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Deshmukh A, Chang K, Cuala J, Campos MJH, Mahmood S, Verma R, Georgia S, Loconte V, White KL. Secretory stimuli distinctly regulate insulin secretory granule maturation through structural remodeling. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2025:2025.03.29.644646. [PMID: 40235991 PMCID: PMC11996419 DOI: 10.1101/2025.03.29.644646] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 04/17/2025]
Abstract
Insulin secretory granule (ISG) maturation is a crucial aspect of insulin secretion and glucose homeostasis. The regulation of this maturation remains poorly understood, especially how secretory stimuli affect ISG maturity and subcellular localization. In this study, we used soft X-tomography (SXT) to quantitatively map ISG morphology, density, and location in single INS-1E and mouse pancreatic β-cells under the effect of various secretory stimuli. We found that the activation of glucokinase (GK), gastric inhibitory polypeptide receptor (GIPR), glucagon-like peptide-1 receptor (GLP-1R), and G-protein coupled receptor 40 (GPR40) promote ISG maturation. Each stimulus induces unique structural remodeling in ISGs, by altering size and density, depending on the specific signaling cascades activated. These distinct ISG subpopulations mobilize and redistribute in the cell altering overall cellular structural organization. Our results provide insight into how current diabetes and obesity therapies impact ISG maturation and may inform the development of future treatments that target maturation specifically.
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Jiao H, Li X, Li Y, Guo Z, Yang Y, Luo Y, Hu X, Yu L. Packaged release and targeted delivery of cytokines by migrasomes in circulation. Cell Discov 2024; 10:121. [PMID: 39648224 PMCID: PMC11625823 DOI: 10.1038/s41421-024-00749-x] [Citation(s) in RCA: 4] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2024] [Accepted: 10/31/2024] [Indexed: 12/10/2024] Open
Abstract
In dynamic systems like the circulatory system, establishing localized cytokine gradients is challenging. Upon lipopolysaccharide (LPS) stimulation, we observed that monocytes release numerous migrasomes enriched with inflammatory cytokines, such as TNF-α and IL-6. These cytokines are transported into migrasomes via secretory carriers, leading to their immediate exocytosis or eventual release from detached migrasomes. We successfully isolated TNF-α and IL-6-enriched, monocyte-derived migrasomes from the blood of LPS-treated mice. Total secretion analysis revealed a substantial amount of TNF-α and IL-6 released in a migrasome-packaged form. Thus, detached, monocyte-derived migrasomes represent a type of extracellular vesicle highly enriched with cytokines. Physiologically, these cytokine-laden migrasomes rapidly accumulate at local sites of inflammation, effectively creating a concentrated source of cytokines. Our research uncovers novel mechanisms for cytokine release and delivery, providing new insights into immune response modulation.
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Affiliation(s)
- Haifeng Jiao
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Xiaopeng Li
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Ying Li
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Ziyi Guo
- Institute for Immunology, Tsinghua University-Peking University Joint Centre for Life Sciences, School of Medicine, Tsinghua University, Beijing, China
| | - Yuzhuo Yang
- Institute for Immunology, Tsinghua University-Peking University Joint Centre for Life Sciences, School of Medicine, Tsinghua University, Beijing, China
| | - Yiqun Luo
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Xiaoyu Hu
- Institute for Immunology, Tsinghua University-Peking University Joint Centre for Life Sciences, School of Medicine, Tsinghua University, Beijing, China
| | - Li Yu
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China.
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Siehler J, Bilekova S, Chapouton P, Dema A, Albanese P, Tamara S, Jain C, Sterr M, Enos SJ, Chen C, Malhotra C, Villalba A, Schomann L, Bhattacharya S, Feng J, Akgün Canan M, Ribaudo F, Ansarullah, Burtscher I, Ahlbrecht C, Plettenburg O, Kurth T, Scharfmann R, Speier S, Scheltema RA, Lickert H. Inceptor binds to and directs insulin towards lysosomal degradation in β cells. Nat Metab 2024; 6:2374-2390. [PMID: 39587340 DOI: 10.1038/s42255-024-01164-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/01/2024] [Accepted: 10/07/2024] [Indexed: 11/27/2024]
Abstract
Blunted first-phase insulin secretion and insulin deficiency are indicators of β cell dysfunction and diabetes manifestation. Therefore, insights into molecular mechanisms that regulate insulin homeostasis might provide entry sites to replenish insulin content and restore β cell function. Here, we identify the insulin inhibitory receptor (inceptor; encoded by the gene IIR/ELAPOR1) as an insulin-binding receptor that regulates insulin stores by lysosomal degradation. Using human induced pluripotent stem cell (SC)-derived islets, we show that IIR knockout (KO) results in enhanced SC β cell differentiation and survival. Strikingly, extended in vitro culture of IIR KO SC β cells leads to greatly increased insulin content and glucose-stimulated insulin secretion (GSIS). We find that inceptor localizes to clathrin-coated vesicles close to the plasma membrane and in the trans-Golgi network as well as in secretory granules, where it acts as a sorting receptor to direct proinsulin and insulin towards lysosomal degradation. Targeting inceptor using a monoclonal antibody increases proinsulin and insulin content and improves SC β cell GSIS. Altogether, our findings reveal the basic mechanisms of β cell insulin turnover and identify inceptor as an insulin degradation receptor.
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Affiliation(s)
- Johanna Siehler
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Sara Bilekova
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Prisca Chapouton
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Alessandro Dema
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Pascal Albanese
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research, Utrecht University, Utrecht, The Netherlands
- Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, Netherlands
- Proteomics Centre, Utrecht, The Netherlands
- Université Grenoble Alpes, IRIG, CEA, CNRS, INRAE, UMR 5168, 38000, Grenoble, France
- Université Grenoble Alpes, INSERM, IRIG, CEA, UA13 BGE, CNRS, CEA, FR2048 ProFI, 38000, Grenoble, France
| | - Sem Tamara
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research, Utrecht University, Utrecht, The Netherlands
- Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, Netherlands
- Proteomics Centre, Utrecht, The Netherlands
| | - Chirag Jain
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Michael Sterr
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Stephen J Enos
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- Institute of Physiology, Faculty of Medicine Carl Gustav Carus, Dresden University of Technology, Dresden, Germany
- Paul Langerhans Institute Dresden (PLID) of the Helmholtz Zentrum München at the University Clinic Carl Gustav Carus of Dresden University of Technology, Dresden, Germany
| | - Chunguang Chen
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- Institute of Physiology, Faculty of Medicine Carl Gustav Carus, Dresden University of Technology, Dresden, Germany
- Paul Langerhans Institute Dresden (PLID) of the Helmholtz Zentrum München at the University Clinic Carl Gustav Carus of Dresden University of Technology, Dresden, Germany
| | - Chetna Malhotra
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- TUM School of Life Sciences, Technische Universität München, Freising, Germany
| | - Adrian Villalba
- Université Paris Cité, Institut Cochin, CNRS, INSERM, Paris, France
| | - Leopold Schomann
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Sreya Bhattacharya
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Jin Feng
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Melis Akgün Canan
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- School of Medicine and Health, Technische Universität München, Munich, Germany
| | - Federico Ribaudo
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Ansarullah
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Ingo Burtscher
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany
- German Center for Diabetes Research (DZD), Neuherberg, Germany
| | - Christin Ahlbrecht
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- Institute for Medicinal Chemistry, Molecular Targets and Therapeutics Center, Helmholtz Zentrum München, Neuherberg, Germany
- Institute of Organic Chemistry (BMWZ) and Laboratory for Nano- and Quantum Engineering (LNQE), Center of Biomolecular Research, Leibniz, University Hannover, Hannover, Germany
| | - Oliver Plettenburg
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- Institute for Medicinal Chemistry, Molecular Targets and Therapeutics Center, Helmholtz Zentrum München, Neuherberg, Germany
- Institute of Organic Chemistry (BMWZ) and Laboratory for Nano- and Quantum Engineering (LNQE), Center of Biomolecular Research, Leibniz, University Hannover, Hannover, Germany
- Institute of Lung Health (ILH), Giessen, Germany
| | - Thomas Kurth
- Center for Molecular and Cellular Bioengineering (CMCB), Technology Platform, Core Facility Electron Microscopy and Histology, Dresden University of Technology, Dresden, Germany
| | | | - Stephan Speier
- German Center for Diabetes Research (DZD), Neuherberg, Germany
- Institute of Physiology, Faculty of Medicine Carl Gustav Carus, Dresden University of Technology, Dresden, Germany
- Paul Langerhans Institute Dresden (PLID) of the Helmholtz Zentrum München at the University Clinic Carl Gustav Carus of Dresden University of Technology, Dresden, Germany
| | - Richard A Scheltema
- Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research, Utrecht University, Utrecht, The Netherlands
- Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, Netherlands
- Proteomics Centre, Utrecht, The Netherlands
- Center for Proteome, Research and Department of Biochemistry, University of Liverpool, Liverpool, UK
| | - Heiko Lickert
- Institute of Diabetes and Regeneration Research, Helmholtz Diabetes Center, Helmholtz Zentrum München, Neuherberg, Germany.
- German Center for Diabetes Research (DZD), Neuherberg, Germany.
- School of Medicine and Health, Technische Universität München, Munich, Germany.
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Gukovskaya AS, Lerch MM, Mayerle J, Sendler M, Ji B, Saluja AK, Gorelick FS, Gukovsky I. Trypsin in pancreatitis: The culprit, a mediator, or epiphenomenon? World J Gastroenterol 2024; 30:4417-4438. [PMID: 39534420 PMCID: PMC11551668 DOI: 10.3748/wjg.v30.i41.4417] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/23/2024] [Revised: 06/19/2024] [Accepted: 07/16/2024] [Indexed: 10/23/2024] Open
Abstract
Pancreatitis is a common, life-threatening inflammatory disease of the exocrine pancreas. Its pathogenesis remains obscure, and no specific or effective treatment is available. Gallstones and alcohol excess are major etiologies of pancreatitis; in a small portion of patients the disease is hereditary. Pancreatitis is believed to be initiated by injured acinar cells (the main exocrine pancreas cell type), leading to parenchymal necrosis and local and systemic inflammation. The primary function of these cells is to produce, store, and secrete a variety of enzymes that break down all categories of nutrients. Most digestive enzymes, including all proteases, are secreted by acinar cells as inactive proforms (zymogens) and in physiological conditions are only activated when reaching the intestine. The generation of trypsin from inactive trypsinogen in the intestine plays a critical role in physiological activation of other zymogens. It was proposed that pancreatitis results from proteolytic autodigestion of the gland, mediated by premature/inappropriate trypsinogen activation within acinar cells. The intra-acinar trypsinogen activation is observed in experimental models of acute and chronic pancreatitis, and in human disease. On the basis of these observations, it has been considered the central pathogenic mechanism of pancreatitis - a concept with a century-old history. This review summarizes the data on trypsinogen activation in experimental and genetic rodent models of pancreatitis, particularly the more recent genetically engineered mouse models that mimic mutations associated with hereditary pancreatitis; analyzes the mechanisms mediating trypsinogen activation and protecting the pancreas against its' damaging effects; discusses the gaps in our knowledge, potential therapeutic approaches, and directions for future research. We conclude that trypsin is not the culprit in the disease pathogenesis but, at most, a mediator of some pancreatitis responses. Therefore, the search for effective therapies should focus on approaches to prevent or normalize other intra-acinar pathologic processes, such as defective autophagy leading to parenchymal cell death and unrelenting inflammation.
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Affiliation(s)
- Anna S Gukovskaya
- Department of Medicine, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90073, United States
- Department of Medicine, VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073, United States
| | - Markus M Lerch
- Department of Medicine, Ludwig Maximilian University Hospital, Munich 81377, Germany
| | - Julia Mayerle
- Department of Medicine II, Ludwig Maximilian University of Munich, Munich 81377, Germany
| | - Matthias Sendler
- Department of Medicine A, University of Greifswald, Greifswald 17475, Germany
| | - Baoan Ji
- Department of Cancer Biology, Mayo Clinic, Jacksonville, FL 32224, United States
| | - Ashok K Saluja
- Department of Surgery, University of Miami Miller School of Medicine, Miami, FL 33136, United States
| | - Fred S Gorelick
- Departments of Cell Biology and Internal Medicine, Yale University School of Medicine and VA West Haven, New Haven, CT 06519, United States
| | - Ilya Gukovsky
- Department of Medicine, David Geffen School of Medicine, University of California at Los Angeles, Los Angeles, CA 90073, United States
- Department of Medicine, VA Greater Los Angeles Healthcare System, Los Angeles, CA 90073, United States
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5
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Jiao H, Li X, Li Y, Guo Y, Hu X, Sho T, Luo Y, Wang J, Cao H, Du W, Li D, Yu L. Localized, highly efficient secretion of signaling proteins by migrasomes. Cell Res 2024; 34:572-585. [PMID: 38918584 PMCID: PMC11291916 DOI: 10.1038/s41422-024-00992-7] [Citation(s) in RCA: 11] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2024] [Accepted: 06/11/2024] [Indexed: 06/27/2024] Open
Abstract
Migrasomes, enriched with signaling molecules such as chemokines, cytokines and angiogenic factors, play a pivotal role in the spatially defined delivery of these molecules, influencing critical physiological processes including organ morphogenesis and angiogenesis. The mechanism governing the accumulation of signaling molecules in migrasomes has been elusive. In this study, we show that secretory proteins, including signaling proteins, are transported into migrasomes by secretory carriers via both the constitutive and regulated secretion pathways. During cell migration, a substantial portion of these carriers is redirected to the rear of the cell and actively transported into migrasomes, driven by the actin-dependent motor protein Myosin-5a. Once at the migrasomes, these carriers fuse with the migrasome membrane through SNARE-mediated mechanisms. Inhibiting migrasome formation significantly reduces secretion, suggesting migrasomes as a principal secretion route in migrating cells. Our findings reveal a specialized, highly localized secretion paradigm in migrating cells, conceptually paralleling the targeted neurotransmitter release observed in neuronal systems.
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Affiliation(s)
- Haifeng Jiao
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China.
| | - Xiaopeng Li
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Ying Li
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Yuting Guo
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Xiaoyu Hu
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Takami Sho
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Yiqun Luo
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Jinyu Wang
- SLSTU-Nikon Biological Imaging Center, Center of Biomedical Analysis, Tsinghua University, Beijing, China
| | - Huizhen Cao
- SLSTU-Nikon Biological Imaging Center, Center of Biomedical Analysis, Tsinghua University, Beijing, China
| | - Wanqing Du
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
| | - Dong Li
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Li Yu
- State Key Laboratory of Membrane Biology, Tsinghua University-Peking University Joint Centre for Life Sciences, Beijing Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China.
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Isola M, Maxia C, Murtas D, Ekström J, Isola R, Loy F. Prostate-specific antigen: An unfamiliar protein in the human salivary glands. J Anat 2024; 244:873-881. [PMID: 38111134 PMCID: PMC11021670 DOI: 10.1111/joa.13996] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/09/2023] [Revised: 11/09/2023] [Accepted: 12/04/2023] [Indexed: 12/20/2023] Open
Abstract
OBJECTIVES The presence of prostate-specific antigen (PSA) in saliva and salivary glands has been reported. Nevertheless, its release pathway in these glands remains to be elucidated. Here, we showed PSA subcellular distribution focusing on its plausible route in human salivary parenchyma. MATERIALS AND METHODS Sections of parotid and submandibular glands were subjected to the immunohistochemical demonstration of PSA by the streptavidin-biotin method revealed by alkaline phosphatase. Moreover, ultrathin sections were collected on nickel grids and processed for immunocytochemical analysis, to visualize the intracellular distribution pattern of PSA through the observation by transmission electron microscopy. RESULTS By immunohistochemistry, in both parotid and submandibular glands PSA expression was detected in serous secretory acini and striated ducts. By immunocytochemistry, immunoreactivity was retrieved in the cytoplasmic compartment of acinar and ductal cells, often associated with small cytoplasmic vesicles. PSA labeling appeared also on rough endoplasmic reticulum and in the acini's lumen. A negligible PSA labeling appeared in most of the secretory granules of both glands. CONCLUSIONS Our findings clearly support that human parotid and submandibular glands are involved in PSA secretion. Moreover, based on the immunoreactivity pattern, its release in oral cavity would probably occur by minor regulated secretory or constitutive-like secretory pathways.
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Affiliation(s)
- Michela Isola
- Department of Biomedical SciencesUniversity of CagliariCagliariItaly
| | - Cristina Maxia
- Department of Biomedical SciencesUniversity of CagliariCagliariItaly
| | - Daniela Murtas
- Department of Biomedical SciencesUniversity of CagliariCagliariItaly
| | - Jörgen Ekström
- Division of Pharmacology, Institute of Neuroscience and PhysiologySahlgrenska Academy at the University of GothenburgGöteborgSweden
| | - Raffaella Isola
- Department of Biomedical SciencesUniversity of CagliariCagliariItaly
| | - Francesco Loy
- Department of Biomedical SciencesUniversity of CagliariCagliariItaly
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Campelo F, Tian M, von Blume J. Rediscovering the intricacies of secretory granule biogenesis. Curr Opin Cell Biol 2023; 85:102231. [PMID: 37657367 DOI: 10.1016/j.ceb.2023.102231] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2023] [Revised: 08/01/2023] [Accepted: 08/07/2023] [Indexed: 09/03/2023]
Abstract
Regulated secretion, an essential cellular process, relies on secretory granules (SGs) for the controlled release of a diverse range of cargo molecules, including proteins, peptides, hormones, enzymes, and neurotransmitters. SG biogenesis encompasses cargo selection, sorting, packaging, and trafficking, with the trans-Golgi Network (TGN) playing a central role. Research in the last three decades has revealed significant components required for SG biogenesis; however, no cargo receptor transferring granule cargo from the TGN to immature SGs (ISGs) has yet been identified. Consequently, recent research has devoted significant attention to studying receptor-independent cargo sorting mechanisms, shedding new light on the complexities of regulated secretion. Understanding the underlying molecular and biophysical mechanisms behind cargo sorting into ISGs holds great promise for advancing our knowledge of cellular communication and disease mechanisms.
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Affiliation(s)
- Felix Campelo
- ICFO-Institut de Ciencies Fotoniques, The Barcelona Institute of Science and Technology, 08860, Castelldefels, Barcelona, Spain
| | - Meng Tian
- Department of Cell Biology, Yale University School of Medicine, New Haven, CT, USA
| | - Julia von Blume
- Department of Cell Biology, Yale University School of Medicine, New Haven, CT, USA.
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Loh YP, Xiao L, Park JJ. Trafficking of hormones and trophic factors to secretory and extracellular vesicles: a historical perspective and new hypothesis. EXTRACELLULAR VESICLES AND CIRCULATING NUCLEIC ACIDS 2023; 4:568-587. [PMID: 38435713 PMCID: PMC10906782 DOI: 10.20517/evcna.2023.34] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Indexed: 03/05/2024]
Abstract
It is well known that peptide hormones and neurotrophic factors are intercellular messengers that are packaged into secretory vesicles in endocrine cells and neurons and released by exocytosis upon the stimulation of the cells in a calcium-dependent manner. These secreted molecules bind to membrane receptors, which then activate signal transduction pathways to mediate various endocrine/trophic functions. Recently, there is evidence that these molecules are also in extracellular vesicles, including small extracellular vesicles (sEVs), which appear to be taken up by recipient cells. This finding raised the hypothesis that they may have functions differentiated from their classical secretory hormone/neurotrophic factor actions. In this article, the historical perspective and updated mechanisms for the sorting and packaging of hormones and neurotrophic factors into secretory vesicles and their transport in these organelles for release at the plasma membrane are reviewed. In contrast, little is known about the packaging of hormones and neurotrophic factors into extracellular vesicles. One proposal is that these molecules could be sorted at the trans-Golgi network, which then buds to form Golgi-derived vesicles that can fuse to endosomes and subsequently form intraluminal vesicles. They are then taken up by multivesicular bodies to form extracellular vesicles, which are subsequently released. Other possible mechanisms for packaging RSP proteins into sEVs are discussed. We highlight some studies in the literature that suggest the dual vesicular pathways for the release of hormones and neurotrophic factors from the cell may have some physiological significance in intercellular communication.
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Affiliation(s)
- Y. Peng Loh
- Section on Cellular Neurobiology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA
| | - Lan Xiao
- Section on Cellular Neurobiology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA
| | - Joshua J. Park
- Scientific Review Branch, National Institute on Aging, National Institutes of Health, Bethesda, MD 20892, USA
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9
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(Flintoaca) Alexandru PR, Chiritoiu GN, Lixandru D, Zurac S, Ionescu-Targoviste C, Petrescu SM. EDEM1 regulates the insulin mRNA level by inhibiting the endoplasmic reticulum stress-induced IRE1/JNK/c-Jun pathway. iScience 2023; 26:107956. [PMID: 37822496 PMCID: PMC10562789 DOI: 10.1016/j.isci.2023.107956] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2023] [Revised: 07/22/2023] [Accepted: 09/14/2023] [Indexed: 10/13/2023] Open
Abstract
Pancreatic beta cells produce and secrete insulin as a response to rises in blood glucose. Despite the advances in understanding glucose-regulated insulin transcription and translation the mechanisms triggering the synthesis of new insulin molecules are still incompletely described. In this report, we identify EDEM1 as a new modulator of insulin synthesis and secretion. In the presence of EDEM1, INS-1E cells secrete significantly more insulin upon glucose stimulation compared to control cells. We found that overexpression of EDEM1 inhibited the IRE1/JNK/c-Jun pathway, leading to an increase in the insulin mRNA level. Similarly, EDEM1 transduced human islets secreted significantly more insulin upon stimulation. Furthermore, EDEM1 improved insulin secretion restoring normoglycemia and glucose tolerance in diabetic rats. We propose EDEM1 as a regulator of the UPR via IRE1/XBP1s and IRE1/JNK/c-Jun signaling cascades and insulin transcription in pancreatic β-cells, supporting EDEM1 as a potential target for the treatment of diabetes.
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Affiliation(s)
| | - Gabriela N. Chiritoiu
- Department of Molecular Cell Biology, Institute of Biochemistry, Romanian Academy, 060031 Bucharest, Romania
| | - Daniela Lixandru
- Department of Biochemistry, “Carol Davila” University of Medicine and Pharmacy, 050474 Bucharest, Romania
| | - Sabina Zurac
- Department of Physiology, “Carol Davila” University of Medicine and Pharmacy, 050474 Bucharest, Romania
| | | | - Stefana M. Petrescu
- Department of Molecular Cell Biology, Institute of Biochemistry, Romanian Academy, 060031 Bucharest, Romania
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10
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Iyer DR, Venkatraman J, Tanguy E, Vitale N, Mahapatra NR. Chromogranin A and its derived peptides: potential regulators of cholesterol homeostasis. Cell Mol Life Sci 2023; 80:271. [PMID: 37642733 PMCID: PMC11072126 DOI: 10.1007/s00018-023-04908-3] [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: 03/03/2023] [Revised: 08/02/2023] [Accepted: 08/03/2023] [Indexed: 08/31/2023]
Abstract
Chromogranin A (CHGA), a member of the granin family of proteins, has been an attractive therapeutic target and candidate biomarker for several cardiovascular, neurological, and inflammatory disorders. The prominence of CHGA stems from the pleiotropic roles of several bioactive peptides (e.g., catestatin, pancreastatin, vasostatins) generated by its proteolytic cleavage and by their wide anatomical distribution. These peptides are emerging as novel modulators of cardiometabolic diseases that are often linked to high blood cholesterol levels. However, their impact on cholesterol homeostasis is poorly understood. The dynamic nature of cholesterol and its multitudinous roles in almost every aspect of normal body function makes it an integral component of metabolic physiology. A tightly regulated coordination of cholesterol homeostasis is imperative for proper functioning of cellular and metabolic processes. The deregulation of cholesterol levels can result in several pathophysiological states. Although studies till date suggest regulatory roles for CHGA and its derived peptides on cholesterol levels, the mechanisms by which this is achieved still remain unclear. This review aims to aggregate and consolidate the available evidence linking CHGA with cholesterol homeostasis in health and disease. In addition, we also look at common molecular regulatory factors (viz., transcription factors and microRNAs) which could govern the expression of CHGA and genes involved in cholesterol homeostasis under basal and pathological conditions. In order to gain further insights into the pathways mediating cholesterol regulation by CHGA/its derived peptides, a few prospective signaling pathways are explored, which could act as primers for future studies.
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Affiliation(s)
- Dhanya R Iyer
- Department of Biotechnology, Bhupat and Jyoti Mehta School of Biosciences, Indian Institute of Technology Madras, Chennai, 600036, India
| | - Janani Venkatraman
- Department of Biotechnology, Bhupat and Jyoti Mehta School of Biosciences, Indian Institute of Technology Madras, Chennai, 600036, India
| | - Emeline Tanguy
- Institut des Neurosciences Cellulaires et Intégratives, CNRS UPR 3212 and Université de Strasbourg, 5 Rue Blaise Pascal, 67000, Strasbourg, France
| | - Nicolas Vitale
- Institut des Neurosciences Cellulaires et Intégratives, CNRS UPR 3212 and Université de Strasbourg, 5 Rue Blaise Pascal, 67000, Strasbourg, France.
| | - Nitish R Mahapatra
- Department of Biotechnology, Bhupat and Jyoti Mehta School of Biosciences, Indian Institute of Technology Madras, Chennai, 600036, India.
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11
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Horváth D, Dürvanger Z, K Menyhárd D, Sulyok-Eiler M, Bencs F, Gyulai G, Horváth P, Taricska N, Perczel A. Polymorphic amyloid nanostructures of hormone peptides involved in glucose homeostasis display reversible amyloid formation. Nat Commun 2023; 14:4621. [PMID: 37528104 PMCID: PMC10394066 DOI: 10.1038/s41467-023-40294-x] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Accepted: 07/21/2023] [Indexed: 08/03/2023] Open
Abstract
A large group of hormones are stored as amyloid fibrils in acidic secretion vesicles before they are released into the bloodstream and readopt their functional state. Here, we identify an evolutionarily conserved hexapeptide sequence as the major aggregation-prone region (APR) of gastrointestinal peptides of the glucagon family: xFxxWL. We determine nine polymorphic crystal structures of the APR segments of glucagon-like peptides 1 and 2, and exendin and its derivatives. We follow amyloid formation by CD, FTIR, ThT assays, and AFM. We propose that the pH-dependent changes of the protonation states of glutamate/aspartate residues of APRs initiate switching between the amyloid and the folded, monomeric forms of the hormones. We find that pH sensitivity diminishes in the absence of acidic gatekeepers and amyloid formation progresses over a broad pH range. Our results highlight the dual role of short aggregation core motifs in reversible amyloid formation and receptor binding.
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Affiliation(s)
- Dániel Horváth
- ELKH-ELTE Protein Modeling Research Group ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Zsolt Dürvanger
- ELKH-ELTE Protein Modeling Research Group ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
- Laboratory of Structural Chemistry and Biology ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Dóra K Menyhárd
- ELKH-ELTE Protein Modeling Research Group ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
- Laboratory of Structural Chemistry and Biology ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Máté Sulyok-Eiler
- Laboratory of Structural Chemistry and Biology ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
- Hevesy György PhD School of Chemistry, ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Fruzsina Bencs
- Laboratory of Structural Chemistry and Biology ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
- Hevesy György PhD School of Chemistry, ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Gergő Gyulai
- Laboratory of Interfaces and Nanostructures, Institute of Chemistry, Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - Péter Horváth
- Department of Pharmaceutical Chemistry, Semmelweis University, Hőgyes Endre utca 9, Budapest, 1092, Hungary
| | - Nóra Taricska
- ELKH-ELTE Protein Modeling Research Group ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary
| | - András Perczel
- ELKH-ELTE Protein Modeling Research Group ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary.
- Laboratory of Structural Chemistry and Biology ELTE Eötvös Loránd University, Pázmány Péter sétány 1/A, Budapest, H-1117, Hungary.
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12
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Boyer CK, Bauchle CJ, Zhang J, Wang Y, Stephens SB. Synchronized proinsulin trafficking reveals delayed Golgi export accompanies β-cell secretory dysfunction in rodent models of hyperglycemia. Sci Rep 2023; 13:5218. [PMID: 36997560 PMCID: PMC10063606 DOI: 10.1038/s41598-023-32322-z] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2022] [Accepted: 03/25/2023] [Indexed: 04/03/2023] Open
Abstract
The pancreatic islet β-cell's preference for release of newly synthesized insulin requires careful coordination of insulin exocytosis with sufficient insulin granule production to ensure that insulin stores exceed peripheral demands for glucose homeostasis. Thus, the cellular mechanisms regulating insulin granule production are critical to maintaining β-cell function. In this report, we utilized the synchronous protein trafficking system, RUSH, in primary β-cells to evaluate proinsulin transit through the secretory pathway leading to insulin granule formation. We demonstrate that the trafficking, processing, and secretion of the proinsulin RUSH reporter, proCpepRUSH, are consistent with current models of insulin maturation and release. Using both a rodent dietary and genetic model of hyperglycemia and β-cell dysfunction, we show that proinsulin trafficking is impeded at the Golgi and coincides with the decreased appearance of nascent insulin granules at the plasma membrane. Ultrastructural analysis of β-cells from diabetic leptin receptor deficient mice revealed gross morphological changes in Golgi structure, including shortened and swollen cisternae, and partial Golgi vesiculation, which are consistent with defects in secretory protein export. Collectively, this work highlights the utility of the proCpepRUSH reporter in studying proinsulin trafficking dynamics and suggests that altered Golgi export function contributes to β-cell secretory defects in the pathogenesis of Type 2 diabetes.
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Affiliation(s)
- Cierra K Boyer
- Department of Neuroscience and Pharmacology, University of Iowa, Iowa City, IA, USA
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, 52246, USA
| | - Casey J Bauchle
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, 52246, USA
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Iowa, Iowa City, IA, 52246, USA
| | - Jianchao Zhang
- Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, MI, USA
| | - Yanzhuang Wang
- Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, MI, USA
- Department of Neurology, School of Medicine, University of Michigan, Ann Arbor, MI, USA
| | - Samuel B Stephens
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA, 52246, USA.
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Iowa, Iowa City, IA, 52246, USA.
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13
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Liu Y, Liu J, He X. Different p Ka Shifts of Internal GLU8 in Human β-Endorphin Amyloid Revealing a Coupling of Internal Ionization and Stepwise Fibril Disassembly. J Phys Chem B 2023; 127:1089-1096. [PMID: 36696655 DOI: 10.1021/acs.jpcb.2c06706] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/27/2023]
Abstract
As a functional amyloid, human β-endorphin amyloid fibril features a β-solenoid conformation and store peptide hormones within acidic secretory granules, which would be released into the blood through fibril disassembly when the cellular milieu pH increases from acidic to neutral level on exocytosis. To gain detailed atomic mechanism of β-endorphin amyloid fibrils' pH-responsive disassembly, we conduct constant pH molecular dynamics simulations to investigate the structural and dynamical properties of β-endorphin amyloid fibrils in experiencing the environmental pH changes. Our results demonstrate a clear pKa shift of the internal ionizable residue of GLU8, and this shift becomes even more pronounced when it is buried more deeply in the amyloid fibrils. The unusual pKa of GLU8 reveals that its protonation state changes from the protonated state in the acidic secretory granule to the deprotonated state in the neutral pH conditions in the blood, where the deprotonation of GLU8 leads to unfavorable interactions within the hydrophobic core of the amyloid and subsequent fibril disassembly. The different pKa shifts of GLU8 relative to its positions in the amyloid fibril indicate that the β-endorphin amyloid fibril disassembly is a stepwise process, accounting for the experimental observation that the disassembly always initiates from the outermost layer. This study reveals the critical role of the protonation state of GLU8 in amyloid fibrils' pH-responsive disassembly, and provides clear insights for understanding the structural transitions of amyloids in hormone secretion. This study also provides theoretical basis for designing pH-sensitive biological tools for specific use with precise positioning of ionizable residues into the hydrophobic interior of proteins.
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Affiliation(s)
- Yiwei Liu
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, Shanghai Frontiers Science Center of Molecule Intelligent Syntheses, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai, 200062, China
| | - Jinfeng Liu
- Department of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, China
| | - Xiao He
- Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development, Shanghai Frontiers Science Center of Molecule Intelligent Syntheses, School of Chemistry and Molecular Engineering, East China Normal University, Shanghai, 200062, China.,New York University-East China Normal University Center for Computational Chemistry, New York University Shanghai, Shanghai, 200062, China
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14
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Galli A, Arunagiri A, Dule N, Castagna M, Marciani P, Perego C. Cholesterol Redistribution in Pancreatic β-Cells: A Flexible Path to Regulate Insulin Secretion. Biomolecules 2023; 13:224. [PMID: 36830593 PMCID: PMC9953638 DOI: 10.3390/biom13020224] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2022] [Revised: 01/17/2023] [Accepted: 01/21/2023] [Indexed: 01/26/2023] Open
Abstract
Pancreatic β-cells, by secreting insulin, play a key role in the control of glucose homeostasis, and their dysfunction is the basis of diabetes development. The metabolic milieu created by high blood glucose and lipids is known to play a role in this process. In the last decades, cholesterol has attracted significant attention, not only because it critically controls β-cell function but also because it is the target of lipid-lowering therapies proposed for preventing the cardiovascular complications in diabetes. Despite the remarkable progress, understanding the molecular mechanisms responsible for cholesterol-mediated β-cell function remains an open and attractive area of investigation. Studies indicate that β-cells not only regulate the total cholesterol level but also its redistribution within organelles, a process mediated by vesicular and non-vesicular transport. The aim of this review is to summarize the most current view of how cholesterol homeostasis is maintained in pancreatic β-cells and to provide new insights on the mechanisms by which cholesterol is dynamically distributed among organelles to preserve their functionality. While cholesterol may affect virtually any activity of the β-cell, the intent of this review is to focus on early steps of insulin synthesis and secretion, an area still largely unexplored.
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Affiliation(s)
- Alessandra Galli
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), Università degli Studi di Milano, 20134 Milan, Italy
| | - Anoop Arunagiri
- Division of Metabolism, Endocrinology & Diabetes, Department of Internal Medicine, University of Michigan, Ann Arbor, MA 48106, USA
| | - Nevia Dule
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), Università degli Studi di Milano, 20134 Milan, Italy
| | - Michela Castagna
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), Università degli Studi di Milano, 20134 Milan, Italy
| | - Paola Marciani
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), Università degli Studi di Milano, 20134 Milan, Italy
| | - Carla Perego
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), Università degli Studi di Milano, 20134 Milan, Italy
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15
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Li M, Feng F, Feng H, Hu P, Xue Y, Xu T, Song E. VAMP4 regulates insulin levels by targeting secretory granules to lysosomes. J Cell Biol 2022; 221:213439. [PMID: 36053215 PMCID: PMC9441717 DOI: 10.1083/jcb.202110164] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2021] [Revised: 06/28/2022] [Accepted: 07/25/2022] [Indexed: 11/22/2022] Open
Abstract
Insulin levels are essential for the maintenance of glucose homeostasis, and deviations lead to pathoglycemia or diabetes. However, the metabolic mechanism controlling insulin quantity and quality is poorly understood. In pancreatic β cells, insulin homeostasis and release are tightly governed by insulin secretory granule (ISG) trafficking, but the required regulators and mechanisms are largely unknown. Here, we identified that VAMP4 controlled the insulin levels in response to glucose challenge. VAMP4 deficiency led to increased blood insulin levels and hyperresponsiveness to glucose. In β cells, VAMP4 is packaged into immature ISGs (iISGs) at trans-Golgi networks and subsequently resorted to clathrin-coated vesicles during granule maturation. VAMP4-positive iISGs and resorted vesicles then fuse with lysosomes facilitated by a SNARE complex consisting of VAMP4, STX7, STX8, and VTI1B, which ensures the breakdown of excess (pro)insulin and obsolete materials and thus maintenance of intracellular insulin homeostasis. Thus, VAMP4 is a key factor regulating the insulin levels and a potential target for the treatment of diabetes.
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Affiliation(s)
- Min Li
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China,Guangzhou Laboratory, Guangzhou, China,Bioland Laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory), Guangzhou, China,Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, China
| | - Fengping Feng
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Han Feng
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Pengkai Hu
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China,College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
| | - Yanhong Xue
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Tao Xu
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China,Guangzhou Laboratory, Guangzhou, China,Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, China,College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China,Dr. Tao Xu:
| | - Eli Song
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China,College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China,Correspondence to Dr. Eli Song:
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16
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Xu H, Chang F, Jain S, Heller BA, Han X, Liu Y, Edwards RH. SNX5 targets a monoamine transporter to the TGN for assembly into dense core vesicles by AP-3. J Cell Biol 2022; 221:e202106083. [PMID: 35426896 PMCID: PMC9016777 DOI: 10.1083/jcb.202106083] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2021] [Revised: 12/06/2021] [Accepted: 02/16/2022] [Indexed: 11/22/2022] Open
Abstract
The time course of signaling by peptide hormones, neural peptides, and other neuromodulators depends on their storage inside dense core vesicles (DCVs). Adaptor protein 3 (AP-3) assembles the membrane proteins that confer regulated release of DCVs and is thought to promote their trafficking from endosomes directly to maturing DCVs. We now find that regulated monoamine release from DCVs requires sorting nexin 5 (SNX5). Loss of SNX5 disrupts trafficking of the vesicular monoamine transporter (VMAT) to DCVs. The mechanism involves a role for SNX5 in retrograde transport of VMAT from endosomes to the TGN. However, this role for SNX5 conflicts with the proposed function of AP-3 in trafficking from endosomes directly to DCVs. We now identify a transient role for AP-3 at the TGN, where it associates with DCV cargo. Thus, retrograde transport from endosomes by SNX5 enables DCV assembly at the TGN by AP-3, resolving the apparent antagonism. A novel role for AP-3 at the TGN has implications for other organelles that also depend on this adaptor.
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Affiliation(s)
- Hongfei Xu
- Departments of Neurology and Physiology, University of California San Francisco, San Francisco, CA
- Jiangsu Key Laboratory of Xenotransplantation, School of Basic Medical Science, Nanjing Medical University, Nanjing, China
| | - Fei Chang
- Jiangsu Key Laboratory of Xenotransplantation, School of Basic Medical Science, Nanjing Medical University, Nanjing, China
| | - Shweta Jain
- Departments of Neurology and Physiology, University of California San Francisco, San Francisco, CA
| | - Bradley Austin Heller
- Departments of Neurology and Physiology, University of California San Francisco, San Francisco, CA
| | - Xu Han
- Jiangsu Key Laboratory of Xenotransplantation, School of Basic Medical Science, Nanjing Medical University, Nanjing, China
| | - Yongjian Liu
- Jiangsu Key Laboratory of Xenotransplantation, School of Basic Medical Science, Nanjing Medical University, Nanjing, China
- Departments of Pharmacology and Biological Chemistry, University of Pittsburgh, Pittsburgh, PA
| | - Robert H. Edwards
- Departments of Neurology and Physiology, University of California San Francisco, San Francisco, CA
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17
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Atiakshin DA, Kostin AA, Trotsenko ID, Shishkina VV, Tiemann M, Buchwalow IB. Carboxypeptidase A3 in the structure of the protease phenotype of mast cells: cytophysiological aspects. RUDN JOURNAL OF MEDICINE 2022. [DOI: 10.22363/2313-0245-2022-26-1-9-33] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/01/2022] Open
Abstract
Carboxypeptidase A3 (CPA3) is a specific protease of mast cells (MC) with variable expression and appears to be one of the preformed components of the secretome. CPA3 is involved in regulation of the state of a specifi tissue microenvironment and components of the integrative-buffer metabolic environment in adaptive and pathological processes; it affects implementation of the innate immunity, mechanisms of angiogenesis, processes of the extracellular matrix remodeling, etc. CPA3 identification using protocols of multiplex immunohistochemistry allows specifying details of the organ-specific mast cell population features, including the protease phenotype, mechanisms of biogenesis with cytoand histotopographic criteria, and features of secretory pathways. Numerous biological effects of CPA3, including participation in the regulation of the pulmonary parenchyma and systemic blood flow, in biogenesis and remodeling of the fibrous component of the extracellular matrix, in epigenetic reprogramming, determine the importance of fundamental investigation of the physiological activity of protease and its involvement in the implementation of pathological processes. Further studies will contribute to the detection of the translational value of the mast cell CPA3 expression features as a prognostic factor and a promising molecular target for treatment of socially significant diseases.
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18
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Atiakshin DA, Kostin AA, Trotsenko ID, Shishkina VV, Tiemann M, Buchwalow IB. Carboxypeptidase A3 in the structure of the protease phenotype of mast cells: cytophysiological aspects. RUDN JOURNAL OF MEDICINE 2022. [DOI: 10.22363/2313-0245-2022-26-1-9-32] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/01/2022] Open
Abstract
Carboxypeptidase A3 (CPA3) is a specific protease of mast cells (MC) with variable expression and appears to be one of the preformed components of the secretome. CPA3 is involved in regulation of the state of a specifi tissue microenvironment and components of the integrative-buffer metabolic environment in adaptive and pathological processes; it affects implementation of the innate immunity, mechanisms of angiogenesis, processes of the extracellular matrix remodeling, etc. CPA3 identification using protocols of multiplex immunohistochemistry allows specifying details of the organ-specific mast cell population features, including the protease phenotype, mechanisms of biogenesis with cyto- and histotopographic criteria, and features of secretory pathways. Numerous biological effects of CPA3, including participation in the regulation of the pulmonary parenchyma and systemic blood flow, in biogenesis and remodeling of the fibrous component of the extracellular matrix, in epigenetic reprogramming, determine the importance of fundamental investigation of the biological activity and regulation of pathological processes of CPA3. Further studies will contribute to the detection of the true value of the mast cell CPA3 expression features as a prognostic factor and a promising molecular target for treatment of socially significant diseases.
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19
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Rohli KE, Boyer CK, Blom SE, Stephens SB. Nutrient Regulation of Pancreatic Islet β-Cell Secretory Capacity and Insulin Production. Biomolecules 2022; 12:335. [PMID: 35204835 PMCID: PMC8869698 DOI: 10.3390/biom12020335] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2022] [Revised: 02/16/2022] [Accepted: 02/17/2022] [Indexed: 01/27/2023] Open
Abstract
Pancreatic islet β-cells exhibit tremendous plasticity for secretory adaptations that coordinate insulin production and release with nutritional demands. This essential feature of the β-cell can allow for compensatory changes that increase secretory output to overcome insulin resistance early in Type 2 diabetes (T2D). Nutrient-stimulated increases in proinsulin biosynthesis may initiate this β-cell adaptive compensation; however, the molecular regulators of secretory expansion that accommodate the increased biosynthetic burden of packaging and producing additional insulin granules, such as enhanced ER and Golgi functions, remain poorly defined. As these adaptive mechanisms fail and T2D progresses, the β-cell succumbs to metabolic defects resulting in alterations to glucose metabolism and a decline in nutrient-regulated secretory functions, including impaired proinsulin processing and a deficit in mature insulin-containing secretory granules. In this review, we will discuss how the adaptative plasticity of the pancreatic islet β-cell's secretory program allows insulin production to be carefully matched with nutrient availability and peripheral cues for insulin signaling. Furthermore, we will highlight potential defects in the secretory pathway that limit or delay insulin granule biosynthesis, which may contribute to the decline in β-cell function during the pathogenesis of T2D.
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Affiliation(s)
- Kristen E. Rohli
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52242, USA; (K.E.R.); (C.K.B.); (S.E.B.)
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Iowa, Iowa City, IA 52242, USA
| | - Cierra K. Boyer
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52242, USA; (K.E.R.); (C.K.B.); (S.E.B.)
- Department of Neuroscience and Pharmacology, University of Iowa, Iowa City, IA 52242, USA
| | - Sandra E. Blom
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52242, USA; (K.E.R.); (C.K.B.); (S.E.B.)
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Iowa, Iowa City, IA 52242, USA
| | - Samuel B. Stephens
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52242, USA; (K.E.R.); (C.K.B.); (S.E.B.)
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Iowa, Iowa City, IA 52242, USA
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20
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Sobiepanek A, Kuryk Ł, Garofalo M, Kumar S, Baran J, Musolf P, Siebenhaar F, Fluhr JW, Kobiela T, Plasenzotti R, Kuchler K, Staniszewska M. The Multifaceted Roles of Mast Cells in Immune Homeostasis, Infections and Cancers. Int J Mol Sci 2022; 23:2249. [PMID: 35216365 PMCID: PMC8875910 DOI: 10.3390/ijms23042249] [Citation(s) in RCA: 23] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2022] [Revised: 02/05/2022] [Accepted: 02/07/2022] [Indexed: 02/07/2023] Open
Abstract
Mast cells (MCs) play important roles in normal immune responses and pathological states. The location of MCs on the boundaries between tissues and the external environment, including gut mucosal surfaces, lungs, skin, and around blood vessels, suggests a multitude of immunological functions. Thus, MCs are pivotal for host defense against different antigens, including allergens and microbial pathogens. MCs can produce and respond to physiological mediators and chemokines to modulate inflammation. As long-lived, tissue-resident cells, MCs indeed mediate acute inflammatory responses such as those evident in allergic reactions. Furthermore, MCs participate in innate and adaptive immune responses to bacteria, viruses, fungi, and parasites. The control of MC activation or stabilization is a powerful tool in regulating tissue homeostasis and pathogen clearance. Moreover, MCs contribute to maintaining the homeostatic equilibrium between host and resident microbiota, and they engage in crosstalk between the resident and recruited hematopoietic cells. In this review, we provide a comprehensive overview of the functions of MCs in health and disease. Further, we discuss how mouse models of MC deficiency have become useful tools for establishing MCs as a potential cellular target for treating inflammatory disorders.
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Affiliation(s)
- Anna Sobiepanek
- Faculty of Chemistry, Warsaw University of Technology, Noakowskiego 3, 00-664 Warsaw, Poland; (A.S.); (J.B.); (P.M.); (T.K.)
| | - Łukasz Kuryk
- National Institute of Public Health NIH—National Institute of Research, 00-791 Warsaw, Poland;
- Clinical Science, Targovax Oy, Lars Sonckin kaari 14, 02600 Espoo, Finland;
| | - Mariangela Garofalo
- Department of Pharmaceutical and Pharmacological Sciences, University of Padova, Via F. Marzolo 5, 35131 Padova, Italy;
| | - Sandeep Kumar
- Clinical Science, Targovax Oy, Lars Sonckin kaari 14, 02600 Espoo, Finland;
| | - Joanna Baran
- Faculty of Chemistry, Warsaw University of Technology, Noakowskiego 3, 00-664 Warsaw, Poland; (A.S.); (J.B.); (P.M.); (T.K.)
| | - Paulina Musolf
- Faculty of Chemistry, Warsaw University of Technology, Noakowskiego 3, 00-664 Warsaw, Poland; (A.S.); (J.B.); (P.M.); (T.K.)
| | - Frank Siebenhaar
- Institute of Allergology, Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, 10117 Berlin, Germany; (F.S.); (J.W.F.)
- Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, Allergology and Immunology, 12203 Berlin, Germany
| | - Joachim Wilhelm Fluhr
- Institute of Allergology, Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, 10117 Berlin, Germany; (F.S.); (J.W.F.)
- Fraunhofer Institute for Translational Medicine and Pharmacology ITMP, Allergology and Immunology, 12203 Berlin, Germany
| | - Tomasz Kobiela
- Faculty of Chemistry, Warsaw University of Technology, Noakowskiego 3, 00-664 Warsaw, Poland; (A.S.); (J.B.); (P.M.); (T.K.)
| | - Roberto Plasenzotti
- Department of Biomedical Research, Medical University of Vienna, Währingergürtel 18-20, 1090 Vienna, Austria;
| | - Karl Kuchler
- Max Perutz Labs Vienna, Center for Medical Biochemistry, Medical University of Vienna, Campus Vienna Biocenter, Dr. Bohr-Gasse 9/2, 1030 Vienna, Austria;
| | - Monika Staniszewska
- Centre for Advanced Materials and Technologies, Warsaw University of Technology, Poleczki 19, 02-822 Warsaw, Poland
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21
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Atiakshin D, Kostin A, Trotsenko I, Samoilova V, Buchwalow I, Tiemann M. Carboxypeptidase A3—A Key Component of the Protease Phenotype of Mast Cells. Cells 2022; 11:cells11030570. [PMID: 35159379 PMCID: PMC8834431 DOI: 10.3390/cells11030570] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2022] [Revised: 02/04/2022] [Accepted: 02/05/2022] [Indexed: 11/16/2022] Open
Abstract
Carboxypeptidase A3 (CPA3) is a specific mast cell (MC) protease with variable expression. This protease is one of the preformed components of the secretome. During maturation of granules, CPA3 becomes an active enzyme with a characteristic localization determining the features of the cytological and ultrastructural phenotype of MC. CPA3 takes part in the regulation of a specific tissue microenvironment, affecting the implementation of innate immunity, the mechanisms of angiogenesis, the processes of remodeling of the extracellular matrix, etc. Characterization of CPA3 expression in MC can be used to refine the MC classification, help in a prognosis, and increase the effectiveness of targeted therapy.
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Affiliation(s)
- Dmitri Atiakshin
- Research and Educational Resource Center for Immunophenotyping, Digital Spatial Profiling and Ultrastructural Analysis Innovative Technologies, Peoples’ Friendship University of Russia, Miklukho-Maklaya Str. 6, 117198 Moscow, Russia; (D.A.); (A.K.); (I.T.)
- Research Institute of Experimental Biology and Medicine, Burdenko Voronezh State Medical University, Studencheskaya Str. 10, 394036 Voronezh, Russia
| | - Andrey Kostin
- Research and Educational Resource Center for Immunophenotyping, Digital Spatial Profiling and Ultrastructural Analysis Innovative Technologies, Peoples’ Friendship University of Russia, Miklukho-Maklaya Str. 6, 117198 Moscow, Russia; (D.A.); (A.K.); (I.T.)
| | - Ivan Trotsenko
- Research and Educational Resource Center for Immunophenotyping, Digital Spatial Profiling and Ultrastructural Analysis Innovative Technologies, Peoples’ Friendship University of Russia, Miklukho-Maklaya Str. 6, 117198 Moscow, Russia; (D.A.); (A.K.); (I.T.)
| | - Vera Samoilova
- Institute for Hematopathology, Fangdieckstr. 75a, 22547 Hamburg, Germany; (V.S.); (M.T.)
| | - Igor Buchwalow
- Research and Educational Resource Center for Immunophenotyping, Digital Spatial Profiling and Ultrastructural Analysis Innovative Technologies, Peoples’ Friendship University of Russia, Miklukho-Maklaya Str. 6, 117198 Moscow, Russia; (D.A.); (A.K.); (I.T.)
- Institute for Hematopathology, Fangdieckstr. 75a, 22547 Hamburg, Germany; (V.S.); (M.T.)
- Correspondence: ; Tel.: +49-(040)-7070-85317; Fax: +49-(040)-7070-85110
| | - Markus Tiemann
- Institute for Hematopathology, Fangdieckstr. 75a, 22547 Hamburg, Germany; (V.S.); (M.T.)
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22
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Reck J, Beuret N, Demirci E, Prescianotto-Baschong C, Spiess M. Small disulfide loops in peptide hormones mediate self-aggregation and secretory granule sorting. Life Sci Alliance 2022; 5:5/5/e202101279. [PMID: 35086936 PMCID: PMC8807871 DOI: 10.26508/lsa.202101279] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2021] [Revised: 01/12/2022] [Accepted: 01/14/2022] [Indexed: 11/24/2022] Open
Abstract
Unlike constitutively secreted proteins, peptide hormones are stored in densely packed secretory granules, before regulated release upon stimulation. Secretory granules are formed at the TGN by self-aggregation of prohormones as functional amyloids. The nonapeptide hormone vasopressin, which forms a small disulfide loop, was shown to be responsible for granule formation of its precursor in the TGN as well as for toxic fibrillar aggregation of unfolded mutants in the ER. Several other hormone precursors also contain similar small disulfide loops suggesting their function as a general device to mediate aggregation for granule sorting. To test this hypothesis, we studied the capacity of small disulfide loops of different hormone precursors to mediate aggregation in the ER and the TGN. They indeed induced ER aggregation in Neuro-2a and COS-1 cells. Fused to a constitutively secreted reporter protein, they also promoted sorting into secretory granules, enhanced stimulated secretion, and increased Lubrol insolubility in AtT20 cells. These results support the hypothesis that small disulfide loops act as novel signals for sorting into secretory granules by self-aggregation.
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23
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Liu Y, Zhang Y, Sun Y, Ding F. A buried glutamate in the cross-β core renders β-endorphin fibrils reversible. NANOSCALE 2021; 13:19593-19603. [PMID: 34812835 PMCID: PMC8674924 DOI: 10.1039/d1nr05679d] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/13/2023]
Abstract
Functional amyloids are abundant in living organisms from prokaryotes to eukaryotes playing diverse biological roles. In contrast to the irreversible aggregation of most known pathological amyloids, we postulate that naturally-occurring functional amyloids are reversible under evolutionary pressure to be able to modulate the fibrillization process, reuse the composite peptides, or perform their biological functions. β-Endorphin, an endogenous opioid peptide hormone, forms such kinds of reversible amyloid fibrils in secretory granules for efficient storage and returns to the functional state of monomers upon release into the blood. The environmental change between low pH in secretory granules and neutral pH in extracellular spaces is believed to drive the reversible fibrillization of β-endorphin. Here, we investigate the critical role of a buried glutamate, Glu8, in the pH-responsive disassembly of β-endorphin fibrils using all-atom molecular dynamics simulations along with structure-based pKa prediction. The fibril was stable at pH 5.5 or lower with all the buried Glu8 residues protonated and neutrally charged. After switching to neutral pH, the Glu8 residues of peptides at the outer layers of the ordered fibrils became deprotonated due to partial solvent exposure, causing sheet-to-coil conformational changes and subsequent exposure of adjacent Glu8 residues in the inner chains. Via iterative deprotonation of Glu8 and induced structural disruption, all Glu8 residues would be progressively deprotonated. Electrostatic repulsion between deprotonated Glu8 residues along with their high solvation tendency disrupted the hydrogen bonding between the β1 strands and increased the solvent exposure of those otherwise buried residues in the cross-β core. Overall, our computational study reveals that the strategic positioning of ionizable residues into the cross-β core is a potential approach for designing reversible amyloid fibrils as pH-responsive smart bio-nanomaterials.
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Affiliation(s)
- Yuying Liu
- School of Physical Science and Technology, Ningbo University, Ningbo 315211, China
| | - Yu Zhang
- School of Physical Science and Technology, Ningbo University, Ningbo 315211, China
| | - Yunxiang Sun
- School of Physical Science and Technology, Ningbo University, Ningbo 315211, China
- Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, USA.
| | - Feng Ding
- Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, USA.
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24
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Germanos M, Gao A, Taper M, Yau B, Kebede MA. Inside the Insulin Secretory Granule. Metabolites 2021; 11:metabo11080515. [PMID: 34436456 PMCID: PMC8401130 DOI: 10.3390/metabo11080515] [Citation(s) in RCA: 28] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/08/2021] [Revised: 08/03/2021] [Accepted: 08/03/2021] [Indexed: 12/19/2022] Open
Abstract
The pancreatic β-cell is purpose-built for the production and secretion of insulin, the only hormone that can remove glucose from the bloodstream. Insulin is kept inside miniature membrane-bound storage compartments known as secretory granules (SGs), and these specialized organelles can readily fuse with the plasma membrane upon cellular stimulation to release insulin. Insulin is synthesized in the endoplasmic reticulum (ER) as a biologically inactive precursor, proinsulin, along with several other proteins that will also become members of the insulin SG. Their coordinated synthesis enables synchronized transit through the ER and Golgi apparatus for congregation at the trans-Golgi network, the initiating site of SG biogenesis. Here, proinsulin and its constituents enter the SG where conditions are optimized for proinsulin processing into insulin and subsequent insulin storage. A healthy β-cell is continually generating SGs to supply insulin in vast excess to what is secreted. Conversely, in type 2 diabetes (T2D), the inability of failing β-cells to secrete may be due to the limited biosynthesis of new insulin. Factors that drive the formation and maturation of SGs and thus the production of insulin are therefore critical for systemic glucose control. Here, we detail the formative hours of the insulin SG from the luminal perspective. We do this by mapping the journey of individual members of the SG as they contribute to its genesis.
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25
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Ma CIJ, Brill JA. Quantitation of Secretory Granule Size in Drosophila Larval Salivary Glands. Bio Protoc 2021; 11:e4039. [PMID: 34250205 DOI: 10.21769/bioprotoc.4039] [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: 01/08/2021] [Revised: 03/14/2021] [Accepted: 03/24/2021] [Indexed: 11/02/2022] Open
Abstract
Maturation of secretory granules is a crucial process that ensures the bioactivity of cargo proteins undergoing regulated secretion. In Drosophila melanogaster, the larval salivary glands produce secretory granules that are up to four-fold larger in cross-sectional area after maturation. Therefore, we developed a live imaging microscopy approach to quantitate the size of secretory granules with a view to identifying genes involved in their maturation. Here, we describe the procedures of larval salivary gland dissection and sample preparation for live imaging with a fluorescence confocal microscope. Furthermore, we describe the workflow for measuring the size of secretory granules by cross-sectional surface area and statistical analysis. Our live imaging microscopy method provides a reliable read-out for the status of secretory granule maturation in Drosophila larval salivary glands.
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Affiliation(s)
- Cheng-I J Ma
- Cell Biology Program, The Hospital for Sick Children, Toronto, Ontario, Canada.,Institute of Medical Science, University of Toronto, Toronto, Canada
| | - Julie A Brill
- Cell Biology Program, The Hospital for Sick Children, Toronto, Ontario, Canada.,Institute of Medical Science, University of Toronto, Toronto, Canada.,Department of Molecular Genetics, University of Toronto, Toronto, Canada
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26
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Isolation and Proteomics of the Insulin Secretory Granule. Metabolites 2021; 11:metabo11050288. [PMID: 33946444 PMCID: PMC8147143 DOI: 10.3390/metabo11050288] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2021] [Revised: 04/28/2021] [Accepted: 04/28/2021] [Indexed: 12/21/2022] Open
Abstract
Insulin, a vital hormone for glucose homeostasis is produced by pancreatic beta-cells and when secreted, stimulates the uptake and storage of glucose from the blood. In the pancreas, insulin is stored in vesicles termed insulin secretory granules (ISGs). In Type 2 diabetes (T2D), defects in insulin action results in peripheral insulin resistance and beta-cell compensation, ultimately leading to dysfunctional ISG production and secretion. ISGs are functionally dynamic and many proteins present either on the membrane or in the lumen of the ISG may modulate and affect different stages of ISG trafficking and secretion. Previously, studies have identified few ISG proteins and more recently, proteomics analyses of purified ISGs have uncovered potential novel ISG proteins. This review summarizes the proteins identified in the current ISG proteomes from rat insulinoma INS-1 and INS-1E cell lines. Here, we also discuss techniques of ISG isolation and purification, its challenges and potential future directions.
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27
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Tokuo H, Komaba S, Coluccio LM. In pancreatic β-cells myosin 1b regulates glucose-stimulated insulin secretion by modulating an early step in insulin granule trafficking from the Golgi. Mol Biol Cell 2021; 32:1210-1220. [PMID: 33826361 PMCID: PMC8351557 DOI: 10.1091/mbc.e21-03-0094] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022] Open
Abstract
Pancreatic β-cells secrete insulin, which controls blood glucose levels, and defects in insulin secretion are responsible for diabetes mellitus. The actin cytoskeleton and some myosins support insulin granule trafficking and release, although a role for the class I myosin Myo1b, an actin- and membrane-associated load-sensitive motor, in insulin biology is unknown. We found by immunohistochemistry that Myo1b is expressed in islet cells of the rat pancreas. In cultured rat insulinoma 832/13 cells, Myo1b localized near actin patches, the trans-Golgi network (TGN) marker TGN38, and insulin granules in the perinuclear region. Myo1b depletion by small interfering RNA in 832/13 cells reduced intracellular proinsulin and insulin content and glucose-stimulated insulin secretion (GSIS) and led to the accumulation of (pro)insulin secretory granules (SGs) at the TGN. Using an in situ fluorescent pulse-chase strategy to track nascent proinsulin, Myo1b depletion in insulinoma cells reduced the number of (pro)insulin-containing SGs budding from the TGN. The studies indicate for the first time that in pancreatic β-cells Myo1b controls GSIS at least in part by mediating an early stage in insulin granule trafficking from the TGN.
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Affiliation(s)
- Hiroshi Tokuo
- Department of Physiology & Biophysics, Boston University School of Medicine, Boston, MA 02118-2518
| | - Shigeru Komaba
- Department of Physiology & Biophysics, Boston University School of Medicine, Boston, MA 02118-2518
| | - Lynne M Coluccio
- Department of Physiology & Biophysics, Boston University School of Medicine, Boston, MA 02118-2518
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28
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Ma CIJ, Burgess J, Brill JA. Maturing secretory granules: Where secretory and endocytic pathways converge. Adv Biol Regul 2021; 80:100807. [PMID: 33866198 DOI: 10.1016/j.jbior.2021.100807] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2021] [Revised: 03/10/2021] [Accepted: 03/18/2021] [Indexed: 10/21/2022]
Abstract
Secretory granules (SGs) are specialized organelles responsible for the storage and regulated release of various biologically active molecules from the endocrine and exocrine systems. Thus, proper SG biogenesis is critical to normal animal physiology. Biogenesis of SGs starts at the trans-Golgi network (TGN), where immature SGs (iSGs) bud off and undergo maturation before fusing with the plasma membrane (PM). How iSGs mature is unclear, but emerging studies have suggested an important role for the endocytic pathway. The requirement for endocytic machinery in SG maturation blurs the line between SGs and another class of secretory organelles called lysosome-related organelles (LROs). Therefore, it is important to re-evaluate the differences and similarities between SGs and LROs.
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Affiliation(s)
- Cheng-I Jonathan Ma
- Cell Biology Program, The Hospital for Sick Children, PGCRL Building, Room 15.9716, 686 Bay Street, Toronto, Ontario, M5G 0A4, Canada; Institute of Medical Science, University of Toronto, Medical Sciences Building, Room 2374, 1 King's College Circle, Toronto, Ontario, M5S 1A8, Canada
| | - Jason Burgess
- Cell Biology Program, The Hospital for Sick Children, PGCRL Building, Room 15.9716, 686 Bay Street, Toronto, Ontario, M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Medical Sciences Building, Room 4396, 1 King's College Circle, Toronto, Ontario, M5S 1A8, Canada
| | - Julie A Brill
- Cell Biology Program, The Hospital for Sick Children, PGCRL Building, Room 15.9716, 686 Bay Street, Toronto, Ontario, M5G 0A4, Canada; Institute of Medical Science, University of Toronto, Medical Sciences Building, Room 2374, 1 King's College Circle, Toronto, Ontario, M5S 1A8, Canada; Department of Molecular Genetics, University of Toronto, Medical Sciences Building, Room 4396, 1 King's College Circle, Toronto, Ontario, M5S 1A8, Canada.
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29
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Seuring C, Verasdonck J, Gath J, Ghosh D, Nespovitaya N, Wälti MA, Maji SK, Cadalbert R, Güntert P, Meier BH, Riek R. The three-dimensional structure of human β-endorphin amyloid fibrils. Nat Struct Mol Biol 2020; 27:1178-1184. [PMID: 33046908 DOI: 10.1038/s41594-020-00515-z] [Citation(s) in RCA: 49] [Impact Index Per Article: 9.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2020] [Accepted: 09/08/2020] [Indexed: 11/09/2022]
Abstract
In the pituitary gland, hormones are stored in a functional amyloid state within acidic secretory granules before they are released into the blood. To gain a detailed understanding of the structure-function relationship of amyloids in hormone secretion, the three-dimensional (3D) structure of the amyloid fibril of the human hormone β-endorphin was determined by solid-state NMR. We find that β-endorphin fibrils are in a β-solenoid conformation with a protonated glutamate residue in their fibrillar core. During exocytosis of the hormone amyloid the pH increases from acidic in the secretory granule to neutral level in the blood, thus it is suggested-and supported with mutagenesis data-that the pH change in the cellular milieu acts through the deprotonation of glutamate 8 to release the hormone from the amyloid. For amyloid disassembly in the blood, it is proposed that the pH change acts together with a buffer composition change and hormone dilution. In the pituitary gland, peptide hormones can be stored as amyloid fibrils within acidic secretory granules before release into the blood stream. Here, we use solid-state NMR to determine the 3D structure of the amyloid fiber formed by the human hormone β-endorphin. We find that β-endorphin fibrils are in a β-solenoid conformation that is generally reminiscent of other functional amyloids. In the β-endorphin amyloid, every layer of the β-solenoid is composed of a single peptide and protonated Glu8 is located in the fibrillar core. The secretory granule has an acidic pH but, on exocytosis, the β-endorphin fibril would encounter neutral pH conditions (pH 7.4) in the blood; this pH change would result in deprotonation of Glu8 to release the hormone peptide from the amyloid. Analyses of β-endorphin variants carrying mutations in Glu8 support the role of the protonation state of this residue in fibril disassembly, among other environmental changes.
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Affiliation(s)
- Carolin Seuring
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland.,Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron, Hamburg, Germany
| | - Joeri Verasdonck
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland
| | - Julia Gath
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland
| | - Dhimam Ghosh
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland.,Department of Biosciences and Bioengineering, IIT Bombay, Powai, Mumbai, India
| | | | | | - Samir K Maji
- Department of Biosciences and Bioengineering, IIT Bombay, Powai, Mumbai, India
| | | | - Peter Güntert
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland.,Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt am Main, Frankfurt am Main, Germany.,Graduate School of Science, Tokyo Metropolitan University, Tokyo, Japan
| | - Beat H Meier
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland.
| | - Roland Riek
- Laboratory of Physical Chemistry, ETH Zürich, Zürich, Switzerland. .,Structural Biology Laboratory, The Salk Institute, La Jolla, CA, USA.
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30
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Chen H, Victor AK, Klein J, Tacer KF, Tai DJ, de Esch C, Nuttle A, Temirov J, Burnett LC, Rosenbaum M, Zhang Y, Ding L, Moresco JJ, Diedrich JK, Yates JR, Tillman HS, Leibel RL, Talkowski ME, Billadeau DD, Reiter LT, Potts PR. Loss of MAGEL2 in Prader-Willi syndrome leads to decreased secretory granule and neuropeptide production. JCI Insight 2020; 5:138576. [PMID: 32879135 PMCID: PMC7526459 DOI: 10.1172/jci.insight.138576] [Citation(s) in RCA: 41] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2020] [Accepted: 07/22/2020] [Indexed: 12/17/2022] Open
Abstract
Prader-Willi syndrome (PWS) is a developmental disorder caused by loss of maternally imprinted genes on 15q11-q13, including melanoma antigen gene family member L2 (MAGEL2). The clinical phenotypes of PWS suggest impaired hypothalamic neuroendocrine function; however, the exact cellular defects are unknown. Here, we report deficits in secretory granule (SG) abundance and bioactive neuropeptide production upon loss of MAGEL2 in humans and mice. Unbiased proteomic analysis of Magel2pΔ/m+ mice revealed a reduction in components of SG in the hypothalamus that was confirmed in 2 PWS patient-derived neuronal cell models. Mechanistically, we show that proper endosomal trafficking by the MAGEL2-regulated WASH complex is required to prevent aberrant lysosomal degradation of SG proteins and reduction of mature SG abundance. Importantly, loss of MAGEL2 in mice, NGN2-induced neurons, and human patients led to reduced neuropeptide production. Thus, MAGEL2 plays an important role in hypothalamic neuroendocrine function, and cellular defects in this pathway may contribute to PWS disease etiology. Moreover, these findings suggest unanticipated approaches for therapeutic intervention.
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Affiliation(s)
- Helen Chen
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - A Kaitlyn Victor
- Department of Neurology, Department of Pediatrics, and Department of Anatomy and Neurobiology, University of Tennessee Health Science Center, Memphis, Tennessee, USA
| | - Jonathon Klein
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - Klementina Fon Tacer
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - Derek Jc Tai
- Center for Genomic Medicine, Department of Neurology, Department of Pathology, and Department of Psychiatry, Massachusetts General Hospital, Boston, Massachusetts, USA.,Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA.,Program in Medical and Population Genetics and Stanley Center for Psychiatric Research, Broad Institute, Cambridge, Massachusetts, USA
| | - Celine de Esch
- Center for Genomic Medicine, Department of Neurology, Department of Pathology, and Department of Psychiatry, Massachusetts General Hospital, Boston, Massachusetts, USA.,Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA.,Program in Medical and Population Genetics and Stanley Center for Psychiatric Research, Broad Institute, Cambridge, Massachusetts, USA
| | - Alexander Nuttle
- Center for Genomic Medicine, Department of Neurology, Department of Pathology, and Department of Psychiatry, Massachusetts General Hospital, Boston, Massachusetts, USA.,Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA.,Program in Medical and Population Genetics and Stanley Center for Psychiatric Research, Broad Institute, Cambridge, Massachusetts, USA
| | - Jamshid Temirov
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - Lisa C Burnett
- Levo Therapeutics, Inc., Skokie, Illinois, USA.,Division of Molecular Genetics, Department of Pediatrics, and Naomi Berrie Diabetes Center, Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, New York, New York, USA
| | - Michael Rosenbaum
- Division of Molecular Genetics, Department of Pediatrics, and Naomi Berrie Diabetes Center, Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, New York, New York, USA
| | - Yiying Zhang
- Division of Molecular Genetics, Department of Pediatrics, and Naomi Berrie Diabetes Center, Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, New York, New York, USA
| | - Li Ding
- Division of Oncology Research and Schulze Center for Novel Therapeutics, Mayo Clinic, Rochester, Minnesota, USA
| | - James J Moresco
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, California, USA
| | - Jolene K Diedrich
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, California, USA
| | - John R Yates
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, California, USA
| | - Heather S Tillman
- Veterinary Pathology Core, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
| | - Rudolph L Leibel
- Division of Molecular Genetics, Department of Pediatrics, and Naomi Berrie Diabetes Center, Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, New York, New York, USA
| | - Michael E Talkowski
- Center for Genomic Medicine, Department of Neurology, Department of Pathology, and Department of Psychiatry, Massachusetts General Hospital, Boston, Massachusetts, USA.,Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA.,Program in Medical and Population Genetics and Stanley Center for Psychiatric Research, Broad Institute, Cambridge, Massachusetts, USA
| | - Daniel D Billadeau
- Division of Oncology Research and Schulze Center for Novel Therapeutics, Mayo Clinic, Rochester, Minnesota, USA
| | - Lawrence T Reiter
- Department of Neurology, Department of Pediatrics, and Department of Anatomy and Neurobiology, University of Tennessee Health Science Center, Memphis, Tennessee, USA
| | - Patrick Ryan Potts
- Department of Cell and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA
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31
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Zhang XW, Hatamie A, Ewing AG. Simultaneous Quantification of Vesicle Size and Catecholamine Content by Resistive Pulses in Nanopores and Vesicle Impact Electrochemical Cytometry. J Am Chem Soc 2020; 142:4093-4097. [PMID: 32069039 PMCID: PMC7108759 DOI: 10.1021/jacs.9b13221] [Citation(s) in RCA: 46] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
![]()
We have developed the means to simultaneously
measure the physical
size and count catecholamine molecules in individual nanometer transmitter
vesicles. This is done by combining resistive pulse (RP) measurements
in a nanopore pipet and vesicle impact electrochemical cytometry (VIEC)
at an electrode as the vesicle exits the nanopore. Analysis of freshly
isolated bovine adrenal vesicles shows that the size and internal
catecholamine concentration of vesicles varies with the occurrence
of a dense core inside the vesicles. These results might benefit the
understanding about the vesicles maturation, especially involving
the “sorting by retention” process and concentration
increase of intravesicular catecholamine. The methodology is applicable
to understanding soft nanoparticle collisions on electrodes, vesicles
in exocytosis and phagocytosis, intracellular vesicle transport, and
analysis of electroactive drugs in exosomes.
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Affiliation(s)
- Xin-Wei Zhang
- Department of Chemistry and Molecular Biology, University of Gothenburg, SE-412 96 Gothenburg, Sweden
| | - Amir Hatamie
- Department of Chemistry and Molecular Biology, University of Gothenburg, SE-412 96 Gothenburg, Sweden
| | - Andrew G Ewing
- Department of Chemistry and Molecular Biology, University of Gothenburg, SE-412 96 Gothenburg, Sweden
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Zhou Y, Liu Z, Zhang S, Zhuang R, Liu H, Liu X, Qiu X, Zhang M, Zheng Y, Li L, Hong W, Wang T. RILP Restricts Insulin Secretion Through Mediating Lysosomal Degradation of Proinsulin. Diabetes 2020; 69:67-82. [PMID: 31624142 DOI: 10.2337/db19-0086] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/24/2019] [Accepted: 10/06/2019] [Indexed: 11/13/2022]
Abstract
Insulin secretion is tightly regulated by membrane trafficking. RILP (Rab7 interacting lysosomal protein) regulates the endocytic trafficking, but its role in insulin secretion has not been investigated. In this study, we found that overexpression of RILP inhibited insulin secretion in both the β-cell lines and freshly isolated islets. Consequently, the expression of RILP in islets suppressed the ability to recover the glucose homeostasis in type 1 diabetes mice upon transplantation. Of physiological relevance is that RILP expression was upregulated in the diabetic mouse islets. Mechanistically, overexpression of RILP induced insulin granule clustering, decreased the number of proinsulin-containing granules in β-cells, and significantly promoted proinsulin degradation. Conversely, RILP depletion sustained proinsulin and increased insulin secretion. The proinsulin degradation induced by RILP expression was inhibited by lysosomal inhibitors and was Rab7-dependent. Finally, we showed that RILP interacts with insulin granule-associated Rab26 to restrict insulin secretion. This study presents a new pathway regulating insulin secretion and mechanically demonstrates a novel function of RILP in modulating insulin secretion through mediating the lysosomal degradation of proinsulin.
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Affiliation(s)
- Yuxia Zhou
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Zhiyu Liu
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Shengmei Zhang
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Ruijuan Zhuang
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Huiying Liu
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Xiaoqing Liu
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Xi Qiu
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Ming Zhang
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Yanpan Zheng
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Liangcheng Li
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
| | - Wanjin Hong
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
- Institute of Molecular and Cell Biology, A STAR (Agency of Science, Technology and Research), Singapore, Singapore
| | - Tuanlao Wang
- School of Pharmaceutical Sciences, State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, Xiamen University, Fujian, China
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Morishita Y, Arvan P. Lessons from animal models of endocrine disorders caused by defects of protein folding in the secretory pathway. Mol Cell Endocrinol 2020; 499:110613. [PMID: 31605742 PMCID: PMC6886696 DOI: 10.1016/j.mce.2019.110613] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/24/2019] [Revised: 09/26/2019] [Accepted: 10/04/2019] [Indexed: 02/06/2023]
Abstract
Most peptide hormones originate from secretory protein precursors synthesized within the endoplasmic reticulum (ER). In this specialized organelle, the newly-made prohormones must fold to their native state. Completion of prohormone folding usually occurs prior to migration through the secretory pathway, as unfolded/misfolded prohormones are retained by mechanisms collectively known as ER quality control. Not only do most monomeric prohormones need to fold properly, but many also dimerize or oligomerize within the ER. If oligomerization occurs before completion of monomer folding then when a poorly folded peptide prohormone is retained by quality control mechanisms, it may confer ER retention upon its oligomerization partners. Conversely, oligomerization between well-folded and improperly folded partners might help to override ER quality control, resulting in rescue of misfolded forms. Both scenarios appear to be possible in different animal models of endocrine disorders caused by genetic defects of protein folding in the secretory pathway. In this paper, we briefly review three such conditions, including familial neurohypophyseal diabetes insipidus, insulin-deficient diabetes mellitus, and hypothyroidism with defective thyroglobulin.
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Affiliation(s)
- Yoshiaki Morishita
- Division of Diabetes, Department of Internal Medicine, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan.
| | - Peter Arvan
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan School of Medicine, Brehm Tower Room 5112, 1000, Wall St., Ann Arbor, MI, USA.
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Abstract
When protein/peptides aggregate, they usually form the amyloid state consisting of cross β-sheet structure built by repetitively stacked β-strands forming long fibrils. Amyloids are usually associated with disease including Alzheimer's. However, amyloid has many useful features. It efficiently transforms protein from the soluble to the insoluble state in an essentially two-state process, while its repetitive structure provides high stability and a robust prion-like replication mechanism. Accordingly, amyloid is used by nature in multifaceted and ingenious ways of life, ranging from bacteria and fungi to mammals. These include (1) Structure: Templating for small chemical molecules (Pmel17), biofilm formation in bacteria (curli), assisting aerial hyphae formation in streptomycetes (chaplins) or monolayer formation at a surface (hydrophobins). (2) Reservoirs: A storage state for peptide/proteins to protect them from their surroundings or vice versa (storage of peptide hormones in mammalian secretory granules or major basic protein in eosinophils). (3) Information carriers: The fungal immune system (HET-s prion in Podospora anserina, yeast prions) or long-term memory (e.g., mnemons in yeast, cytoplasmic polyadenylation element-binding protein in aplysia). Aggregation is also used to (4) "suppress" the function of the soluble protein (e.g., Cdc19 in yeast stress granules), or (5) "signaling" through formation of oligomers (e.g., HET-s prion, necroptosis-related proteins RIP1/RIP3). This review summarizes current knowledge on functional amyloids with a focus on the amyloid systems curli in bacteria, HET-s prion in P. anserina, and peptide hormone storage in mammals together with an attempt to highlight differences between functional and disease-associated amyloids.
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Affiliation(s)
- Daniel Otzen
- iNANO, Aarhus University, DK-8000 Aarhus C, Denmark
| | - Roland Riek
- Laboratory of Physical Chemistry, Swiss Federal Institute of Technology, CH-8093 Zürich, Switzerland
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Carbone E, Borges R, Eiden LE, García AG, Hernández-Cruz A. Chromaffin Cells of the Adrenal Medulla: Physiology, Pharmacology, and Disease. Compr Physiol 2019; 9:1443-1502. [PMID: 31688964 DOI: 10.1002/cphy.c190003] [Citation(s) in RCA: 55] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2025]
Abstract
Chromaffin cells (CCs) of the adrenal gland and the sympathetic nervous system produce the catecholamines (epinephrine and norepinephrine; EPI and NE) needed to coordinate the bodily "fight-or-flight" response to fear, stress, exercise, or conflict. EPI and NE release from CCs is regulated both neurogenically by splanchnic nerve fibers and nonneurogenically by hormones (histamine, corticosteroids, angiotensin, and others) and paracrine messengers [EPI, NE, adenosine triphosphate, opioids, γ-aminobutyric acid (GABA), etc.]. The "stimulus-secretion" coupling of CCs is a Ca2+ -dependent process regulated by Ca2+ entry through voltage-gated Ca2+ channels, Ca2+ pumps, and exchangers and intracellular organelles (RE and mitochondria) and diffusible buffers that provide both Ca2+ -homeostasis and Ca2+ -signaling that ultimately trigger exocytosis. CCs also express Na+ and K+ channels and ionotropic (nAChR and GABAA ) and metabotropic receptors (mACh, PACAP, β-AR, 5-HT, histamine, angiotensin, and others) that make CCs excitable and responsive to autocrine and paracrine stimuli. To maintain high rates of E/NE secretion during stressful conditions, CCs possess a large number of secretory chromaffin granules (CGs) and members of the soluble NSF-attachment receptor complex protein family that allow docking, fusion, and exocytosis of CGs at the cell membrane, and their recycling. This article attempts to provide an updated account of well-established features of the molecular processes regulating CC function, and a survey of the as-yet-unsolved but important questions relating to CC function and dysfunction that have been the subject of intense research over the past 15 years. Examples of CCs as a model system to understand the molecular mechanisms associated with neurodegenerative diseases are also provided. Published 2019. Compr Physiol 9:1443-1502, 2019.
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Affiliation(s)
- Emilio Carbone
- Laboratory of Cellular and Molecular Neuroscience, Department of Drug Science, N.I.S. Centre, University of Torino, Torino, Italy
| | - Ricardo Borges
- Unidad de Farmacología, Facultad de Medicina, Universidad de La Laguna, Tenerife, Spain
| | - Lee E Eiden
- Section on Molecular Neuroscience, National Institute of Mental Health, Bethesda, Maryland, USA
| | - Antonio G García
- Departamento de Farmacología y Terapéutica, Facultad de Medicina, Instituto Teófilo Hernando, Universidad Autónoma de Madrid, Madrid, Spain
| | - Arturo Hernández-Cruz
- Departamento de Neurociencia Cognitiva and Laboratorio Nacional de Canalopatías, Instituto de Fisiología Celular, Universidad Nacional Autonoma de México, Ciudad Universitaria, CDMX, México
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Khawar MB, Gao H, Li W. Mechanism of Acrosome Biogenesis in Mammals. Front Cell Dev Biol 2019; 7:195. [PMID: 31620437 PMCID: PMC6759486 DOI: 10.3389/fcell.2019.00195] [Citation(s) in RCA: 98] [Impact Index Per Article: 16.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2019] [Accepted: 08/29/2019] [Indexed: 11/13/2022] Open
Abstract
During sexual reproduction, two haploid gametes fuse to form the zygote, and the acrosome is essential to this fusion process (fertilization) in animals. The acrosome is a special kind of organelle with a cap-like structure that covers the anterior portion of the head of the spermatozoon. The acrosome is derived from the Golgi apparatus and contains digestive enzymes. With the progress of our understanding of acrosome biogenesis, a number of models have been proposed to address the origin of the acrosome. The acrosome has been regarded as a lysosome-related organelle, and it has been proposed to have originated from the lysosome or the autolysosome. Our review will provide a brief historical overview and highlight recent findings on acrosome biogenesis in mammals.
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Affiliation(s)
- Muhammad Babar Khawar
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Hui Gao
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Wei Li
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
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Bearrows SC, Bauchle CJ, Becker M, Haldeman JM, Swaminathan S, Stephens SB. Chromogranin B regulates early-stage insulin granule trafficking from the Golgi in pancreatic islet β-cells. J Cell Sci 2019; 132:jcs.231373. [PMID: 31182646 DOI: 10.1242/jcs.231373] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2019] [Accepted: 05/31/2019] [Indexed: 12/12/2022] Open
Abstract
Chromogranin B (CgB, also known as CHGB) is abundantly expressed in dense core secretory granules of multiple endocrine tissues and has been suggested to regulate granule biogenesis in some cell types, including the pancreatic islet β-cell, though the mechanisms are poorly understood. Here, we demonstrate a critical role for CgB in regulating secretory granule trafficking in the β-cell. Loss of CgB impairs glucose-stimulated insulin secretion, impedes proinsulin processing to yield increased proinsulin content, and alters the density of insulin-containing granules. Using an in situ fluorescent pulse-chase strategy to track nascent proinsulin, we show that loss of CgB impairs Golgi budding of proinsulin-containing secretory granules, resulting in a substantial delay in trafficking of nascent granules to the plasma membrane with an overall decrease in total plasma membrane-associated granules. These studies demonstrate that CgB is necessary for efficient trafficking of secretory proteins into the budding granule, which impacts the availability of insulin-containing secretory granules for exocytic release.This article has an associated First Person interview with the first author of the paper.
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Affiliation(s)
- Shelby C Bearrows
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52246, USA.,Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Iowa, Iowa City, IA 52246, USA
| | - Casey J Bauchle
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52246, USA.,Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Iowa, Iowa City, IA 52246, USA
| | - McKenzie Becker
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52246, USA.,Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Iowa, Iowa City, IA 52246, USA
| | - Jonathan M Haldeman
- Duke Molecular Physiology Institute, Duke University Medical Center, Durham, NC 27710, USA.,Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA
| | - Svetha Swaminathan
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52246, USA.,Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Iowa, Iowa City, IA 52246, USA
| | - Samuel B Stephens
- Fraternal Order of Eagles Diabetes Research Center, University of Iowa, Iowa City, IA 52246, USA .,Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Iowa, Iowa City, IA 52246, USA
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The Functional Mammalian CRES (Cystatin-Related Epididymal Spermatogenic) Amyloid is Antiparallel β-Sheet Rich and Forms a Metastable Oligomer During Assembly. Sci Rep 2019; 9:9210. [PMID: 31239483 PMCID: PMC6593142 DOI: 10.1038/s41598-019-45545-w] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2019] [Accepted: 06/06/2019] [Indexed: 12/15/2022] Open
Abstract
An amyloid matrix composed of several family 2 cystatins, including the reproductive cystatin CRES, is an integral structure in the mouse epididymal lumen and has proposed functions in sperm maturation and protection. Understanding how CRES amyloid assembles in vitro may provide clues on how the epididymal amyloid matrix forms in vivo. We therefore purified full-length CRES under nondenaturing conditions and followed its aggregation from monomer to amyloid under conditions that may approximate those in the epididymal lumen. CRES transitioned into a metastable oligomer that was resistant to aggregation and only over extended time formed higher-ordered amyloids. High protein concentrations facilitated oligomer assembly and also were required to maintain the metastable state since following dilution the oligomer was no longer detected. Similar to other amyloid precursors, the formation of CRES amyloids correlated with a loss of α-helix and a gain of β-sheet content. However, CRES is unique in that its amyloids are rich in antiparallel β-sheets instead of the more common parallel β-sheets. Taken together, our studies suggest that early metastable oligomers may serve as building blocks for functional amyloid assembly and further reveal that antiparallel β-sheet-rich amyloids can be functional forms.
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Schönherr R, Rudolph JM, Redecke L. Protein crystallization in living cells. Biol Chem 2019; 399:751-772. [PMID: 29894295 DOI: 10.1515/hsz-2018-0158] [Citation(s) in RCA: 46] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2018] [Accepted: 05/07/2018] [Indexed: 11/15/2022]
Abstract
Protein crystallization in living cells has been observed surprisingly often as a native assembly process during the past decades, and emerging evidence indicates that this phenomenon is also accessible for recombinant proteins. But only recently the advent of high-brilliance synchrotron sources, X-ray free-electron lasers, and improved serial data collection strategies has allowed the use of these micrometer-sized crystals for structural biology. Thus, in cellulo crystallization could offer exciting new possibilities for proteins that do not crystallize applying conventional approaches. In this review, we comprehensively summarize the current knowledge of intracellular protein crystallization. This includes an overview of the cellular functions, the physical properties, and, if known, the mode of regulation of native in cellulo crystal formation, complemented with a discussion of the reported crystallization events of recombinant proteins and the current method developments to successfully collect X-ray diffraction data from in cellulo crystals. Although the intracellular protein self-assembly mechanisms are still poorly understood, regulatory differences between native in cellulo crystallization linked to a specific function and accidently crystallizing proteins, either disease associated or recombinantly introduced, become evident. These insights are important to systematically exploit living cells as protein crystallization chambers in the future.
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Affiliation(s)
- Robert Schönherr
- Institute of Biochemistry, Center for Structural and Cell Biology in Medicine, University of Lübeck, Ratzeburger Allee 160, D-23562 Lübeck, Germany.,Deutsches Elektronen Synchrotron (DESY), Notkestrasse 85, D-22607 Hamburg, Germany
| | - Janine Mia Rudolph
- Institute of Biochemistry, Center for Structural and Cell Biology in Medicine, University of Lübeck, Ratzeburger Allee 160, D-23562 Lübeck, Germany.,Center for Free-Electron Laser Science (CFEL), DESY, Notkestrasse 85, D-22607 Hamburg, Germany
| | - Lars Redecke
- Institute of Biochemistry, Center for Structural and Cell Biology in Medicine, University of Lübeck, Ratzeburger Allee 160, D-23562 Lübeck, Germany.,Deutsches Elektronen Synchrotron (DESY), Notkestrasse 85, D-22607 Hamburg, Germany
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40
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Pakdel M, von Blume J. Exploring new routes for secretory protein export from the trans-Golgi network. Mol Biol Cell 2019; 29:235-240. [PMID: 29382805 PMCID: PMC5996961 DOI: 10.1091/mbc.e17-02-0117] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2017] [Revised: 11/30/2017] [Accepted: 12/01/2017] [Indexed: 12/21/2022] Open
Abstract
Sorting of soluble proteins for transport to intracellular compartments and for secretion from cells is essential for cell and tissue homeostasis. The trans-Golgi network (TGN) is a major sorting station that sorts secretory proteins into specific carriers to transport them to their final destinations. The sorting of lysosomal hydrolases at the TGN by the mannose 6-phosphate receptor is well understood. The recent discovery of a Ca2+-based sorting of secretory cargo at the TGN is beginning to uncover the mechanism by which cells sort secretory cargoes from Golgi residents and cargoes destined to the other cellular compartments. This Ca2+-based sorting involves the cytoplasmic actin cytoskeleton, which through membrane anchored Ca2+ ATPase SPCA1 and the luminal Ca2+ binding protein Cab45 sorts of a subset of secretory proteins at the TGN. We present this discovery and highlight important challenges that remain unaddressed in the overall pathway of cargo sorting at the TGN.
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Affiliation(s)
- Mehrshad Pakdel
- Max Planck Institute of Biochemistry, 82152 Martinsried, Germany
| | - Julia von Blume
- Max Planck Institute of Biochemistry, 82152 Martinsried, Germany
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Stevenson MJ, Uyeda KS, Harder NHO, Heffern MC. Metal-dependent hormone function: the emerging interdisciplinary field of metalloendocrinology. Metallomics 2019; 11:85-110. [PMID: 30270362 PMCID: PMC10249669 DOI: 10.1039/c8mt00221e] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2024]
Abstract
For over 100 years, there has been an incredible amount of knowledge amassed concerning hormones in the endocrine system and their central role in human health. Hormones represent a diverse group of biomolecules that are released by glands, communicate signals to their target tissue, and are regulated by feedback loops to maintain organism health. Many disease states, such as diabetes and reproductive disorders, stem from misregulation or dysfunction of hormones. Increasing research is illuminating the intricate roles of metal ions in the endocrine system where they may act advantageously in concert with hormones or deleteriously catalyze hormone-associated disease states. As the critical role of metal ions in the endocrine system becomes more apparent, it is increasingly important to untangle the complex mechanisms underlying the connections between inorganic biochemistry and hormone function to understand and control endocrinological phenomena. This tutorial review harmonizes the interdisciplinary fields of endocrinology and inorganic chemistry in the newly-termed field of "metalloendocrinology". We describe examples linking metals to both normal and aberrant hormone function with a focus on highlighting insight to molecular mechanisms. Hormone activities related to both essential metal micronutrients, such as copper, iron, zinc, and calcium, and disruptive nonessential metals, such as lead and cadmium are discussed.
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Affiliation(s)
- Michael J Stevenson
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, USA.
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42
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Protein Nanofibrils as Storage Forms of Peptide Drugs and Hormones. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2019; 1174:265-290. [PMID: 31713202 DOI: 10.1007/978-981-13-9791-2_8] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Amyloids are highly organized cross β-sheet protein nanofibrils that are associated with both diseases and functions. Thermodynamically amyloids are stable structures as they represent the lowest free energy state that proteins can attain. However, recent studies suggest that amyloid fibrils can be dissociated by a change in environmental parameters such as pH and ionic strength. This reversibility of amyloids can not only be associated with disease, but function as well. In disease-associated amyloids, fibrils can act as reservoirs of cytotoxic oligomers. Recently, in higher organisms such as mammals, hormones were found to be stored in amyloid-like state, where these were reported to act as a reservoir of functional monomers. These hormone amyloids can dissociate to monomers upon release from the secretory granules, and subsequently bind to their respective receptors and perform their functions. In this book chapter, we describe in detail how these protein nanofibrils represent the densest possible peptide packing and are suitable for long-term storage. Thus, mimicking the feature of amyloids to release functional monomers, it is possible to formulate amyloid-based peptide/protein drugs, which can be used for sustained release.
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43
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Dufurrena Q, Bäck N, Mains R, Hodgson L, Tanowitz H, Mandela P, Eipper B, Kuliawat R. Kalirin/Trio Rho GDP/GTP exchange factors regulate proinsulin and insulin secretion. J Mol Endocrinol 2018; 62:JME-18-0048.R2. [PMID: 30407917 PMCID: PMC6494717 DOI: 10.1530/jme-18-0048] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/15/2018] [Accepted: 11/05/2018] [Indexed: 12/31/2022]
Abstract
Key features for progression to pancreatic β-cell failure and disease are loss of glucose responsiveness and an increased ratio of secreted proinsulin to insulin. Proinsulin and insulin are stored in secretory granules (SGs) and the fine-tuning of hormone output requires signal mediated recruitment of select SG populations according to intracellular location and age. The GTPase Rac1 coordinates multiple signaling pathways that specify SG release and Rac1 activity is controlled in part by GDP/GTP exchange factors (GEFs). To explore the function of two large multidomain GEFs, Kalirin and Trio in β-cells, we manipulated their Rac1-specific GEF1 domain activity by using small molecule inhibitors and by genetically ablating Kalirin. We examined age related secretory granule behavior employing radiolabeling protocols. Loss of Kalirin/Trio function attenuated radioactive proinsulin release by reducing constitutive-like secretion and exocytosis of 2-hour old granules. At later chase times or at steady state, Kalirin/Trio manipulations decreased glucose stimulated insulin output. Finally, use of a Rac1 FRET biosensor with cultured β-cell lines, demonstrated that Kalirin/Trio GEF1 activity was required for normal rearrangement of Rac1 to the plasma membrane in response to glucose. Rac1 activation can be evoked by both glucose metabolism and signaling through the incretin glucagon-like peptide 1 (GLP-1) receptor. GLP-1 addition restored Rac1 localization/activity and insulin secretion in the absence of Kalirin, thereby assigning Kalirin's participation to stimulatory glucose signaling.
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Affiliation(s)
- Quinn Dufurrena
- Department of Medicine, Stony Brook University School of Medicine, Stony Brook, NY
| | - Nils Bäck
- Department of Anatomy, Faculty of Medicine, University of Helsinki, Helsinki, Finland
| | - Richard Mains
- Department of Neuroscience, University of Connecticut Health Center, Farmington, CT
| | - Louis Hodgson
- Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY
| | - Herbert Tanowitz
- Departments of Pathology, Medicine, Albert Einstein College of Medicine, Bronx, NY
| | | | - Betty Eipper
- Department of Molecular Biology and Biophysics, University of Connecticut Health Center, Farmington, CT
| | - Regina Kuliawat
- Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, NY
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44
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Venugopal K, Marion S. Secretory organelle trafficking in Toxoplasma gondii: A long story for a short travel. Int J Med Microbiol 2018; 308:751-760. [DOI: 10.1016/j.ijmm.2018.07.007] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2018] [Revised: 07/10/2018] [Accepted: 07/15/2018] [Indexed: 12/15/2022] Open
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45
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Liu M, Weiss MA, Arunagiri A, Yong J, Rege N, Sun J, Haataja L, Kaufman RJ, Arvan P. Biosynthesis, structure, and folding of the insulin precursor protein. Diabetes Obes Metab 2018; 20 Suppl 2:28-50. [PMID: 30230185 PMCID: PMC6463291 DOI: 10.1111/dom.13378] [Citation(s) in RCA: 140] [Impact Index Per Article: 20.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/25/2018] [Revised: 05/04/2018] [Accepted: 05/23/2018] [Indexed: 02/06/2023]
Abstract
Insulin synthesis in pancreatic β-cells is initiated as preproinsulin. Prevailing glucose concentrations, which oscillate pre- and postprandially, exert major dynamic variation in preproinsulin biosynthesis. Accompanying upregulated translation of the insulin precursor includes elements of the endoplasmic reticulum (ER) translocation apparatus linked to successful orientation of the signal peptide, translocation and signal peptide cleavage of preproinsulin-all of which are necessary to initiate the pathway of proper proinsulin folding. Evolutionary pressures on the primary structure of proinsulin itself have preserved the efficiency of folding ("foldability"), and remarkably, these evolutionary pressures are distinct from those protecting the ultimate biological activity of insulin. Proinsulin foldability is manifest in the ER, in which the local environment is designed to assist in the overall load of proinsulin folding and to favour its disulphide bond formation (while limiting misfolding), all of which is closely tuned to ER stress response pathways that have complex (beneficial, as well as potentially damaging) effects on pancreatic β-cells. Proinsulin misfolding may occur as a consequence of exuberant proinsulin biosynthetic load in the ER, proinsulin coding sequence mutations, or genetic predispositions that lead to an altered ER folding environment. Proinsulin misfolding is a phenotype that is very much linked to deficient insulin production and diabetes, as is seen in a variety of contexts: rodent models bearing proinsulin-misfolding mutants, human patients with Mutant INS-gene-induced Diabetes of Youth (MIDY), animal models and human patients bearing mutations in critical ER resident proteins, and, quite possibly, in more common variety type 2 diabetes.
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Affiliation(s)
- Ming Liu
- Department of Endocrinology and Metabolism, Tianjin Medical University General Hospital, Tianjin, China 300052
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical School, Ann Arbor 48105 MI USA
| | - Michael A. Weiss
- Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis 46202 IN USA
- Department of Biochemistry, Case-Western Reserve University, Cleveland 44016 OH USA
| | - Anoop Arunagiri
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical School, Ann Arbor 48105 MI USA
| | - Jing Yong
- Degenerative Diseases Program, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA 92307 USA
| | - Nischay Rege
- Department of Biochemistry, Case-Western Reserve University, Cleveland 44016 OH USA
| | - Jinhong Sun
- Department of Endocrinology and Metabolism, Tianjin Medical University General Hospital, Tianjin, China 300052
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical School, Ann Arbor 48105 MI USA
| | - Leena Haataja
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical School, Ann Arbor 48105 MI USA
| | - Randal J. Kaufman
- Degenerative Diseases Program, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA 92307 USA
| | - Peter Arvan
- Division of Metabolism, Endocrinology & Diabetes, University of Michigan Medical School, Ann Arbor 48105 MI USA
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Li M, Du W, Zhou M, Zheng L, Song E, Hou J. Proteomic analysis of insulin secretory granules in INS-1 cells by protein correlation profiling. BIOPHYSICS REPORTS 2018; 4:329-338. [PMID: 30596141 PMCID: PMC6276070 DOI: 10.1007/s41048-018-0061-3] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2018] [Accepted: 07/08/2018] [Indexed: 11/30/2022] Open
Abstract
Abstract Insulin secretory granules (ISGs), a group of distinguishing organelles in pancreatic β cells, are responsible for the storage and secretion of insulin to maintain blood glucose homeostasis. The molecular mechanisms of ISG biogenesis, maturation, transportation, and exocytosis are still largely unknown because the proteins involved in these distinct steps have not been fully identified. Subcellular fractionation by density gradient centrifugation has been successfully employed to analyze the proteomes of numerous organelles. However, use of this method to elucidate the ISG proteome is limited by co-fractionated contaminants because ISGs are very dynamic and have abundant exchanges or contacts with other organelles, such as the Golgi apparatus, lysosomes, and endosomes. In this study, we developed a new strategy for identifying ISG proteins by protein correlation profiling (PCP)-based proteomics, which included ISG purification by OptiPrep density gradient centrifugation, label-free quantitative proteome, and identification of ISG proteins by correlating fractionation profiles between candidates and known ISG markers. Using this approach, we were able to identify 81 ISG proteins. Among them, TM9SF3, a nine-transmembrane protein, was considered a high confidence ISG candidate protein highlighted in the PCP network. Further biochemical and immunofluorescence assays indicated that TM9SF3 localized in ISGs, suggesting that it is a potential new ISG marker. Graphical abstract ![]()
Electronic supplementary material The online version of this article (10.1007/s41048-018-0061-3) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Min Li
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China.,2College of Life Sciences, University of Chinese Academy of Sciences, Beijing, 100049 China
| | - Wen Du
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China
| | - Maoge Zhou
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China
| | - Li Zheng
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China
| | - Eli Song
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China
| | - Junjie Hou
- 1National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, 100101 China
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Kostenko S, Heu CC, Yaron JR, Singh G, de Oliveira C, Muller WJ, Singh VP. c-Src regulates cargo transit via the Golgi in pancreatic acinar cells. Sci Rep 2018; 8:11903. [PMID: 30093675 PMCID: PMC6085363 DOI: 10.1038/s41598-018-30370-4] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2018] [Accepted: 07/17/2018] [Indexed: 12/11/2022] Open
Abstract
The exocrine pancreatic acinar cell is unique for its rapid protein synthesis and packaging in zymogen granules (ZGs). However, while crucial to the pathogenesis of pancreatitis, the signaling involved in the transit of proteins via the Golgi is poorly understood in these cells. Noting the evidence of c-Src in regulating transit of cargo via the Golgi in other systems, we explored this in acinar cells. Stimulation of ZG formation with dexamethasone activated Src and increased the Golgi area in acinar cells. c-Src localized to the microsomes of acinar cells on immunofluorescence and subcellular fractionation. While other Src family members had no effect on the Golgi markers P115 and GM130, active c-Src increased the Golgi area these stained, extending them into the ER. Src inhibition reduced amylase staining outside the Golgi and increased it in a stack like Golgi morphology. In vivo pharmacologic inhibition or acinar specific genetic deletion of c-Src reduced ZG number and staining of amylase in ZGs along with increasing amylase retention in the microsomal fraction. Morphologically this was associated with smaller Golgi stacks, and dilation of the endoplasmic reticulum. Therefore the role c-Src regulated Golgi function, ZG formation and microsomal zymogen transit in acinar cells needs to be explored in pancreatitis.
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Affiliation(s)
- Sergiy Kostenko
- Department of Medicine, Mayo Clinic, Scottsdale, Arizona, USA
| | - Chan C Heu
- Department of Medicine, Mayo Clinic, Scottsdale, Arizona, USA
| | - Jordan R Yaron
- Department of Medicine, Mayo Clinic, Scottsdale, Arizona, USA
| | - Garima Singh
- Department of Medicine, Mayo Clinic, Scottsdale, Arizona, USA
| | | | - William J Muller
- Goodman Cancer Research Center and Department of Biology, McGill University, Montreal, QC, H3A 1A3, Canada
| | - Vijay P Singh
- Department of Medicine, Mayo Clinic, Scottsdale, Arizona, USA.
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Jurek B, Neumann ID. The Oxytocin Receptor: From Intracellular Signaling to Behavior. Physiol Rev 2018; 98:1805-1908. [DOI: 10.1152/physrev.00031.2017] [Citation(s) in RCA: 601] [Impact Index Per Article: 85.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
The many facets of the oxytocin (OXT) system of the brain and periphery elicited nearly 25,000 publications since 1930 (see FIGURE 1 , as listed in PubMed), which revealed central roles for OXT and its receptor (OXTR) in reproduction, and social and emotional behaviors in animal and human studies focusing on mental and physical health and disease. In this review, we discuss the mechanisms of OXT expression and release, expression and binding of the OXTR in brain and periphery, OXTR-coupled signaling cascades, and their involvement in behavioral outcomes to assemble a comprehensive picture of the central and peripheral OXT system. Traditionally known for its role in milk let-down and uterine contraction during labor, OXT also has implications in physiological, and also behavioral, aspects of reproduction, such as sexual and maternal behaviors and pair bonding, but also anxiety, trust, sociability, food intake, or even drug abuse. The many facets of OXT are, on a molecular basis, brought about by a single receptor. The OXTR, a 7-transmembrane G protein-coupled receptor capable of binding to either Gαior Gαqproteins, activates a set of signaling cascades, such as the MAPK, PKC, PLC, or CaMK pathways, which converge on transcription factors like CREB or MEF-2. The cellular response to OXT includes regulation of neurite outgrowth, cellular viability, and increased survival. OXTergic projections in the brain represent anxiety and stress-regulating circuits connecting the paraventricular nucleus of the hypothalamus, amygdala, bed nucleus of the stria terminalis, or the medial prefrontal cortex. Which OXT-induced patterns finally alter the behavior of an animal or a human being is still poorly understood, and studying those OXTR-coupled signaling cascades is one initial step toward a better understanding of the molecular background of those behavioral effects.
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Affiliation(s)
- Benjamin Jurek
- Department of Behavioural and Molecular Neurobiology, Institute of Zoology, University of Regensburg, Regensburg, Germany
| | - Inga D. Neumann
- Department of Behavioural and Molecular Neurobiology, Institute of Zoology, University of Regensburg, Regensburg, Germany
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Genetic dissection of neuropeptide cell biology at high and low activity in a defined sensory neuron. Proc Natl Acad Sci U S A 2018; 115:E6890-E6899. [PMID: 29959203 PMCID: PMC6055185 DOI: 10.1073/pnas.1714610115] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Neuropeptides are ubiquitous modulators of behavior and physiology. They are packaged in specialized secretory organelles called dense core vesicles (DCVs) that are released upon neural stimulation. Whereas local recycling of synaptic vesicles has been investigated intensively, there are few studies on recycling of DCV proteins. We set up a paradigm to study DCVs in a neuron whose activity we can control. We validate our model by confirming many previous observations on DCV cell biology. We identify a set of genes involved in recycling of DCV proteins. We also find evidence that different mechanisms of DCV priming and exocytosis may operate at high and low neural activity. Neuropeptides are ubiquitous modulators of behavior and physiology. They are packaged in specialized secretory organelles called dense core vesicles (DCVs) that are released upon neural stimulation. Unlike synaptic vesicles, which can be recycled and refilled close to release sites, DCVs must be replenished by de novo synthesis in the cell body. Here, we dissect DCV cell biology in vivo in a Caenorhabditis elegans sensory neuron whose tonic activity we can control using a natural stimulus. We express fluorescently tagged neuropeptides in the neuron and define parameters that describe their subcellular distribution. We measure these parameters at high and low neural activity in 187 mutants defective in proteins implicated in membrane traffic, neuroendocrine secretion, and neuronal or synaptic activity. Using unsupervised hierarchical clustering methods, we analyze these data and identify 62 groups of genes with similar mutant phenotypes. We explore the function of a subset of these groups. We recapitulate many previous findings, validating our paradigm. We uncover a large battery of proteins involved in recycling DCV membrane proteins, something hitherto poorly explored. We show that the unfolded protein response promotes DCV production, which may contribute to intertissue communication of stress. We also find evidence that different mechanisms of priming and exocytosis may operate at high and low neural activity. Our work provides a defined framework to study DCV biology at different neural activity levels.
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ER Lipid Defects in Neuropeptidergic Neurons Impair Sleep Patterns in Parkinson's Disease. Neuron 2018; 98:1155-1169.e6. [PMID: 29887339 DOI: 10.1016/j.neuron.2018.05.022] [Citation(s) in RCA: 74] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2017] [Revised: 04/16/2018] [Accepted: 05/14/2018] [Indexed: 01/23/2023]
Abstract
Parkinson's disease patients report disturbed sleep patterns long before motor dysfunction. Here, in parkin and pink1 models, we identify circadian rhythm and sleep pattern defects and map these to specific neuropeptidergic neurons in fly models and in hypothalamic neurons differentiated from patient induced pluripotent stem cells (iPSCs). Parkin and Pink1 control the clearance of mitochondria by protein ubiquitination. Although we do not observe major defects in mitochondria of mutant neuropeptidergic neurons, we do find an excess of endoplasmic reticulum-mitochondrial contacts. These excessive contact sites cause abnormal lipid trafficking that depletes phosphatidylserine from the endoplasmic reticulum (ER) and disrupts the production of neuropeptide-containing vesicles. Feeding mutant animals phosphatidylserine rescues neuropeptidergic vesicle production and acutely restores normal sleep patterns in mutant animals. Hence, sleep patterns and circadian disturbances in Parkinson's disease models are explained by excessive ER-mitochondrial contacts, and blocking their formation or increasing phosphatidylserine levels rescues the defects in vivo.
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