterial/pharmacology
- Aspergillus fumigatus/metabolism
- Calcitriol/antagonists & inhibitors
- Calcitriol/metabolism
- Cells, Cultured
- Cyclosporine/pharmacology
- Enzyme-Linked Immunosorbent Assay
- Epithelium, Corneal/cytology
- Epithelium, Corneal/drug effects
- Epithelium, Corneal/metabolism
- Gene Expression/drug effects
- Humans
- Interleukin-1beta/analysis
- Interleukin-1beta/genetics
- Interleukin-1beta/metabolism
- Lectins, C-Type/antagonists & inhibitors
- Lectins, C-Type/genetics
- Lectins, C-Type/metabolism
- RNA, Messenger/metabolism
- Receptors, Calcitriol/antagonists & inhibitors
- Receptors, Calcitriol/metabolism
- Signal Transduction/drug effects
- Tumor Necrosis Factor-alpha/analysis
- Tumor Necrosis Factor-alpha/genetics
- Tumor Necrosis Factor-alpha/metabolism
- beta-Glucans/pharmacology
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Affiliation(s)
- Yiping Xia
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
| | - Guiqiu Zhao
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
- * E-mail:
| | - Jing Lin
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
| | - Cui Li
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
| | - Lin Cong
- Shandong Eye Institute, Qingdao, Shandong, China
| | - Nan Jiang
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
| | - Qiang Xu
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
| | - Qian Wang
- Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China
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