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Zhao M, Cao N, Gu H, Xu J, Xu W, Zhang D, Wei TYW, Wang K, Guo R, Cui H, Wang X, Guo X, Li Z, He K, Li Z, Zhang Y, Shyy JYJ, Dong E, Xiao H. AMPK Attenuation of β-Adrenergic Receptor-Induced Cardiac Injury via Phosphorylation of β-Arrestin-1-ser330. Circ Res 2024; 135:651-667. [PMID: 39082138 DOI: 10.1161/circresaha.124.324762] [Citation(s) in RCA: 5] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/21/2024] [Revised: 07/16/2024] [Accepted: 07/22/2024] [Indexed: 09/01/2024]
Abstract
BACKGROUND β-adrenergic receptor (β-AR) overactivation is a major pathological cue associated with cardiac injury and diseases. AMPK (AMP-activated protein kinase), a conserved energy sensor, regulates energy metabolism and is cardioprotective. However, whether AMPK exerts cardioprotective effects via regulating the signaling pathway downstream of β-AR remains unclear. METHODS Using immunoprecipitation, mass spectrometry, site-specific mutation, in vitro kinase assay, and in vivo animal studies, we determined whether AMPK phosphorylates β-arrestin-1 at serine (Ser) 330. Wild-type mice and mice with site-specific mutagenesis (S330A knock-in [KI]/S330D KI) were subcutaneously injected with the β-AR agonist isoproterenol (5 mg/kg) to evaluate the causality between β-adrenergic insult and β-arrestin-1 Ser330 phosphorylation. Cardiac transcriptomics was used to identify changes in gene expression from β-arrestin-1-S330A/S330D mutation and β-adrenergic insult. RESULTS Metformin could decrease cAMP/PKA (protein kinase A) signaling induced by isoproterenol. AMPK bound to β-arrestin-1 and phosphorylated Ser330 with the highest phosphorylated mass spectrometry score. AMPK activation promoted β-arrestin-1 Ser330 phosphorylation in vitro and in vivo. Neonatal mouse cardiomyocytes overexpressing β-arrestin-1-S330D (active form) inhibited the β-AR/cAMP/PKA axis by increasing PDE (phosphodiesterase) 4 expression and activity. Cardiac transcriptomics revealed that the differentially expressed genes between isoproterenol-treated S330A KI and S330D KI mice were mainly involved in immune processes and inflammatory response. β-arrestin-1 Ser330 phosphorylation inhibited isoproterenol-induced reactive oxygen species production and NLRP3 (NOD-like receptor protein 3) inflammasome activation in neonatal mouse cardiomyocytes. In S330D KI mice, the β-AR-activated cAMP/PKA pathways were attenuated, leading to repressed inflammasome activation, reduced expression of proinflammatory cytokines, and mitigated macrophage infiltration. Compared with S330A KI mice, S330D KI mice showed diminished cardiac fibrosis and improved cardiac function upon isoproterenol exposure. However, the cardiac protection exerted by AMPK was abolished in S330A KI mice. CONCLUSIONS AMPK phosphorylation of β-arrestin-1 Ser330 potentiated PDE4 expression and activity, thereby inhibiting β-AR/cAMP/PKA activation. Subsequently, β-arrestin-1 Ser330 phosphorylation blocks β-AR-induced cardiac inflammasome activation and remodeling.
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Affiliation(s)
- Mingming Zhao
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Ning Cao
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Department of Cardiology, Cardiovascular Center, Beijing Friendship Hospital (N.C.), Capital Medical University, Beijing, China
| | - Huijun Gu
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Jiachao Xu
- Laboratory for Clinical Medicine (N.C.), Capital Medical University, Beijing, China
| | - Wenli Xu
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Center for Cardiopulmonary Rehabilitation, University of Health and Rehabilitation Sciences Qingdao Hospital (Qingdao Municipal Hospital), School of Health and Life Sciences, University of Health and Rehabilitation Sciences, Qingdao, China (W.X., E.D., H.X.)
| | - Di Zhang
- Center for Quantitative Biology, Academy for Advanced Interdisciplinary Studies (D.Z., Zhiyuan Li), Peking University, Beijing, China
| | - Tong-You Wade Wei
- Division of Cardiology, Department of Medicine, University of California, San Diego, La Jolla (T.-Y.W.W., J.Y.-J.S.)
| | - Kang Wang
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Ruiping Guo
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Hongtu Cui
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Xiaofeng Wang
- Laboratory of Inflammation and Vaccines, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China (X.W.)
| | - Xin Guo
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Zhiyuan Li
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- Center for Quantitative Biology, Academy for Advanced Interdisciplinary Studies (D.Z., Zhiyuan Li), Peking University, Beijing, China
| | - Kangmin He
- State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China (J.X., K.H.)
- University of Chinese Academy of Sciences, Beijing, China (K.H.)
| | - Zijian Li
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - Youyi Zhang
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
| | - John Y-J Shyy
- Division of Cardiology, Department of Medicine, University of California, San Diego, La Jolla (T.-Y.W.W., J.Y.-J.S.)
| | - Erdan Dong
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- Institute of Cardiovascular Sciences (E.D.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Center for Cardiopulmonary Rehabilitation, University of Health and Rehabilitation Sciences Qingdao Hospital (Qingdao Municipal Hospital), School of Health and Life Sciences, University of Health and Rehabilitation Sciences, Qingdao, China (W.X., E.D., H.X.)
| | - Han Xiao
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- State Key Laboratory of Vascular Homeostasis and Remodeling, Institute of Advanced Clinical Medicine (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.), Peking University, Beijing, China
- National Health Commission (NHC) Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Beijing Key Laboratory of Cardiovascular Receptors Research, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Unit of Medical Science Research Management/Basic and Clinical Research of Metabolic Cardiovascular Diseases, Chinese Academy of Medical Sciences, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Haihe Laboratory of Cell Ecosystem, Beijing, China (M.Z., N.C., H.G., W.X., K.W., R.G., H.C., X.G., Zijian Li, Y.Z., E.D., H.X.)
- Research Center for Cardiopulmonary Rehabilitation, University of Health and Rehabilitation Sciences Qingdao Hospital (Qingdao Municipal Hospital), School of Health and Life Sciences, University of Health and Rehabilitation Sciences, Qingdao, China (W.X., E.D., H.X.)
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Shpakov AO. Hormonal and Allosteric Regulation of the Luteinizing Hormone/Chorionic Gonadotropin Receptor. FRONT BIOSCI-LANDMRK 2024; 29:313. [PMID: 39344322 DOI: 10.31083/j.fbl2909313] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2024] [Revised: 06/20/2024] [Accepted: 07/10/2024] [Indexed: 10/01/2024]
Abstract
Luteinizing hormone (LH) and human chorionic gonadotropin (CG), like follicle-stimulating hormone, are the most important regulators of the reproductive system. They exert their effect on the cell through the LH/CG receptor (LHCGR), which belongs to the family of G protein-coupled receptors. Binding to gonadotropin induces the interaction of LHCGR with various types of heterotrimeric G proteins (Gs, Gq/11, Gi) and β-arrestins, which leads to stimulation (Gs) or inhibition (Gi) of cyclic adenosine monophosphate-dependent cascades, activation of the phospholipase pathway (Gq/11), and also to the formation of signalosomes that mediate the stimulation of mitogen-activated protein kinases (β-arrestins). The efficiency and selectivity of activation of intracellular cascades by different gonadotropins varies, which is due to differences in their interaction with the ligand-binding site of LHCGR. Gonadotropin signaling largely depends on the status of N- and O-glycosylation of LH and CG, on the formation of homo- and heterodimeric receptor complexes, on the cell-specific microenvironment of LHCGR and the presence of autoantibodies to it, and allosteric mechanisms are important in the implementation of these influences, which is due to the multiplicity of allosteric sites in different loci of the LHCGR. The development of low-molecular-weight allosteric regulators of LHCGR with different profiles of pharmacological activity, which can be used in medicine for the correction of reproductive disorders and in assisted reproductive technologies, is promising. These and other issues regarding the hormonal and allosteric regulation of LHCGR are summarized and discussed in this review.
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Affiliation(s)
- Alexander O Shpakov
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 194223 St. Petersburg, Russia
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Strauss A, Gonzalez-Hernandez AJ, Lee J, Abreu N, Selvakumar P, Salas-Estrada L, Kristt M, Arefin A, Huynh K, Marx DC, Gilliland K, Melancon BJ, Filizola M, Meyerson J, Levitz J. Structural basis of positive allosteric modulation of metabotropic glutamate receptor activation and internalization. Nat Commun 2024; 15:6498. [PMID: 39090128 PMCID: PMC11294631 DOI: 10.1038/s41467-024-50548-x] [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: 03/21/2024] [Accepted: 07/12/2024] [Indexed: 08/04/2024] Open
Abstract
The metabotropic glutamate receptors (mGluRs) are neuromodulatory family C G protein coupled receptors which assemble as dimers and allosterically couple extracellular ligand binding domains (LBDs) to transmembrane domains (TMDs) to drive intracellular signaling. Pharmacologically, mGluRs can be targeted at the LBDs by glutamate and synthetic orthosteric compounds or at the TMDs by allosteric modulators. Despite the potential of allosteric compounds as therapeutics, an understanding of the functional and structural basis of their effects is limited. Here we use multiple approaches to dissect the functional and structural effects of orthosteric versus allosteric ligands. We find, using electrophysiological and live cell imaging assays, that both agonists and positive allosteric modulators (PAMs) can drive activation and internalization of group II and III mGluRs. The effects of PAMs are pleiotropic, boosting the maximal response to orthosteric agonists and serving independently as internalization-biased agonists across mGluR subtypes. Motivated by this and intersubunit FRET analyses, we determine cryo-electron microscopy structures of mGluR3 in the presence of either an agonist or antagonist alone or in combination with a PAM. These structures reveal PAM-driven re-shaping of intra- and inter-subunit conformations and provide evidence for a rolling TMD dimer interface activation pathway that controls G protein and beta-arrestin coupling.
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Affiliation(s)
- Alexa Strauss
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
- Tri-Institutional Program in Chemical Biology, New York, NY, 10065, USA
| | | | - Joon Lee
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Nohely Abreu
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Purushotham Selvakumar
- Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Leslie Salas-Estrada
- Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, 10029, USA
| | - Melanie Kristt
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Anisul Arefin
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Kevin Huynh
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Dagan C Marx
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Kristen Gilliland
- Warren Center for Neuroscience Drug Discovery at Vanderbilt University, Vanderbilt University, Nashville, TN, 37232, USA
| | - Bruce J Melancon
- Warren Center for Neuroscience Drug Discovery at Vanderbilt University, Vanderbilt University, Nashville, TN, 37232, USA
- Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN, 37232, USA
| | - Marta Filizola
- Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, 10029, USA
| | - Joel Meyerson
- Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY, 10065, USA
| | - Joshua Levitz
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, 10065, USA.
- Tri-Institutional Program in Chemical Biology, New York, NY, 10065, USA.
- Department of Psychiatry, Weill Cornell Medicine, New York, NY, 10065, USA.
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4
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Savransky S, White AD, Vilardaga JP. Deciphering the role of glycosaminoglycans in GPCR signaling. Cell Signal 2024; 118:111149. [PMID: 38522808 PMCID: PMC10999332 DOI: 10.1016/j.cellsig.2024.111149] [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: 01/28/2024] [Revised: 03/11/2024] [Accepted: 03/21/2024] [Indexed: 03/26/2024]
Abstract
G protein-coupled receptors (GPCR) and glycosaminoglycans (GAGs) are two essential components of the cell surface that regulate physiological processes in the body. GPCRs are the most extensive family of transmembrane receptors that control cellular responses to extracellular stimuli, while GAGs are polysaccharides that contribute to the function of the extracellular matrix (ECM). Due to their proximity to the plasma membrane, GAGs participate in signal transduction by interacting with various extracellular molecules and cell surface receptors. GAGs can directly interact with certain GPCRs or their ligands (chemokines, peptide hormones and neuropeptides, structural proteins, and enzymes) from the glutamate receptor family, the rhodopsin receptor family, the adhesion receptor family, and the secretin receptor family. These interactions have recently become an emerging topic, providing a new avenue for understanding how GPCR signaling is regulated. This review discusses our current state of knowledge about the role of GAGs in GPCR signaling and function.
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Affiliation(s)
- Sofya Savransky
- Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA; Graduate Program in Molecular Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
| | - Alex D White
- Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA; Graduate Program in Molecular Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Jean-Pierre Vilardaga
- Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
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5
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Reja SI, Minoshima M, Hori Y, Kikuchi K. Recent advancements of fluorescent biosensors using semisynthetic probes. Biosens Bioelectron 2024; 247:115862. [PMID: 38147718 DOI: 10.1016/j.bios.2023.115862] [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: 07/15/2022] [Revised: 11/07/2023] [Accepted: 11/17/2023] [Indexed: 12/28/2023]
Abstract
Fluorescent biosensors are crucial experimental tools for live-cell imaging and the quantification of different biological analytes. Fluorescent protein (FP)-based biosensors are widely used for imaging applications in living systems. However, the use of FP-based biosensors is hindered by their large size, poor photostability, and laborious genetic manipulations required to improve their properties. Recently, semisynthetic fluorescent biosensors have been developed to address the limitations of FP-based biosensors using chemically modified fluorescent probes and self-labeling protein tag/peptide tags or DNA/RNA-based hybrid systems. Semisynthetic biosensors have unique advantages, as they can be easily modified using different probes. Moreover, the self-labeling protein tag, which labels synthetically developed ligands via covalent bonds, has immense potential for biosensor development. This review discusses the recent progress in different types of fluorescent biosensors for metabolites, protein aggregation and degradation, DNA methylation, endocytosis and exocytosis, membrane tension, and cellular viscosity. Here, we explain in detail the design strategy and working principle of these biosensors. The information presented will help the reader to create new biosensors using self-labeling protein tags for various applications.
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Affiliation(s)
- Shahi Imam Reja
- Immunology Frontier Research Center (IFReC), Osaka University, Suita, Osaka, 565-0871, Japan
| | - Masafumi Minoshima
- Division of Applied Chemistry, Graduate School of Engineering, Osaka University, Suita, Osaka, 565-0871, Japan
| | - Yuichiro Hori
- Department of Chemistry, Faculty of Science, Kyushu University, Fukuoka, 819-0395, Japan
| | - Kazuya Kikuchi
- Immunology Frontier Research Center (IFReC), Osaka University, Suita, Osaka, 565-0871, Japan; Division of Applied Chemistry, Graduate School of Engineering, Osaka University, Suita, Osaka, 565-0871, Japan.
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6
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Kee TR, Khan SA, Neidhart MB, Masters BM, Zhao VK, Kim YK, McGill Percy KC, Woo JAA. The multifaceted functions of β-arrestins and their therapeutic potential in neurodegenerative diseases. Exp Mol Med 2024; 56:129-141. [PMID: 38212557 PMCID: PMC10834518 DOI: 10.1038/s12276-023-01144-4] [Citation(s) in RCA: 14] [Impact Index Per Article: 14.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2023] [Revised: 10/18/2023] [Accepted: 10/20/2023] [Indexed: 01/13/2024] Open
Abstract
Arrestins are multifunctional proteins that regulate G-protein-coupled receptor (GPCR) desensitization, signaling, and internalization. The arrestin family consists of four subtypes: visual arrestin1, β-arrestin1, β-arrestin2, and visual arrestin-4. Recent studies have revealed the multifunctional roles of β-arrestins beyond GPCR signaling, including scaffolding and adapter functions, and physically interacting with non-GPCR receptors. Increasing evidence suggests that β-arrestins are involved in the pathogenesis of a variety of neurodegenerative diseases, including Alzheimer's disease (AD), frontotemporal dementia (FTD), and Parkinson's disease (PD). β-arrestins physically interact with γ-secretase, leading to increased production and accumulation of amyloid-beta in AD. Furthermore, β-arrestin oligomers inhibit the autophagy cargo receptor p62/SQSTM1, resulting in tau accumulation and aggregation in FTD. In PD, β-arrestins are upregulated in postmortem brain tissue and an MPTP model, and the β2AR regulates SNCA gene expression. In this review, we aim to provide an overview of β-arrestin1 and β-arrestin2, and describe their physiological functions and roles in neurodegenerative diseases. The multifaceted roles of β-arrestins and their involvement in neurodegenerative diseases suggest that they may serve as promising therapeutic targets.
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Affiliation(s)
- Teresa R Kee
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
- Department of Molecular Medicine, USF Health College of Medicine, Tampa, FL, 33613, USA
| | - Sophia A Khan
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
| | - Maya B Neidhart
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
| | - Brianna M Masters
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
| | - Victoria K Zhao
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
| | - Yenna K Kim
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA
| | | | - Jung-A A Woo
- Department of Pathology, CWRU School of Medicine, Cleveland, OH, 44106, USA.
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7
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Lee J, Gonzalez-Hernandez AJ, Kristt M, Abreu N, Roßmann K, Arefin A, Marx DC, Broichhagen J, Levitz J. Distinct beta-arrestin coupling and intracellular trafficking of metabotropic glutamate receptor homo- and heterodimers. SCIENCE ADVANCES 2023; 9:eadi8076. [PMID: 38055809 PMCID: PMC10699790 DOI: 10.1126/sciadv.adi8076] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/18/2023] [Accepted: 11/03/2023] [Indexed: 12/08/2023]
Abstract
The metabotropic glutamate receptors (mGluRs) are family C, dimeric G protein-coupled receptors (GPCRs), which play critical roles in synaptic transmission. Despite an increasing appreciation of the molecular diversity of this family, how distinct mGluR subtypes are regulated remains poorly understood. We reveal that different group II/III mGluR subtypes show markedly different beta-arrestin (β-arr) coupling and endocytic trafficking. While mGluR2 is resistant to internalization and mGluR3 shows transient β-arr coupling, which enables endocytosis and recycling, mGluR8 and β-arr form stable complexes, which leads to efficient lysosomal targeting and degradation. Using chimeras and mutagenesis, we pinpoint carboxyl-terminal domain regions that control β-arr coupling and trafficking, including the identification of an mGluR8 splice variant with impaired internalization. We then use a battery of high-resolution fluorescence assays to find that heterodimerization further expands the diversity of mGluR regulation. Together, this work provides insight into the relationship between GPCR/β-arr complex formation and trafficking while revealing diversity and intricacy in the regulation of mGluRs.
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Affiliation(s)
- Joon Lee
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
| | | | - Melanie Kristt
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
| | - Nohely Abreu
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
| | - Kilian Roßmann
- Leibniz-Forschungsinstitut für Molekulare Pharmakologie, 13125 Berlin, Germany
| | - Anisul Arefin
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
| | - Dagan C. Marx
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
| | | | - Joshua Levitz
- Department of Biochemistry, Weill Cornell Medicine, New York, NY 10065, USA
- Department of Psychiatry, Weill Cornell Medicine, New York, NY 10065, USA
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8
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Strauss A, Gonzalez-Hernandez AJ, Lee J, Abreu N, Selvakumar P, Salas-Estrada L, Kristt M, Marx DC, Gilliland K, Melancon BJ, Filizola M, Meyerson J, Levitz J. Structural basis of allosteric modulation of metabotropic glutamate receptor activation and desensitization. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.08.13.552748. [PMID: 37645747 PMCID: PMC10461995 DOI: 10.1101/2023.08.13.552748] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/31/2023]
Abstract
The metabotropic glutamate receptors (mGluRs) are neuromodulatory family C G protein coupled receptors which assemble as dimers and allosterically couple extracellular ligand binding domains (LBDs) to transmembrane domains (TMDs) to drive intracellular signaling. Pharmacologically, mGluRs can be targeted either at the LBDs by glutamate and synthetic orthosteric compounds or at the TMDs by allosteric modulators. Despite the potential of allosteric TMD-targeting compounds as therapeutics, an understanding of the functional and structural basis of their effects on mGluRs is limited. Here we use a battery of approaches to dissect the distinct functional and structural effects of orthosteric versus allosteric ligands. We find using electrophysiological and live cell imaging assays that both agonists and positive allosteric modulators (PAMs) can drive activation and desensitization of mGluRs. The effects of PAMs are pleiotropic, including both the ability to boost the maximal response to orthosteric agonists and to serve independently as desensitization-biased agonists across mGluR subtypes. Conformational sensors reveal PAM-driven inter-subunit re-arrangements at both the LBD and TMD. Motivated by this, we determine cryo-electron microscopy structures of mGluR3 in the presence of either an agonist or antagonist alone or in combination with a PAM. These structures reveal PAM-driven re-shaping of intra- and inter-subunit conformations and provide evidence for a rolling TMD dimer interface activation pathway that controls G protein and beta-arrestin coupling. Highlights -Agonists and PAMs drive mGluR activation, desensitization, and endocytosis-PAMs are desensitization-biased and synergistic with agonists-Four combinatorial ligand conditions reveal an ensemble of full-length mGluR structures with novel interfaces-Activation and desensitization involve rolling TMD interfaces which are re-shaped by PAM.
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9
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Vilardaga JP, Clark LJ, White AD, Sutkeviciute I, Lee JY, Bahar I. Molecular Mechanisms of PTH/PTHrP Class B GPCR Signaling and Pharmacological Implications. Endocr Rev 2023; 44:474-491. [PMID: 36503956 PMCID: PMC10461325 DOI: 10.1210/endrev/bnac032] [Citation(s) in RCA: 14] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/13/2022] [Revised: 11/14/2022] [Accepted: 12/05/2022] [Indexed: 12/14/2022]
Abstract
The classical paradigm of G protein-coupled receptor (GPCR) signaling via G proteins is grounded in a view that downstream responses are relatively transient and confined to the cell surface, but this notion has been revised in recent years following the identification of several receptors that engage in sustained signaling responses from subcellular compartments following internalization of the ligand-receptor complex. This phenomenon was initially discovered for the parathyroid hormone (PTH) type 1 receptor (PTH1R), a vital GPCR for maintaining normal calcium and phosphate levels in the body with the paradoxical ability to build or break down bone in response to PTH binding. The diverse biological processes regulated by this receptor are thought to depend on its capacity to mediate diverse modes of cyclic adenosine monophosphate (cAMP) signaling. These include transient signaling at the plasma membrane and sustained signaling from internalized PTH1R within early endosomes mediated by PTH. Here we discuss recent structural, cell signaling, and in vivo studies that unveil potential pharmacological outputs of the spatial versus temporal dimension of PTH1R signaling via cAMP. Notably, the combination of molecular dynamics simulations and elastic network model-based methods revealed how precise modulation of PTH signaling responses is achieved through structure-encoded allosteric coupling within the receptor and between the peptide hormone binding site and the G protein coupling interface. The implications of recent findings are now being explored for addressing key questions on how location bias in receptor signaling contributes to pharmacological functions, and how to drug a difficult target such as the PTH1R toward discovering nonpeptidic small molecule candidates for the treatment of metabolic bone and mineral diseases.
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Affiliation(s)
- Jean-Pierre Vilardaga
- Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Lisa J Clark
- Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Alex D White
- Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Ieva Sutkeviciute
- Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Ji Young Lee
- Department of Computational and Systems Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Ivet Bahar
- Department of Computational and Systems Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
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10
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Tang X, Bian J, Li Z. Post-Translational Modifications in GPCR Internalization. Am J Physiol Cell Physiol 2022; 323:C84-C94. [PMID: 35613355 DOI: 10.1152/ajpcell.00015.2022] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
G protein-coupled receptors (GPCRs) are the largest family of membrane receptors that serve as the most important drug targets. Classically, GPCR internalization has been considered to lead to receptor desensitization. However, many studies over the past decade have reported that internalized membrane receptors can trigger distinct signal activation. The "internalized activation" provides a completely new understanding for the receptor internalization, the mechanism of physiology/pathology and novel drug targets for precision medicine. GPCR internalization undergoes a series of strict regulations, especially by post-translational modifications (PTMs). Here, this review summarizes different PTMs in GPCR internalization and analyzes their significance in GPCR internalization dynamics, internalization routes, post-internalization fates and related diseases, which will offer new insights into the regulatory mechanism of GPCR signaling and novel drug targets for precision medicine.
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Affiliation(s)
- Xueqing Tang
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing Key Laboratory of Cardiovascular Receptors Research, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Ministry of Health, Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education, Beijing, China
| | - Jingwei Bian
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing Key Laboratory of Cardiovascular Receptors Research, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Ministry of Health, Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education, Beijing, China
| | - Zijian Li
- Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital, Beijing Key Laboratory of Cardiovascular Receptors Research, Key Laboratory of Cardiovascular Molecular Biology and Regulatory Peptides, Ministry of Health, Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education, Beijing, China.,Department of Pharmacy, Peking University Third Hospital, Beijing, China
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11
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Wedegaertner H, Pan WA, Gonzalez CC, Gonzalez DJ, Trejo J. The α-Arrestin ARRDC3 Is an Emerging Multifunctional Adaptor Protein in Cancer. Antioxid Redox Signal 2022; 36:1066-1079. [PMID: 34465145 PMCID: PMC9127825 DOI: 10.1089/ars.2021.0193] [Citation(s) in RCA: 19] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/18/2021] [Accepted: 08/21/2021] [Indexed: 01/04/2023]
Abstract
Significance: Adaptor proteins control the spatiotemporal dynamics of cellular signaling. Dysregulation of adaptor protein function can cause aberrant cell signaling and promote cancer. The arrestin family of adaptor proteins are known to regulate signaling by the superfamily of G protein-coupled receptors (GPCRs). The GPCRs are highly druggable and implicated in cancer progression. However, the molecular mechanisms responsible for arrestin dysregulation and the impact on GPCR function in cancer have yet to be fully elucidated. Recent Advances: A new family of mammalian arrestins, termed the α-arrestins, was recently discovered. The α-arrestin, arrestin domain-containing protein 3 (ARRDC3), in particular, has been identified as a tumor suppressor and is reported to control cellular signaling of GPCRs in cancer. Critical Issues: Compared with the extensively studied mammalian β-arrestins, there is limited information regarding the regulatory mechanisms that control α-arrestin activation and function. Here, we discuss the molecular mechanisms that regulate ARRDC3, which include post-translational modifications such as phosphorylation and ubiquitination. We also provide evidence that ARRDC3 can interact with a wide array of proteins that control diverse biological functions. Future Directions: ARRDC3 interacts with numerous proteins and is likely to display diverse functions in cancer, metabolic disease, and other syndromes. Thus, understanding the regulatory mechanisms of ARRDC3 activity in various cellular contexts is critically important. Recent studies suggest that α-arrestins may be regulated through post-translational modification, which is known to impact adaptor protein function. However, additional studies are needed to determine how these regulatory mechanisms affect ARRDC3 tumor suppressor function. Antioxid. Redox Signal. 36, 1066-1079.
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Affiliation(s)
- Helen Wedegaertner
- Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla, California, USA
- Biomedical Sciences Graduate Program, University of California, San Diego, La Jolla, California, USA
| | - Wen-An Pan
- Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla, California, USA
| | - Carlos C. Gonzalez
- Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla, California, USA
| | - David J. Gonzalez
- Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla, California, USA
- Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California, USA
| | - JoAnn Trejo
- Department of Pharmacology, School of Medicine, University of California, San Diego, La Jolla, California, USA
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12
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Che F, Xu Q, Li Q, Huang ZX, Yang CW, Wang LY, Wei Y, Shi YJ, Song B. Radiomics signature: A potential biomarker for β-arrestin1 phosphorylation prediction in hepatocellular carcinoma. World J Gastroenterol 2022; 28:1479-1493. [PMID: 35582676 PMCID: PMC9048469 DOI: 10.3748/wjg.v28.i14.1479] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/09/2021] [Revised: 01/22/2022] [Accepted: 03/06/2022] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND The phosphorylation status of β-arrestin1 influences its function as a signal strongly related to sorafenib resistance. This retrospective study aimed to develop and validate radiomics-based models for predicting β-arrestin1 phosphorylation in hepatocellular carcinoma (HCC) using whole-lesion radiomics and visual imaging features on preoperative contrast-enhanced computed tomography (CT) images. AIM To develop and validate radiomics-based models for predicting β-arrestin1 phosphorylation in HCC using radiomics with contrast-enhanced CT. METHODS Ninety-nine HCC patients (training cohort: n = 69; validation cohort: n = 30) receiving systemic sorafenib treatment after surgery were enrolled in this retrospective study. Three-dimensional whole-lesion regions of interest were manually delineated along the tumor margins on portal venous CT images. Radiomics features were generated and selected to build a radiomics score using logistic regression analysis. Imaging features were evaluated by two radiologists independently. All these features were combined to establish clinico-radiological (CR) and clinico-radiological-radiomics (CRR) models by using multivariable logistic regression analysis. The diagnostic performance and clinical usefulness of the models were measured by receiver operating characteristic and decision curves, and the area under the curve (AUC) was determined. Their association with prognosis was evaluated using the Kaplan-Meier method. RESULTS Four radiomics features were selected to construct the radiomics score. In the multivariate analysis, alanine aminotransferase level, tumor size and tumor margin on portal venous phase images were found to be significant independent factors for predicting β-arrestin1 phosphorylation-positive HCC and were included in the CR model. The CRR model integrating the radiomics score with clinico-radiological risk factors showed better discriminative performance (AUC = 0.898, 95%CI, 0.820 to 0.977) than the CR model (AUC = 0.794, 95%CI, 0.686 to 0.901; P = 0.011), with increased clinical usefulness confirmed in both the training and validation cohorts using decision curve analysis. The risk of β-arrestin1 phosphorylation predicted by the CRR model was significantly associated with overall survival in the training and validation cohorts (log-rank test, P < 0.05). CONCLUSION The radiomics signature is a reliable tool for evaluating β-arrestin1 phosphorylation which has prognostic significance for HCC patients, providing the potential to better identify patients who would benefit from sorafenib treatment.
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Affiliation(s)
- Feng Che
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Qing Xu
- Institute of Clinical Pathology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Qian Li
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Zi-Xing Huang
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Cai-Wei Yang
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Li Ye Wang
- Department of Research and Development, Shanghai United Imaging Intelligence Co., Ltd, Shanghai 200232, China
| | - Yi Wei
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Yu-Jun Shi
- Institute of Clinical Pathology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
| | - Bin Song
- Department of Radiology, West China Hospital, Sichuan University, Chengdu 610041, Sichuan Province, China
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13
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Reiter E. [β-arrestins, their mechanisms of action and multiple roles in the biology of G protein-coupled receptors]. Biol Aujourdhui 2022; 215:107-118. [PMID: 35275055 DOI: 10.1051/jbio/2021010] [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: 09/07/2021] [Indexed: 06/14/2023]
Abstract
The stimulation of G protein-coupled receptors (GPCRs) induces biological responses to a wide range of extracellular cues. The heterotrimeric G proteins, which are recruited to the active conformation of GPCRs, lead to the generation of various diffusible second messengers. Only two other families of proteins exhibit the remarkable characteristic of recognizing and binding to the active conformation of most GPCRs: GPCR kinases (GRKs) and β-arrestins. These two families of proteins were initially identified as key players in the desensitization of G protein activation by GPCRs. Over the years, β-arrestins have been implicated in an increasing number of interactions with non-receptor proteins, expanding the range of cellular functions in which they are involved. It is now well established that β-arrestins, by scaffolding and recruiting protein complexes in an agonist-dependent manner, directly regulate the trafficking and signaling of GPCRs. Remarkable advances have been made in recent years which have made it possible i) to identify biased ligands capable, by stabilizing particular conformations of a growing number of GPCRs, of activating or blocking the action of β-arrestins independently of that of G proteins, some of these ligands holding great therapeutic interest; ii) to demonstrate β-arrestins' role in the compartmentalization of GPCR signaling within the cell, and iii) to understand the molecular details of their interaction with GPCRs and of their activation through structural and biophysical approaches.
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Affiliation(s)
- Eric Reiter
- CNRS, IFCE, INRAE, Université de Tours, PRC, 37380 Nouzilly, France - Inria, Centre de recherche Inria Saclay-Île-de-France, 91120 Palaiseau, France
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14
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Redpath G, Deo N. Serotonin: an overlooked regulator of endocytosis and endosomal sorting? Biol Open 2022; 11:bio059057. [PMID: 35076063 PMCID: PMC8801889 DOI: 10.1242/bio.059057] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2021] [Accepted: 12/09/2021] [Indexed: 12/23/2022] Open
Abstract
Serotonin is a neurotransmitter and a hormone that is typically associated with regulating our mood. However, the serotonin transporter and receptors are expressed throughout the body, highlighting the much broader, systemic role of serotonin in regulating human physiology. A substantial body of data strongly implicates serotonin as a fundamental regulator of endocytosis and endocytic sorting. Serotonin has the potential to enhance endocytosis through three distinct mechanisms - serotonin signalling, serotonylation and insertion into the plasma membrane - although the interplay and relationship between these mechanisms has not yet been explored. Endocytosis is central to the cellular response to the extracellular environment, controlling receptor distribution on the plasma membrane to modulate signalling, neurotransmitter release and uptake, circulating protein and lipid cargo uptake, and amino acid internalisation for cell proliferation. Uncovering the range of cellular and physiological circumstances in which serotonin regulates endocytosis is of great interest for our understanding of how serotonin regulates mood, and also the fundamental understanding of endocytosis and its regulation throughout the body. This article has an associated Future Leader to Watch interview with the first author of the paper.
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Affiliation(s)
- Gregory Redpath
- EMBL Australia Node in Single Molecule Science, School of Medical Sciences and the ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney 2052, Australia
| | - Nikita Deo
- Department of Biochemistry, University of Otago, Dunedin 9016, New Zealand
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15
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Patel M, Matti C, Grimsey NL, Legler DF, Javitch JA, Finlay DB, Glass M. Delineating the interactions between the cannabinoid CB 2 receptor and its regulatory effectors; β-arrestins and G protein-coupled receptor kinases. Br J Pharmacol 2021; 179:2223-2239. [PMID: 34811740 DOI: 10.1111/bph.15748] [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: 08/20/2021] [Revised: 10/20/2021] [Accepted: 11/10/2021] [Indexed: 11/27/2022] Open
Abstract
BACKGROUND AND PURPOSE The cannabinoid CB2 receptor (CB2 ) is a promising therapeutic target for modulating inflammation. However, little is known surrounding the mechanisms underpinning CB2 desensitisation and regulation, particularly the role of G protein-coupled receptor kinases (GRKs). Here, we evaluated the role of six GRK isoforms in β-arrestin recruitment to CB2 . Mutagenesis of several distal C-terminal aspartic acid residues was also performed in an attempt to delineate additional structural elements involved in the regulation of CB2 . EXPERIMENTAL APPROACH In CB2 -expressing HEK 293 cells, β-arrestin translocation was measured using real-time BRET assays. G protein dissociation BRET assays were performed to assess the activation and desensitisation of CB2 in the presence of β-arrestin 2. KEY RESULTS Overexpression of GRK isoforms 1-6 failed to considerably improve translocation of either β-arrestin 1 or β-arrestin 2 to CB2 . Consistent with this, inhibition of endogenous GRK2/3 did not substantially reduce β-arrestin 2 translocation. Mutagenesis of C-terminal aspartic acid residues resulted in attenuation of β-arrestin 2 translocation, which translated to a reduction in desensitisation of G protein activation. CONCLUSION AND IMPLICATIONS Our findings suggest that CB2 does not adhere to the classical GPCR regulatory paradigm, entailing GRK- and β-arrestin-mediated desensitisation. Instead, C-terminal aspartic acid residues may act as phospho-mimics to induce β-arrestin activation. This study provides novel insights into the regulatory mechanisms of CB2 , which may aid in our understanding of drug tolerance and dependence.
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Affiliation(s)
- Monica Patel
- Department of Pharmacology and Toxicology, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand
| | - Christoph Matti
- Biotechnology Institute Thurgau (BITg) at the University of Konstanz, Kreuzlingen, Switzerland
| | - Natasha L Grimsey
- Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand
| | - Daniel F Legler
- Biotechnology Institute Thurgau (BITg) at the University of Konstanz, Kreuzlingen, Switzerland.,Faculty of Biology, University of Konstanz, Konstanz, Germany.,Theodor Kocher Institute, University of Bern, Bern, Switzerland
| | - Jonathan A Javitch
- Department of Psychiatry and Pharmacology, Columbia University Vagelos College of Physicians and Surgeons, New York, New York, United States.,Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, New York, United States
| | - David B Finlay
- Department of Pharmacology and Toxicology, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand
| | - Michelle Glass
- Department of Pharmacology and Toxicology, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand
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16
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Pathological AT1R-B2R Protein Aggregation and Preeclampsia. Cells 2021; 10:cells10102609. [PMID: 34685589 PMCID: PMC8533718 DOI: 10.3390/cells10102609] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2021] [Revised: 09/22/2021] [Accepted: 09/27/2021] [Indexed: 01/07/2023] Open
Abstract
Preeclampsia is one of the most frequent and severe complications of pregnancy. Symptoms of preeclampsia usually occur after 20 weeks of pregnancy and include hypertension and kidney dysfunction with proteinuria. Up to now, delivery of the infant has been the most effective and life-saving treatment to alleviate symptoms of preeclampsia because a causative treatment does not exist, which could prolong a pregnancy complicated with preeclampsia. Preeclampsia is a complex medical condition, which is attributed to a variety of different risk factors and causes. Risk factors account for insufficient placentation and impaired vasculogenesis and finally culminate in this life-threatening condition of pregnancy. Despite progress, many pathomechanisms and causes of preeclampsia are still incompletely understood. In recent years, it was found that excessive protein complex formation between G-protein-coupled receptors is a common sign of preeclampsia. Specifically, the aberrant heteromerization of two vasoactive G-protein-coupled receptors (GPCRs), the angiotensin II AT1 receptor and the bradykinin B2 receptor, is a causative factor of preeclampsia symptoms. Based on this knowledge, inhibition of abnormal GPCR protein complex formation is an experimental treatment approach of preeclampsia. This review summarizes the impact of pathological GPCR protein aggregation on symptoms of preeclampsia and delineates potential new therapeutic targets.
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17
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Jean-Alphonse FG, Sposini S. Confocal and TIRF microscopy based approaches to visualize arrestin trafficking in living cells. Methods Cell Biol 2021; 166:179-203. [PMID: 34752332 DOI: 10.1016/bs.mcb.2021.06.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023]
Abstract
Arrestins are key proteins that serve as versatile scaffolds to control and mediate G protein coupled receptors (GPCR) activity. Arrestin control of GPCR functions involves their recruitment from the cytosol to plasma membrane-localized GPCRs and to endosomal compartments, where they mediate internalization, sorting and signaling of GPCRs. Several methods can be used to monitor trafficking of arrestins; however, live fluorescence imaging remains the method of choice to both assess arrestin recruitment to ligand-activated receptors and to monitor its dynamic subcellular localization. Here, we present two approaches based on Total Internal Fluorescence (TIRF) microscopy and confocal microscopy to visualize arrestin trafficking in live cells in real time and to assess their co-localization with the GPCR of interest and their localization at specific subcellular locations.
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Affiliation(s)
- Frédéric Gaëtan Jean-Alphonse
- CNRS, IFCE, INRAE, Université de Tours, PRC, Nouzilly, France; Université Paris-Saclay, Inria, Inria Saclay-Île-de-France, Palaiseau, France
| | - Silvia Sposini
- Department of Metabolism, Digestion and Reproduction, Institute of Reproductive and Developmental Biology, Imperial College London, London, United Kingdom; University of Bordeaux, CNRS, Interdisciplinary Institute for Neuroscience, Bordeaux, France.
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18
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Fang Y, Jiang Q, Li S, Zhu H, Xu R, Song N, Ding X, Liu J, Chen M, Song M, Ding J, Lu M, Wu G, Hu G. Opposing functions of β-arrestin 1 and 2 in Parkinson's disease via microglia inflammation and Nprl3. Cell Death Differ 2021; 28:1822-1836. [PMID: 33686256 PMCID: PMC8184754 DOI: 10.1038/s41418-020-00704-9] [Citation(s) in RCA: 36] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2020] [Revised: 11/25/2020] [Accepted: 11/30/2020] [Indexed: 01/14/2023] Open
Abstract
Although β-arrestins (ARRBs) regulate diverse physiological and pathophysiological processes, their functions and regulation in Parkinson's disease (PD) remain poorly defined. In this study, we show that the expression of β-arrestin 1 (ARRB1) and β-arrestin 2 (ARRB2) is reciprocally regulated in PD mouse models, particularly in microglia. ARRB1 ablation ameliorates, whereas ARRB2 knockout aggravates, the pathological features of PD, including dopaminergic neuron loss, neuroinflammation and microglia activation in vivo, and microglia-mediated neuron damage in vitro. We also demonstrate that ARRB1 and ARRB2 produce adverse effects on inflammation and activation of the inflammatory STAT1 and NF-κB pathways in primary cultures of microglia and macrophages and that two ARRBs competitively interact with the activated form of p65, a component of the NF-κB pathway. We further find that ARRB1 and ARRB2 differentially regulate the expression of nitrogen permease regulator-like 3 (Nprl3), a functionally poorly characterized protein, as revealed by RNA sequencing, and that in the gain- and loss-of-function studies, Nprl3 mediates the functions of both ARRBs in microglia inflammatory responses. Collectively, these data demonstrate that two closely related ARRBs exert opposite functions in microglia-mediated inflammation and the pathogenesis of PD which are mediated at least in part through Nprl3 and provide novel insights into the understanding of the functional divergence of ARRBs in PD.
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Affiliation(s)
- Yinquan Fang
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Qingling Jiang
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Shanshan Li
- Department of Pharmacology, Nanjing University of Chinese Medicine, 138 Xianlin Avenue, Nanjing, 210023, Jiangsu, China
| | - Hong Zhu
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Rong Xu
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Nanshan Song
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Xiao Ding
- Department of Pharmacology, Nanjing University of Chinese Medicine, 138 Xianlin Avenue, Nanjing, 210023, Jiangsu, China
| | - Jiaqi Liu
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Miaomiao Chen
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Mengmeng Song
- Department of Pharmacology, Nanjing University of Chinese Medicine, 138 Xianlin Avenue, Nanjing, 210023, Jiangsu, China
| | - Jianhua Ding
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Ming Lu
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China
| | - Guangyu Wu
- Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta University, 1459 Laney Walker Blvd., Augusta, GA, 30912, USA
| | - Gang Hu
- Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical University, 818 Tianyuan East Road, Nanjing, 211166, Jiangsu, China.
- Department of Pharmacology, Nanjing University of Chinese Medicine, 138 Xianlin Avenue, Nanjing, 210023, Jiangsu, China.
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19
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Abreu N, Acosta-Ruiz A, Xiang G, Levitz J. Mechanisms of differential desensitization of metabotropic glutamate receptors. Cell Rep 2021; 35:109050. [PMID: 33910009 PMCID: PMC9750234 DOI: 10.1016/j.celrep.2021.109050] [Citation(s) in RCA: 35] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2021] [Revised: 03/04/2021] [Accepted: 04/07/2021] [Indexed: 12/16/2022] Open
Abstract
G protein-coupled receptors (GPCRs) interact with intracellular transducers to control both signal initiation and desensitization, but the distinct mechanisms that control the regulation of different GPCR subtypes are unclear. Here we use fluorescence imaging and electrophysiology to examine the metabotropic glutamate receptor (mGluR) family. We find distinct properties across subtypes in both rapid desensitization and internalization, with striking differences between the group II mGluRs. mGluR3, but not mGluR2, undergoes glutamate-dependent rapid desensitization, internalization, trafficking, and recycling. We map differences between mGluRs to variable Ser/Thr-rich sequences in the C-terminal domain (CTD) that control interaction with both GPCR kinases and β-arrestins. Finally, we identify a cancer-associated mutation, G848E, within the mGluR3 CTD that enhances β-arrestin coupling and internalization, enabling an analysis of mGluR3 β-arrestin-coupling properties and revealing biased variants. Together, this work provides a framework for understanding the distinct regulation and functional roles of mGluR subtypes.
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Affiliation(s)
- Nohely Abreu
- Biochemistry, Cell and Molecular Biology Graduate Program, Weill Cornell Medicine, New York, NY, USA; Department of Biochemistry, Weill Cornell Medicine, New York, NY, USA
| | - Amanda Acosta-Ruiz
- Biochemistry, Cell and Molecular Biology Graduate Program, Weill Cornell Medicine, New York, NY, USA; Department of Biochemistry, Weill Cornell Medicine, New York, NY, USA
| | - Guoqing Xiang
- Department of Biochemistry, Weill Cornell Medicine, New York, NY, USA
| | - Joshua Levitz
- Biochemistry, Cell and Molecular Biology Graduate Program, Weill Cornell Medicine, New York, NY, USA; Department of Biochemistry, Weill Cornell Medicine, New York, NY, USA.
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20
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Bondar A, Jang W, Sviridova E, Lambert NA. Components of the G s signaling cascade exhibit distinct changes in mobility and membrane domain localization upon β 2 -adrenergic receptor activation. Traffic 2021; 21:324-332. [PMID: 32096320 DOI: 10.1111/tra.12724] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2019] [Revised: 02/21/2020] [Accepted: 02/21/2020] [Indexed: 12/12/2022]
Abstract
The G protein signaling cascade is a key player in cell signaling. Cascade activation leads to a redistribution of its members in various cellular compartments. These changes are likely related to the "second wave" of signaling from endosomes. Here, we set out to determine whether Gs signaling cascade members expressed at very low levels exhibit altered mobility and localize in clathrin-coated structures (CCSs) or caveolae upon activation by β2 -adrenergic receptors (β2 AR). Activated β2 AR showed decreased mobility and sustained accumulation in CCSs but not in caveolae. Arrestin 3 translocated to the plasma membrane after β2 AR activation and showed very low mobility and pronounced accumulation in CCSs. In contrast, Gαs and Gγ2 exhibited a modest reduction in mobility but no detectable accumulation in or exclusion from CCSs or caveolae. The effector adenylyl cyclase 5 (AC5) showed a slight mobility increase upon β2 AR stimulation, no redistribution to CCSs, and weak activation-independent accumulation in caveolae. Our findings show an overall decrease in the mobility of most activated Gs signaling cascade members and confirm that β2 AR and arrestin 3 accumulate in CCSs, while Gαs , Gγ2 and AC5 can transiently enter CCSs and caveolae but do not accumulate in and are not excluded from these domains.
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Affiliation(s)
- Alexey Bondar
- Department of Pharmacology and Toxicology, Augusta University, Augusta, Georgia, USA.,Center for Nanobiology and Structural Biology, Institute of Microbiology of the Academy of Sciences of the Czech Republic, Czech Republic.,Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Prague, Czech Republic.,University of South Bohemia, Czech Republic
| | - Wonjo Jang
- Department of Pharmacology and Toxicology, Augusta University, Augusta, Georgia, USA
| | - Ekaterina Sviridova
- Center for Nanobiology and Structural Biology, Institute of Microbiology of the Academy of Sciences of the Czech Republic, Czech Republic
| | - Nevin A Lambert
- Department of Pharmacology and Toxicology, Augusta University, Augusta, Georgia, USA
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21
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Aydin Y, Coin I. Biochemical insights into structure and function of arrestins. FEBS J 2021; 288:2529-2549. [DOI: 10.1111/febs.15811] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2020] [Revised: 02/26/2021] [Accepted: 03/09/2021] [Indexed: 12/13/2022]
Affiliation(s)
- Yasmin Aydin
- Institute of Biochemistry Faculty of Life Sciences University of Leipzig Germany
| | - Irene Coin
- Institute of Biochemistry Faculty of Life Sciences University of Leipzig Germany
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22
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Dissecting the structural features of β-arrestins as multifunctional proteins. BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS 2021; 1869:140603. [PMID: 33421644 DOI: 10.1016/j.bbapap.2021.140603] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 11/06/2020] [Revised: 12/21/2020] [Accepted: 01/04/2021] [Indexed: 02/08/2023]
Abstract
β-arrestins bind active G protein-coupled receptors (GPCRs) and play a crucial role in receptor desensitization and internalization. The classical paradigm of arrestin function has been expanded with the identification of many non-receptor-binding partners, which indicated the multifunctional role of β-arrestins in cellular functions. To elucidate the molecular mechanism of β-arrestin-mediated signaling, the structural features of β-arrestins were investigated using X-ray crystallography and cryogenic electron microscopy (cryo-EM). However, the intrinsic conformational flexibility of β-arrestins hampers the elucidation of structural interactions between β-arrestins and their binding partners using conventional structure determination tools. Therefore, structural information obtained using complementary structure analysis techniques would be necessary in combination with X-ray crystallography and cryo-EM data. In this review, we describe how β-arrestins interact with their binding partners from a structural point of view, as elucidated by both traditional methods (X-ray crystallography and cryo-EM) and complementary structure analysis techniques.
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23
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Lee C, Viswanathan G, Choi I, Jassal C, Kohlmann T, Rajagopal S. Beta-Arrestins and Receptor Signaling in the Vascular Endothelium. Biomolecules 2020; 11:biom11010009. [PMID: 33374806 PMCID: PMC7824595 DOI: 10.3390/biom11010009] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2020] [Revised: 12/13/2020] [Accepted: 12/19/2020] [Indexed: 12/17/2022] Open
Abstract
The vascular endothelium is the innermost layer of blood vessels and is a key regulator of vascular tone. Endothelial function is controlled by receptor signaling through G protein-coupled receptors, receptor tyrosine kinases and receptor serine-threonine kinases. The β-arrestins, multifunctional adapter proteins, have the potential to regulate all of these receptor families, although it is unclear as to whether they serve to integrate signaling across all of these different axes. Notably, the β-arrestins have been shown to regulate signaling by a number of receptors important in endothelial function, such as chemokine receptors and receptors for vasoactive substances such as angiotensin II, endothelin-1 and prostaglandins. β-arrestin-mediated signaling pathways have been shown to play central roles in pathways that control vasodilation, cell proliferation, migration, and immune function. At this time, the physiological impact of this signaling has not been studied in detail, but a deeper understanding of it could lead to the development of novel therapies for the treatment of vascular disease.
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Affiliation(s)
- Claudia Lee
- Department of Biochemistry, School of Medicine, Duke University, Durham, NC 27710, USA;
| | - Gayathri Viswanathan
- Medical Center, Department of Medicine, Division of Cardiology, Duke University, Durham, NC 27710, USA; (G.V.); (I.C.)
| | - Issac Choi
- Medical Center, Department of Medicine, Division of Cardiology, Duke University, Durham, NC 27710, USA; (G.V.); (I.C.)
| | - Chanpreet Jassal
- College of Arts and Sciences, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA;
| | - Taylor Kohlmann
- Trinity College of Arts and Sciences, Duke University, Durham, NC 27708, USA;
| | - Sudarshan Rajagopal
- Department of Biochemistry, School of Medicine, Duke University, Durham, NC 27710, USA;
- Medical Center, Department of Medicine, Division of Cardiology, Duke University, Durham, NC 27710, USA; (G.V.); (I.C.)
- Correspondence:
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24
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Ozawa K, Tsumoto H, Miura Y, Yamaguchi J, Iguchi-Ariga SMM, Sakuma T, Yamamoto T, Uchiyama Y. DJ-1 is indispensable for the S-nitrosylation of Parkin, which maintains function of mitochondria. Sci Rep 2020; 10:4377. [PMID: 32152416 PMCID: PMC7062835 DOI: 10.1038/s41598-020-61287-6] [Citation(s) in RCA: 34] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2019] [Accepted: 02/20/2020] [Indexed: 01/10/2023] Open
Abstract
The DJ-1 gene, a causative gene for familial Parkinson’s disease (PD), has been reported to have various functions, including transcriptional regulation, antioxidant response, and chaperone and protease functions; however, the molecular mechanism associated with the pathogenesis of PD remains elusive. To further explore the molecular function of DJ-1 in the pathogenesis of PD, we compared protein expression profiles in brain tissues from wild-type and DJ-1-deficient mice. Two-dimensional difference gel electrophoresis analysis and subsequent analysis using data mining methods revealed alterations in the expression of molecules associated with energy production. We demonstrated that DJ-1 deletion inhibited S-nitrosylation of endogenous Parkin as well as overexpressed Parkin in neuroblastoma cells and mouse brain tissues. Thus, we used genome editing to generate neuroblastoma cells with DJ-1 deletion or S-nitrosylated cysteine mutation in Parkin and demonstrated that these cells exhibited similar phenotypes characterized by enhancement of cell death under mitochondrial depolarization and dysfunction of mitochondria. Our data indicate that DJ-1 is required for the S-nitrosylation of Parkin, which positively affects mitochondrial function, and suggest that the denitrosylation of Parkin via DJ-1 inactivation might contribute to PD pathogenesis and act as a therapeutic target.
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Affiliation(s)
- Kentaro Ozawa
- Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology, 35-2 Sakae-cho, Itabashi-ku, Tokyo, 173-0015, Japan. .,Department of Pharmacology, Nara Medical University School of Medicine, Kashihara City, Nara, 634-8521, Japan. .,Department of Cellular and Molecular Neuropathology, Juntendo University Graduate School of Medicine, Bunkyo-Ku 2-1-1, Tokyo, 113-8421, Japan. .,Asakayama General Hospital, Sakai-ku, Sakai City, Osaka, 590-0018, Japan.
| | - Hiroki Tsumoto
- Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology, 35-2 Sakae-cho, Itabashi-ku, Tokyo, 173-0015, Japan
| | - Yuri Miura
- Research Team for Mechanism of Aging, Tokyo Metropolitan Institute of Gerontology, 35-2 Sakae-cho, Itabashi-ku, Tokyo, 173-0015, Japan
| | - Junji Yamaguchi
- Department of Cellular and Molecular Neuropathology, Juntendo University Graduate School of Medicine, Bunkyo-Ku 2-1-1, Tokyo, 113-8421, Japan
| | - Sanae M M Iguchi-Ariga
- Faculty of Pharmaceutical Sciences, Hokkaido University, Kita 12, Nishi 6, Kita-ku, Sapporo, 060-0812, Japan
| | - Tetsushi Sakuma
- Division of Integrated Sciences for Life, Graduate School of Integrated Sciences for Life, Hiroshima University, Hiroshima, 739-8526, Japan
| | - Takashi Yamamoto
- Division of Integrated Sciences for Life, Graduate School of Integrated Sciences for Life, Hiroshima University, Hiroshima, 739-8526, Japan
| | - Yasuo Uchiyama
- Department of Cellular and Molecular Neuropathology, Juntendo University Graduate School of Medicine, Bunkyo-Ku 2-1-1, Tokyo, 113-8421, Japan
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25
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Conformational Dynamics and Functional Implications of Phosphorylated β-Arrestins. Structure 2020; 28:314-323.e3. [DOI: 10.1016/j.str.2019.12.008] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2019] [Revised: 11/25/2019] [Accepted: 12/21/2019] [Indexed: 12/18/2022]
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26
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Myagmar BE, Ismaili T, Swigart PM, Raghunathan A, Baker AJ, Sahdeo S, Blevitt JM, Milla ME, Simpson PC. Coupling to Gq Signaling Is Required for Cardioprotection by an Alpha-1A-Adrenergic Receptor Agonist. Circ Res 2019; 125:699-706. [PMID: 31426700 DOI: 10.1161/circresaha.118.314416] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
RATIONALE Gq signaling in cardiac myocytes is classically considered toxic. Targeting Gq directly to test this is problematic, because cardiac myocytes have many Gq-coupled receptors. OBJECTIVE Test whether Gq coupling is required for the cardioprotective effects of an alpha-1A-AR (adrenergic receptor) agonist. METHODS AND RESULTS In recombinant cells, a mouse alpha-1A-AR with a 6-residue substitution in the third intracellular loop does not couple to Gq signaling. Here we studied a knockin mouse with this alpha-1A-AR mutation. Heart alpha-1A receptor levels and antagonist affinity in the knockin were identical to wild-type. In wild-type cardiac myocytes, the selective alpha-1A agonist A61603-stimulated phosphoinositide-phospholipase C and myocyte contraction. In myocytes with the alpha-1A knockin, both A61603 effects were absent, indicating that Gq coupling was absent. Surprisingly, A61603 activation of cardioprotective ERK (extracellular signal-regulated kinase) was markedly impaired in the KI mutant myocytes, and A61603 did not protect mutant myocytes from doxorubicin toxicity in vitro. Similarly, mice with the α1A KI mutation had increased mortality after transverse aortic constriction, and A61603 did not rescue cardiac function in mice with the Gq coupling-defective alpha-1A receptor. CONCLUSIONS Gq coupling is required for cardioprotection by an alpha-1A-AR agonist. Gq signaling can be adaptive.
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Affiliation(s)
- Bat-Erdene Myagmar
- From the VA Medical Center, San Francisco, CA (B.-E.M., P.M.S., A.R., A.J.B., P.C.S.).,University of California, San Francisco (B.-E.M., A.J.B., P.C.S.)
| | - Taylor Ismaili
- Janssen Research and Development, San Diego, CA (T.I., S.S., J.M.B.)
| | - Philip M Swigart
- From the VA Medical Center, San Francisco, CA (B.-E.M., P.M.S., A.R., A.J.B., P.C.S.)
| | - Anaha Raghunathan
- From the VA Medical Center, San Francisco, CA (B.-E.M., P.M.S., A.R., A.J.B., P.C.S.)
| | - Anthony J Baker
- From the VA Medical Center, San Francisco, CA (B.-E.M., P.M.S., A.R., A.J.B., P.C.S.).,University of California, San Francisco (B.-E.M., A.J.B., P.C.S.)
| | - Sunil Sahdeo
- Janssen Research and Development, San Diego, CA (T.I., S.S., J.M.B.)
| | | | | | - Paul C Simpson
- From the VA Medical Center, San Francisco, CA (B.-E.M., P.M.S., A.R., A.J.B., P.C.S.).,University of California, San Francisco (B.-E.M., A.J.B., P.C.S.)
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27
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Gurevich VV, Gurevich EV, Uversky VN. Arrestins: structural disorder creates rich functionality. Protein Cell 2018; 9:986-1003. [PMID: 29453740 PMCID: PMC6251804 DOI: 10.1007/s13238-017-0501-8] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2017] [Accepted: 11/27/2017] [Indexed: 01/14/2023] Open
Abstract
Arrestins are soluble relatively small 44-46 kDa proteins that specifically bind hundreds of active phosphorylated GPCRs and dozens of non-receptor partners. There are binding partners that demonstrate preference for each of the known arrestin conformations: free, receptor-bound, and microtubule-bound. Recent evidence suggests that conformational flexibility in every functional state is the defining characteristic of arrestins. Flexibility, or plasticity, of proteins is often described as structural disorder, in contrast to the fixed conformational order observed in high-resolution crystal structures. However, protein-protein interactions often involve highly flexible elements that can assume many distinct conformations upon binding to different partners. Existing evidence suggests that arrestins are no exception to this rule: their flexibility is necessary for functional versatility. The data on arrestins and many other multi-functional proteins indicate that in many cases, "order" might be artificially imposed by highly non-physiological crystallization conditions and/or crystal packing forces. In contrast, conformational flexibility (and its extreme case, intrinsic disorder) is a more natural state of proteins, representing true biological order that underlies their physiologically relevant functions.
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Affiliation(s)
- Vsevolod V Gurevich
- Department of Pharmacology, Vanderbilt University, Nashville, TN, 37232, USA.
| | - Eugenia V Gurevich
- Department of Pharmacology, Vanderbilt University, Nashville, TN, 37232, USA
| | - Vladimir N Uversky
- Department of Molecular Medicine and USF Health Byrd Alzheimer's Research Institute, Morsani College of Medicine, University of South Florida, Tampa, FL, 33612, USA
- Institute for Biological Instrumentation, Russian Academy of Sciences, Pushchino, Moscow Region, Russia, 142290
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Quitterer U, Fu X, Pohl A, Bayoumy KM, Langer A, AbdAlla S. Beta-Arrestin1 Prevents Preeclampsia by Downregulation of Mechanosensitive AT1-B2 Receptor Heteromers. Cell 2018; 176:318-333.e19. [PMID: 30503206 DOI: 10.1016/j.cell.2018.10.050] [Citation(s) in RCA: 49] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2018] [Revised: 08/26/2018] [Accepted: 10/23/2018] [Indexed: 12/20/2022]
Abstract
Preeclampsia is the most frequent pregnancy-related complication worldwide with no cure. While a number of molecular features have emerged, the underlying causal mechanisms behind the disorder remain obscure. Here, we find that increased complex formation between angiotensin II AT1 and bradykinin B2, two G protein-coupled receptors with opposing effects on blood vessel constriction, triggers symptoms of preeclampsia in pregnant mice. Aberrant heteromerization of AT1-B2 led to exaggerated calcium signaling and high vascular smooth muscle mechanosensitivity, which could explain the onset of preeclampsia symptoms at late-stage pregnancy as mechanical forces increase with fetal mass. AT1-B2 receptor aggregation was inhibited by beta-arrestin-mediated downregulation. Importantly, symptoms of preeclampsia were prevented by transgenic ARRB1 expression or a small-molecule drug. Because AT1-B2 heteromerization was found to occur in human placental biopsies from pregnancies complicated by preeclampsia, specifically targeting AT1-B2 heteromerization and its downstream consequences represents a promising therapeutic approach.
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Affiliation(s)
- Ursula Quitterer
- Molecular Pharmacology, Department of Chemistry and Applied Biosciences, ETH Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland; Department of Medicine, Institute of Pharmacology and Toxicology, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland.
| | - Xuebin Fu
- Department of Microbiology and Immunology, Stanford University, 299 Campus Dr., Stanford, CA 94305, USA
| | - Armin Pohl
- Roche Diagnostics International AG, Forrenstrasse 2, 6343 Rotkreuz, Switzerland
| | - Karam M Bayoumy
- Clinic of Obstetrics and Gynecology, Ain Shams University Hospitals, Cairo 11566, Egypt
| | - Andreas Langer
- Molecular Pharmacology, Department of Chemistry and Applied Biosciences, ETH Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland
| | - Said AbdAlla
- Molecular Pharmacology, Department of Chemistry and Applied Biosciences, ETH Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland
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Intrathymic Notch3 and CXCR4 combinatorial interplay facilitates T-cell leukemia propagation. Oncogene 2018; 37:6285-6298. [PMID: 30038265 PMCID: PMC6284016 DOI: 10.1038/s41388-018-0401-2] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2017] [Revised: 05/20/2018] [Accepted: 06/10/2018] [Indexed: 12/27/2022]
Abstract
Notch hyperactivation dominates T-cell acute lymphoblastic leukemia development, but the mechanisms underlying “pre-leukemic” cell dissemination are still unclear. Here we describe how deregulated Notch3 signaling enhances CXCR4 cell-surface expression and migratory ability of CD4+CD8+ thymocytes, possibly contributing to “pre-leukemic” cell propagation, early in disease progression. In transgenic mice overexpressing the constitutively active Notch3 intracellular domain, we detect the progressive increase in circulating blood and bone marrow of CD4+CD8+ cells, characterized by high and combined surface expression of Notch3 and CXCR4. We report for the first time that transplantation of such CD4+CD8+ cells reveals their competence in infiltrating spleen and bone marrow of immunocompromised recipient mice. We also show that CXCR4 surface expression is central to the migratory ability of CD4+CD8+ cells and such an expression is regulated by Notch3 through β-arrestin in human leukemia cells. De novo, we propose that hyperactive Notch3 signaling by boosting CXCR4-dependent migration promotes anomalous egression of CD4+CD8+ cells from the thymus in early leukemia stages. In fact, in vivo CXCR4 antagonism prevents bone marrow colonization by such CD4+CD8+ cells in young Notch3 transgenic mice. Therefore, our data suggest that combined therapies precociously counteracting intrathymic Notch3/CXCR4 crosstalk may prevent dissemination of “pre-leukemic” CD4+CD8+ cells, by a “thymus-autonomous” mechanism.
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30
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Indrischek H, Prohaska SJ, Gurevich VV, Gurevich EV, Stadler PF. Uncovering missing pieces: duplication and deletion history of arrestins in deuterostomes. BMC Evol Biol 2017; 17:163. [PMID: 28683816 PMCID: PMC5501109 DOI: 10.1186/s12862-017-1001-4] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2016] [Accepted: 06/19/2017] [Indexed: 12/13/2022] Open
Abstract
BACKGROUND The cytosolic arrestin proteins mediate desensitization of activated G protein-coupled receptors (GPCRs) via competition with G proteins for the active phosphorylated receptors. Arrestins in active, including receptor-bound, conformation are also transducers of signaling. Therefore, this protein family is an attractive therapeutic target. The signaling outcome is believed to be a result of structural and sequence-dependent interactions of arrestins with GPCRs and other protein partners. Here we elucidated the detailed evolution of arrestins in deuterostomes. RESULTS Identity and number of arrestin paralogs were determined searching deuterostome genomes and gene expression data. In contrast to standard gene prediction methods, our strategy first detects exons situated on different scaffolds and then solves the problem of assigning them to the correct gene. This increases both the completeness and the accuracy of the annotation in comparison to conventional database search strategies applied by the community. The employed strategy enabled us to map in detail the duplication- and deletion history of arrestin paralogs including tandem duplications, pseudogenizations and the formation of retrogenes. The two rounds of whole genome duplications in the vertebrate stem lineage gave rise to four arrestin paralogs. Surprisingly, visual arrestin ARR3 was lost in the mammalian clades Afrotheria and Xenarthra. Duplications in specific clades, on the other hand, must have given rise to new paralogs that show signatures of diversification in functional elements important for receptor binding and phosphate sensing. CONCLUSION The current study traces the functional evolution of deuterostome arrestins in unprecedented detail. Based on a precise re-annotation of the exon-intron structure at nucleotide resolution, we infer the gain and loss of paralogs and patterns of conservation, co-variation and selection.
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Affiliation(s)
- Henrike Indrischek
- Computational EvoDevo Group, Department of Computer Science, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany.
- Bioinformatics Group, Department of Computer Science, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany.
- Interdisciplinary Center for Bioinformatics, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany.
| | - Sonja J Prohaska
- Computational EvoDevo Group, Department of Computer Science, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany
- Interdisciplinary Center for Bioinformatics, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany
| | - Vsevolod V Gurevich
- Department of Pharmacology, Vanderbilt University, 2200 Pierce Ave, Nashville, TN 37232, USA
| | - Eugenia V Gurevich
- Department of Pharmacology, Vanderbilt University, 2200 Pierce Ave, Nashville, TN 37232, USA
| | - Peter F Stadler
- Bioinformatics Group, Department of Computer Science, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany
- Interdisciplinary Center for Bioinformatics, Universität Leipzig, Härtelstraße 16-18, Leipzig, D-04107, Germany
- Max Planck Institute for Mathematics in the Sciences, Inselstraße 22, Leipzig, D-04103, Germany
- Fraunhofer Institute for Cell Therapy and Immunology, Perlickstraße 1, Leipzig, D-04103, Germany
- Department of Theoretical Chemistry, University of Vienna, Währinger Straße 17, Vienna, A-1090, Austria
- Center for non-coding RNA in Technology and Health, Grønegårdsvej 3, Frederiksberg C, DK-1870, Denmark
- Santa Fe Institute, 1399 Hyde Park Rd., Santa Fe, NM 87501, USA
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Peterson YK, Luttrell LM. The Diverse Roles of Arrestin Scaffolds in G Protein-Coupled Receptor Signaling. Pharmacol Rev 2017; 69:256-297. [PMID: 28626043 PMCID: PMC5482185 DOI: 10.1124/pr.116.013367] [Citation(s) in RCA: 324] [Impact Index Per Article: 40.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
The visual/β-arrestins, a small family of proteins originally described for their role in the desensitization and intracellular trafficking of G protein-coupled receptors (GPCRs), have emerged as key regulators of multiple signaling pathways. Evolutionarily related to a larger group of regulatory scaffolds that share a common arrestin fold, the visual/β-arrestins acquired the capacity to detect and bind activated GPCRs on the plasma membrane, which enables them to control GPCR desensitization, internalization, and intracellular trafficking. By acting as scaffolds that bind key pathway intermediates, visual/β-arrestins both influence the tonic level of pathway activity in cells and, in some cases, serve as ligand-regulated scaffolds for GPCR-mediated signaling. Growing evidence supports the physiologic and pathophysiologic roles of arrestins and underscores their potential as therapeutic targets. Circumventing arrestin-dependent GPCR desensitization may alleviate the problem of tachyphylaxis to drugs that target GPCRs, and find application in the management of chronic pain, asthma, and psychiatric illness. As signaling scaffolds, arrestins are also central regulators of pathways controlling cell growth, migration, and survival, suggesting that manipulating their scaffolding functions may be beneficial in inflammatory diseases, fibrosis, and cancer. In this review we examine the structure-function relationships that enable arrestins to perform their diverse roles, addressing arrestin structure at the molecular level, the relationship between arrestin conformation and function, and sites of interaction between arrestins, GPCRs, and nonreceptor-binding partners. We conclude with a discussion of arrestins as therapeutic targets and the settings in which manipulating arrestin function might be of clinical benefit.
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Affiliation(s)
- Yuri K Peterson
- Department of Drug Discovery and Biomedical Sciences, College of Pharmacy (Y.K.P.), and Departments of Medicine and Biochemistry and Molecular Biology (L.M.L.), Medical University of South Carolina, Charleston, South Carolina; and Ralph H. Johnson Veterans Affairs Medical Center, Charleston, South Carolina (L.M.L.)
| | - Louis M Luttrell
- Department of Drug Discovery and Biomedical Sciences, College of Pharmacy (Y.K.P.), and Departments of Medicine and Biochemistry and Molecular Biology (L.M.L.), Medical University of South Carolina, Charleston, South Carolina; and Ralph H. Johnson Veterans Affairs Medical Center, Charleston, South Carolina (L.M.L.)
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Ahmadzai MM, Broadbent D, Occhiuto C, Yang C, Das R, Subramanian H. Canonical and Noncanonical Signaling Roles of β-Arrestins in Inflammation and Immunity. Adv Immunol 2017; 136:279-313. [PMID: 28950948 DOI: 10.1016/bs.ai.2017.05.004] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
β-Arrestins are a highly conserved family of cytosolic adaptor proteins that contribute to many immune functions by orchestrating the desensitization and internalization of cell-surface G protein-coupled receptors (GPCRs) via well-studied canonical interactions. In cells of the innate and adaptive immune system, β-arrestins also subserve a parallel but less understood role in which they propagate, rather than terminate, intracellular signal transduction cascades. Because β-arrestins are promiscuous in their binding, they are capable of interacting with several different GPCRs and downstream effectors; in doing so, they vastly expand the repertoire of cellular responses evoked by agonist binding and the scope of responses that may contribute to inflammation during infectious and sterile insults. In this chapter, we attempt to provide an overview of the canonical and noncanonical roles of β-arrestins in inflammatory diseases.
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Affiliation(s)
| | | | | | - Canchai Yang
- Michigan State University, East Lansing, MI, United States
| | - Rupali Das
- Michigan State University, East Lansing, MI, United States
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Functional antagonism of β-arrestin isoforms balance IGF-1R expression and signalling with distinct cancer-related biological outcomes. Oncogene 2017; 36:5734-5744. [PMID: 28581517 PMCID: PMC5658667 DOI: 10.1038/onc.2017.179] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2017] [Revised: 04/09/2017] [Accepted: 05/01/2017] [Indexed: 12/21/2022]
Abstract
With very similar 3D structures, the widely expressed β-arrestin isoforms 1 and 2 play at times identical, distinct or even opposing roles in regulating various aspects of G protein-coupled receptors (GPCR) expression and signalling. Recent evidence recognizes the β-arrestin system as a key regulator of not only GPCRs, but also receptor tyrosine kinases, including the highly cancer relevant insulin-like growth factor type 1 receptor (IGF-1R). Binding of β-arrestin1 to IGF-1R leads to ligand-dependent degradation of the receptor and generates additional MAPK/ERK signalling, protecting cancer cells against anti-IGF-1R therapy. Because the interplay between β-arrestin isoforms governs the biological effects for most GPCRs, as yet unexplored for the IGF-1R, we sought to investigate specifically the regulatory roles of the β-arrestin2 isoform on expression and function of the IGF-1R. Results from controlled expression of either β-arrestin isoform demonstrate that β-arrestin2 acts in an opposite manner to β-arrestin1 by promoting degradation of an unstimulated IGF-1R, but protecting the receptor against agonist-induced degradation. Although both isoforms co-immunoprecipitate with IGF-1R, the ligand-occupied receptor has greater affinity for β-arrestin1; this association lasts longer, sustains MAPK/ERK signalling and mitigates p53 activation. Conversely, β-arrestin2 has greater affinity for the ligand-unoccupied receptor; this interaction is transient, triggers receptor ubiquitination and degradation without signalling activation, and leads to a lack of responsiveness to IGF-1, cell cycle arrest and decreased viability of cancer cells. This study reveals contrasting abilities of IGF-1R to interact with each β-arrestin isoform, depending on the presence of the ligand and demonstrates the antagonism between the two β-arrestin isoforms in controlling IGF-1R expression and function, which could be developed into a practical anti-IGF-1R strategy for cancer therapy.
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34
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Ferrão FM, Cardoso LHD, Drummond HA, Li XC, Zhuo JL, Gomes DS, Lara LS, Vieyra A, Lowe J. Luminal ANG II is internalized as a complex with AT 1R/AT 2R heterodimers to target endoplasmic reticulum in LLC-PK 1 cells. Am J Physiol Renal Physiol 2017; 313:F440-F449. [PMID: 28468964 DOI: 10.1152/ajprenal.00261.2016] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2016] [Revised: 04/18/2017] [Accepted: 04/23/2017] [Indexed: 12/28/2022] Open
Abstract
ANG II has many biological effects in renal physiology, particularly in Ca2+ handling in the regulation of fluid and solute reabsorption. It involves the systemic endocrine renin-angiotensin system (RAS), but tissue and intracrine ANG II are also known. We have shown that ANG II induces heterodimerization of its AT1 and AT2 receptors (AT1R and AT2R) to stimulate sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) activity. Thus, we investigated whether ANG II-AT1R/AT2R complex is formed and internalized, and also examined the intracellular localization of this complex to determine how its effect might be exerted on renal intracrine RAS. Living cell imaging of LLC-PK1 cells, quantification of extracellular ANG II, and use of the receptor antagonists, losartan and PD123319, showed that ANG II is internalized with AT1R/AT2R heterodimers as a complex in a microtubule-dependent and clathrin-independent manner, since colchicine-but not Pitstop2-blocked this process. This result was confirmed by an increase of β-arrestin phosphorylation after ANG II treatment, clathrin-mediated endocytosis being dependent on dephosphorylation of β-arrestin. Internalized ANG II colocalized with an endoplasmic reticulum (ER) marker and increased levels of AT1R, AT2R, and PKCα in ER-enriched membrane fractions. This novel evidence suggests the internalization of an ANG II-AT1/AT2 complex to target ER, where it might trigger intracellular Ca2+ responses.
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Affiliation(s)
- Fernanda M Ferrão
- Laboratório de Físico-Química Biológica Aída Hassón-Voloch, Instituto de Biofísica Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Luiza H D Cardoso
- Laboratório de Físico-Química Biológica Aída Hassón-Voloch, Instituto de Biofísica Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Heather A Drummond
- Department of Physiology and Biophysics and the Center for Excellence in Cardiovascular-Renal Research, University of Mississippi Medical Center, Jackson, Mississippi
| | - Xiao C Li
- Department of Pharmacology and Toxicology, University of Mississippi Medical Center, Jackson, Mississippi
| | - Jia L Zhuo
- Department of Pharmacology and Toxicology, University of Mississippi Medical Center, Jackson, Mississippi
| | - Dayene S Gomes
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Lucienne S Lara
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Adalberto Vieyra
- Laboratório de Físico-Química Biológica Aída Hassón-Voloch, Instituto de Biofísica Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.,Centro Nacional de Biologia Estrutural e Bioimagem, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; and.,National Institute of Science and Technology for Regenerative Medicine, Rio de Janeiro, Brazil
| | - Jennifer Lowe
- Laboratório de Físico-Química Biológica Aída Hassón-Voloch, Instituto de Biofísica Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; .,Centro Nacional de Biologia Estrutural e Bioimagem, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; and
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35
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Cassier E, Gallay N, Bourquard T, Claeysen S, Bockaert J, Crépieux P, Poupon A, Reiter E, Marin P, Vandermoere F. Phosphorylation of β-arrestin2 at Thr 383 by MEK underlies β-arrestin-dependent activation of Erk1/2 by GPCRs. eLife 2017; 6. [PMID: 28169830 PMCID: PMC5325621 DOI: 10.7554/elife.23777] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2016] [Accepted: 02/06/2017] [Indexed: 01/14/2023] Open
Abstract
In addition to their role in desensitization and internalization of G protein-coupled receptors (GPCRs), β-arrestins are essential scaffolds linking GPCRs to Erk1/2 signaling. However, their role in GPCR-operated Erk1/2 activation differs between GPCRs and the underlying mechanism remains poorly characterized. Here, we show that activation of serotonin 5-HT2C receptors, which engage Erk1/2 pathway via a β-arrestin-dependent mechanism, promotes MEK-dependent β-arrestin2 phosphorylation at Thr383, a necessary step for Erk recruitment to the receptor/β-arrestin complex and Erk activation. Likewise, Thr383 phosphorylation is involved in β-arrestin-dependent Erk1/2 stimulation elicited by other GPCRs such as β2-adrenergic, FSH and CXCR4 receptors, but does not affect the β-arrestin-independent Erk1/2 activation by 5-HT4 receptor. Collectively, these data show that β-arrestin2 phosphorylation at Thr383 underlies β-arrestin-dependent Erk1/2 activation by GPCRs. DOI:http://dx.doi.org/10.7554/eLife.23777.001
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Affiliation(s)
- Elisabeth Cassier
- CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France.,INSERM, U1191, Montpellier, France.,Université de Montpellier, Montpellier, France
| | - Nathalie Gallay
- INRA, UMR85, Unité Physiologie de la Reproduction et des Comportements, Nouzilly, France.,CNRS, UMR7247, Nouzilly, France.,Université François Rabelais, Tours, France
| | - Thomas Bourquard
- INRA, UMR85, Unité Physiologie de la Reproduction et des Comportements, Nouzilly, France.,CNRS, UMR7247, Nouzilly, France.,Université François Rabelais, Tours, France
| | - Sylvie Claeysen
- CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France.,INSERM, U1191, Montpellier, France.,Université de Montpellier, Montpellier, France
| | - Joël Bockaert
- CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France.,INSERM, U1191, Montpellier, France.,Université de Montpellier, Montpellier, France
| | - Pascale Crépieux
- INRA, UMR85, Unité Physiologie de la Reproduction et des Comportements, Nouzilly, France.,CNRS, UMR7247, Nouzilly, France.,Université François Rabelais, Tours, France
| | - Anne Poupon
- INRA, UMR85, Unité Physiologie de la Reproduction et des Comportements, Nouzilly, France.,CNRS, UMR7247, Nouzilly, France.,Université François Rabelais, Tours, France
| | - Eric Reiter
- INRA, UMR85, Unité Physiologie de la Reproduction et des Comportements, Nouzilly, France.,CNRS, UMR7247, Nouzilly, France.,Université François Rabelais, Tours, France
| | - Philippe Marin
- CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France.,INSERM, U1191, Montpellier, France.,Université de Montpellier, Montpellier, France
| | - Franck Vandermoere
- CNRS, UMR-5203, Institut de Génomique Fonctionnelle, Montpellier, France.,INSERM, U1191, Montpellier, France.,Université de Montpellier, Montpellier, France
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36
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Dela Paz NG, Melchior B, Frangos JA. Shear stress induces Gα q/11 activation independently of G protein-coupled receptor activation in endothelial cells. Am J Physiol Cell Physiol 2017; 312:C428-C437. [PMID: 28148497 DOI: 10.1152/ajpcell.00148.2016] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2016] [Revised: 01/25/2017] [Accepted: 01/25/2017] [Indexed: 12/17/2022]
Abstract
Mechanochemical signal transduction occurs when mechanical forces, such as fluid shear stress, are converted into biochemical responses within the cell. The molecular mechanisms by which endothelial cells (ECs) sense/transduce shear stress into biological signals, including the nature of the mechanosensor, are still unclear. G proteins and G protein-coupled receptors (GPCRs) have been postulated independently to mediate mechanotransduction. In this study, we used in situ proximity ligation assay (PLA) to investigate the role of a specific GPCR/Gαq/11 pair in EC shear stress-induced mechanotransduction. We demonstrated that sphingosine 1-phosphate (S1P) stimulation causes a rapid dissociation at 0.5 min of Gαq/11 from its receptor S1P3, followed by an increased association within 2 min of GPCR kinase-2 (GRK2) and β-arrestin-1/2 with S1P3 in human coronary artery ECs, which are consistent with GPCR/Gαq/11 activation and receptor desensitization/internalization. The G protein activator AlF4 resulted in increased dissociation of Gαq/11 from S1P3, but no increase in association between S1P3 and either GRK2 or β-arrestin-1/2. The G protein inhibitor guanosine 5'-(β-thio) diphosphate (GDP-β-S) and the S1P3 antagonist VPC23019 both prevented S1P-induced activation. Shear stress also caused the rapid activation within 7 s of S1P3/Gαq/11 There were no increased associations between S1P3 and GRK2 or S1P3 and β-arrestin-1/2 until 5 min. GDP-β-S, but not VPC23019, prevented dissociation of Gαq/11 from S1P3 in response to shear stress. Shear stress did not induce rapid dephosphorylation of β-arrestin-1 or rapid internalization of S1P3, indicating no GPCR activation. These findings suggest that Gαq/11 participates in the sensing/transducing of shear stress independently of GPCR activation in ECs.
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Sensoy O, Moreira IS, Morra G. Understanding the Differential Selectivity of Arrestins toward the Phosphorylation State of the Receptor. ACS Chem Neurosci 2016; 7:1212-24. [PMID: 27405242 DOI: 10.1021/acschemneuro.6b00073] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Proteins in the arrestin family exhibit a conserved structural fold that nevertheless allows for significant differences in their selectivity for G-protein coupled receptors (GPCRs) and their phosphorylation states. To reveal the mechanism of activation that prepares arrestin for selective interaction with GPCRs, and to understand the basis for these differences, we used unbiased molecular dynamics simulations to compare the structural and dynamic properties of wild type Arr1 (Arr1-WT), Arr3 (Arr3-WT), and a constitutively active Arr1 mutant, Arr1-R175E, characterized by a perturbation of the phosphate recognition region called "polar core". We find that in our simulations the mutant evolves toward a conformation that resembles the known preactivated structures of an Arr1 splice-variant, and the structurally similar phosphopeptide-bound Arr2-WT, while this does not happen for Arr1-WT. Hence, we propose an activation allosteric mechanism connecting the perturbation of the polar core to a global conformational change, including the relative reorientation of N- and C-domains, and the emergence of electrostatic properties of putative binding surfaces. The underlying local structural changes are interpreted as markers of the evolution of an arrestin structure toward an active-like conformation. Similar activation related changes occur in Arr3-WT in the absence of any perturbation of the polar core, suggesting that this system could spontaneously visit preactivated states in solution. This hypothesis is proposed to explain the lower selectivity of Arr3 toward nonphosphorylated receptors. Moreover, by elucidating the allosteric mechanism underlying activation, we identify functionally critical regions on arrestin structure that can be targeted with drugs or chemical tools for functional modulation.
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Affiliation(s)
- Ozge Sensoy
- The School of Engineering and Natural Sciences, Istanbul Medipol University, 34810 Istanbul, Turkey
- Department of Physiology and Biophysics, Weill Cornell Medical College, 1300 York Ave, New York, New York 10065, United States
| | - Irina S. Moreira
- CNC - Center for Neuroscience and Cell Biology; Rua Larga, FMUC, Polo I, 1°andar, Universidade de Coimbra, 3004-517 Coimbra, Portugal
- Bijvoet Center for Biomolecular Research,
Faculty of Science - Chemistry, Utrecht University, Utrecht 3584CH, The Netherlands
| | - Giulia Morra
- Department of Physiology and Biophysics, Weill Cornell Medical College, 1300 York Ave, New York, New York 10065, United States
- ICRM-CNR
Istituto di Chimica del Riconoscimento Molecolare, Consiglio Nazionale delle Ricerche, Via Mario Bianco 9, 20131 Milano, Italia
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38
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Gendron L, Cahill CM, von Zastrow M, Schiller PW, Pineyro G. Molecular Pharmacology of δ-Opioid Receptors. Pharmacol Rev 2016; 68:631-700. [PMID: 27343248 PMCID: PMC4931872 DOI: 10.1124/pr.114.008979] [Citation(s) in RCA: 89] [Impact Index Per Article: 9.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
Opioids are among the most effective analgesics available and are the first choice in the treatment of acute severe pain. However, partial efficacy, a tendency to produce tolerance, and a host of ill-tolerated side effects make clinically available opioids less effective in the management of chronic pain syndromes. Given that most therapeutic opioids produce their actions via µ-opioid receptors (MOPrs), other targets are constantly being explored, among which δ-opioid receptors (DOPrs) are being increasingly considered as promising alternatives. This review addresses DOPrs from the perspective of cellular and molecular determinants of their pharmacological diversity. Thus, DOPr ligands are examined in terms of structural and functional variety, DOPrs' capacity to engage a multiplicity of canonical and noncanonical G protein-dependent responses is surveyed, and evidence supporting ligand-specific signaling and regulation is analyzed. Pharmacological DOPr subtypes are examined in light of the ability of DOPr to organize into multimeric arrays and to adopt multiple active conformations as well as differences in ligand kinetics. Current knowledge on DOPr targeting to the membrane is examined as a means of understanding how these receptors are especially active in chronic pain management. Insight into cellular and molecular mechanisms of pharmacological diversity should guide the rational design of more effective, longer-lasting, and better-tolerated opioid analgesics for chronic pain management.
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Affiliation(s)
- Louis Gendron
- Département de Pharmacologie-Physiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Centre de Recherche du CHU de Sherbrooke, Centre d'excellence en neurosciences de l'Univeristé de Sherbrooke, and Institut de Pharmacologie de Sherbrooke, Sherbrooke, Quebec, Canada (L.G.); Québec Pain Research Network, Sherbrooke, Quebec, Canada (L.G.); Departments of Anesthesiology and Perioperative Care and Pharmacology, University of California, Irvine, California (C.M.C.); Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada (C.M.C.); Departments of Psychiatry and Cellular and Molecular Pharmacology, University of California, San Francisco, California (M.v.Z.); Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montréal, Montreal, Quebec, Canada (P.W.S.); and Departments of Psychiatry, Pharmacology, and Neurosciences, Faculty of Medicine, University of Montréal and Sainte-Justine Hospital Research Center, Montreal, Quebec, Canada (G.P.)
| | - Catherine M Cahill
- Département de Pharmacologie-Physiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Centre de Recherche du CHU de Sherbrooke, Centre d'excellence en neurosciences de l'Univeristé de Sherbrooke, and Institut de Pharmacologie de Sherbrooke, Sherbrooke, Quebec, Canada (L.G.); Québec Pain Research Network, Sherbrooke, Quebec, Canada (L.G.); Departments of Anesthesiology and Perioperative Care and Pharmacology, University of California, Irvine, California (C.M.C.); Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada (C.M.C.); Departments of Psychiatry and Cellular and Molecular Pharmacology, University of California, San Francisco, California (M.v.Z.); Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montréal, Montreal, Quebec, Canada (P.W.S.); and Departments of Psychiatry, Pharmacology, and Neurosciences, Faculty of Medicine, University of Montréal and Sainte-Justine Hospital Research Center, Montreal, Quebec, Canada (G.P.)
| | - Mark von Zastrow
- Département de Pharmacologie-Physiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Centre de Recherche du CHU de Sherbrooke, Centre d'excellence en neurosciences de l'Univeristé de Sherbrooke, and Institut de Pharmacologie de Sherbrooke, Sherbrooke, Quebec, Canada (L.G.); Québec Pain Research Network, Sherbrooke, Quebec, Canada (L.G.); Departments of Anesthesiology and Perioperative Care and Pharmacology, University of California, Irvine, California (C.M.C.); Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada (C.M.C.); Departments of Psychiatry and Cellular and Molecular Pharmacology, University of California, San Francisco, California (M.v.Z.); Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montréal, Montreal, Quebec, Canada (P.W.S.); and Departments of Psychiatry, Pharmacology, and Neurosciences, Faculty of Medicine, University of Montréal and Sainte-Justine Hospital Research Center, Montreal, Quebec, Canada (G.P.)
| | - Peter W Schiller
- Département de Pharmacologie-Physiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Centre de Recherche du CHU de Sherbrooke, Centre d'excellence en neurosciences de l'Univeristé de Sherbrooke, and Institut de Pharmacologie de Sherbrooke, Sherbrooke, Quebec, Canada (L.G.); Québec Pain Research Network, Sherbrooke, Quebec, Canada (L.G.); Departments of Anesthesiology and Perioperative Care and Pharmacology, University of California, Irvine, California (C.M.C.); Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada (C.M.C.); Departments of Psychiatry and Cellular and Molecular Pharmacology, University of California, San Francisco, California (M.v.Z.); Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montréal, Montreal, Quebec, Canada (P.W.S.); and Departments of Psychiatry, Pharmacology, and Neurosciences, Faculty of Medicine, University of Montréal and Sainte-Justine Hospital Research Center, Montreal, Quebec, Canada (G.P.)
| | - Graciela Pineyro
- Département de Pharmacologie-Physiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Centre de Recherche du CHU de Sherbrooke, Centre d'excellence en neurosciences de l'Univeristé de Sherbrooke, and Institut de Pharmacologie de Sherbrooke, Sherbrooke, Quebec, Canada (L.G.); Québec Pain Research Network, Sherbrooke, Quebec, Canada (L.G.); Departments of Anesthesiology and Perioperative Care and Pharmacology, University of California, Irvine, California (C.M.C.); Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada (C.M.C.); Departments of Psychiatry and Cellular and Molecular Pharmacology, University of California, San Francisco, California (M.v.Z.); Laboratory of Chemical Biology and Peptide Research, Clinical Research Institute of Montréal, Montreal, Quebec, Canada (P.W.S.); and Departments of Psychiatry, Pharmacology, and Neurosciences, Faculty of Medicine, University of Montréal and Sainte-Justine Hospital Research Center, Montreal, Quebec, Canada (G.P.)
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39
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Smith JS, Rajagopal S. The β-Arrestins: Multifunctional Regulators of G Protein-coupled Receptors. J Biol Chem 2016; 291:8969-77. [PMID: 26984408 DOI: 10.1074/jbc.r115.713313] [Citation(s) in RCA: 233] [Impact Index Per Article: 25.9] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
The β-arrestins (βarrs) are versatile, multifunctional adapter proteins that are best known for their ability to desensitize G protein-coupled receptors (GPCRs), but also regulate a diverse array of cellular functions. To signal in such a complex fashion, βarrs adopt multiple conformations and are regulated at multiple levels to differentially activate downstream pathways. Recent structural studies have demonstrated that βarrs have a conserved structure and activation mechanism, with plasticity of their structural fold, allowing them to adopt a wide array of conformations. Novel roles for βarrs continue to be identified, demonstrating the importance of these dynamic regulators of cellular signaling.
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Affiliation(s)
| | - Sudarshan Rajagopal
- From the Departments of Biochemistry and Medicine, Duke University Medical Center, Durham, North Carolina 27710
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40
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TRIP6 antagonizes the recruitment of A20 and CYLD to TRAF6 to promote the LPA2 receptor-mediated TRAF6 activation. Cell Discov 2016; 2. [PMID: 27134758 PMCID: PMC4850058 DOI: 10.1038/celldisc.2015.48] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
The elevated lysophosphatidic acid signaling has been causally linked to cancer-associated inflammation and tumorigenesis through upregulation of nuclear factor-κB signaling. However, how this signaling event is regulated has not yet been fully understood. Here we demonstrate that TRIP6, an LPA2 receptor-interacting adaptor protein, functions as a positive regulator of nuclear factor-κB and JNK signaling through direct binding to and activation of the E3 ligase TRAF6. Upon lysophosphatidic acid stimulation, TRIP6 recruits TRAF6 to the LPA2 receptor and promotes lysophosphatidic acid-induced JNK and nuclear factor-κB activation in a TRAF6-dependent manner. TRIP6 antagonizes the recruitment of deubiquitinases A20 and CYLD to TRAF6, thus sustaining the E3 ligase activity of TRAF6 and augmenting lysophosphatidic acid-activated nuclear factor-κB signaling. In contrast, depletion of TRIP6 by TRIP6-specific shRNA or Cas9/sgRNA greatly enhances the association of TRAF6 with A20 and CYLD, and attenuates lysophosphatidic acid-induced muclear factor-κB and JNK/p38 activation in ovarian cancer cells. On the other hand, TRAF6 also regulates TRIP6 by facilitating its binding to nuclear factor-κB p65 and phosphorylation by c-Src. Together, TRIP6 cooperates with TRAF6 to regulate the LPA2 receptor signaling, which may ultimately contribute to chronic inflammation, apoptotic resistance and cell invasion.
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41
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Sharma D, Parameswaran N. Multifaceted role of β-arrestins in inflammation and disease. Genes Immun 2015; 16:499-513. [PMID: 26378652 PMCID: PMC4670277 DOI: 10.1038/gene.2015.37] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2015] [Revised: 07/05/2015] [Accepted: 07/31/2015] [Indexed: 12/19/2022]
Abstract
Arrestins are intracellular scaffolding proteins known to regulate a range of biochemical processes including G protein-coupled receptor (GPCR) desensitization, signal attenuation, receptor turnover and downstream signaling cascades. Their roles in regulation of signaling network have lately been extended to receptors outside of the GPCR family, demonstrating their roles as important scaffolding proteins in various physiological processes including proliferation, differentiation and apoptosis. Recent studies have demonstrated a critical role for arrestins in immunological processes including key functions in inflammatory signaling pathways. In this review, we provide a comprehensive analysis of the different functions of the arrestin family of proteins especially related to immunity and inflammatory diseases.
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Affiliation(s)
- Deepika Sharma
- Department of Physiology and Division of Pathology Michigan State University East Lansing, MI 48824
| | - Narayanan Parameswaran
- Department of Physiology and Division of Pathology Michigan State University East Lansing, MI 48824
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42
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The emerging roles of β-arrestins in fibrotic diseases. Acta Pharmacol Sin 2015; 36:1277-87. [PMID: 26388156 DOI: 10.1038/aps.2015.74] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/03/2015] [Accepted: 06/29/2015] [Indexed: 02/06/2023]
Abstract
β-Arrestins and β-arrestin2 are important adaptor proteins and signal transduction proteins that are mainly involved in the desensitization and internalization of G-protein-coupled receptors. Fibrosis is characterized by accumulation of excess extracellular matrix (ECM) molecules caused by chronic tissue injury. If highly progressive, the fibrotic process leads to organ malfunction and, eventually, death. The incurable lung fibrosis, renal fibrosis and liver fibrosis are among the most common fibrotic diseases. Recent studies show that β-arrestins can activate signaling cascades independent of G-protein activation and scaffold many intracellular signaling networks by diverse types of signaling pathways, including the Hedgehog, Wnt, Notch and transforming growth factor-β pathways, as well as downstream kinases such as MAPK and PI3K. These signaling pathways are involved in the pathological process of fibrosis and fibrotic diseases. This β-arrestin-mediated regulation not only affects cell growth and apoptosis, but also the deposition of ECM, activation of inflammatory response and development of fibrotic diseases. In this review, we survey the involvement of β-arrestins in various signaling pathways and highlight different aspects of their regulation of fibrosis. We also discuss the important roles of β-arrestins in the process of fibrotic diseases by regulating the inflammation and deposit of ECM. It is becoming more evident that targeting β-arrestins may offer therapeutic potential for the treatment of fibrotic diseases.
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43
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Robinson KA, Ou WL, Guan X, Sugamori KS, Bandyopadhyay A, Ernst OP, Mitchell J. The effect of phosphorylation on arrestin-rhodopsin interaction in the squid visual system. J Neurochem 2015; 135:1129-39. [PMID: 26375013 DOI: 10.1111/jnc.13366] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2015] [Revised: 08/07/2015] [Accepted: 09/02/2015] [Indexed: 11/30/2022]
Abstract
Invertebrate visual opsins are G protein-coupled receptors coupled to retinoid chromophores that isomerize reversibly between inactive rhodopsin and active metarhodopsin upon absorption of photons of light. The squid visual system has an arrestin protein that binds to metarhodopsin to block signaling to Gq and activation of phospholipase C. Squid rhodopsin kinase (SQRK) can phosphorylate both metarhodopsin and arrestin, a dual role that is unique among the G protein-coupled receptor kinases. The sites and role of arrestin phosphorylation by SQRK were investigated here using recombinant proteins. Arrestin was phosphorylated on serine 392 and serine 397 in the C-terminus. Unphosphorylated arrestin bound to metarhodopsin and phosphorylated metarhodopsin with similar high affinities (Kd 33 and 21 nM respectively), while phosphorylation of arrestin reduced the affinity 3- to 5-fold (Kd 104 nM). Phosphorylation of metarhodopsin slightly increased the dissociation of arrestin observed during a 1 hour incubation. Together these studies suggest a unique role for SQRK in phosphorylating both receptor and arrestin and inhibiting the binding of these two proteins in the squid visual system. Invertebrate visual systems are inactivated by arrestin binding to metarhodopsin that does not require receptor phosphorylation. Here we show that squid rhodopsin kinase phosphorylates arrestin on two serines (S392,S397) in the C-terminus and phosphorylation decreases the affinity of arrestin for squid metarhodopsin. Metarhodopsin phosphorylation has very little effect on arrestin binding but does increase arrestin dissociation.
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Affiliation(s)
- Kelly A Robinson
- Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario, Canada
| | - Wei-Lin Ou
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada
| | - Xinyu Guan
- Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario, Canada
| | - Kim S Sugamori
- Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario, Canada
| | | | - Oliver P Ernst
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.,Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada
| | - Jane Mitchell
- Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario, Canada
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44
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Trafficking of β-Adrenergic Receptors: Implications in Intracellular Receptor Signaling. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2015; 132:151-88. [PMID: 26055058 DOI: 10.1016/bs.pmbts.2015.03.008] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
β-Adrenergic receptors (βARs), prototypical G-protein-coupled receptors, play a pivotal role in regulating neuronal and cardiovascular responses to catecholamines during stress. Agonist-induced receptor endocytosis is traditionally considered as a primary mechanism to turn off the receptor signaling (or receptor desensitization). However, recent progress suggests that intracellular trafficking of βAR presents a mean to translocate receptor signaling machinery to intracellular organelles/compartments while terminating the signaling at the cell surface. Moreover, the apparent multidimensionality of ligand efficacy in space and time in a cell has forecasted exciting pathophysiological implications, which are just beginning to be explored. As we begin to understand how these pathways impact downstream cellular programs, this will have significant implications for a number of pathophysiological conditions in heart and other systems, that in turn open up new therapeutic opportunities.
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45
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Crudden C, Ilic M, Suleymanova N, Worrall C, Girnita A, Girnita L. The dichotomy of the Insulin-like growth factor 1 receptor: RTK and GPCR: friend or foe for cancer treatment? Growth Horm IGF Res 2015; 25:2-12. [PMID: 25466906 DOI: 10.1016/j.ghir.2014.10.002] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/14/2014] [Revised: 10/06/2014] [Accepted: 10/20/2014] [Indexed: 01/14/2023]
Abstract
The prime position of the insulin-like growth factor 1 receptor (IGF-1R), at the head of the principle mitogenic and anti-apoptotic signalling cascades, along with the resilience to transformation of IGF-1R deficient cells fuelled great excitement for its anti-cancer targeting. Yet its potential has not been fulfilled, as clinical trial results fell far short of expectations. Advancements in understanding of other receptors' function have now begun to shed light on this incongruity, with the now apparent parallels highlighting the immaturity of our understanding of IGF-1R biology, with the model used for drug development now recognised as having been too simplistic. Gathering together the many advancements of the field of IGF-1R research over the past decade, alongside those in the GPCR field, advocates for a major paradigm shift in our appreciation of the subtle workings of this receptor. This review will emphasise the updating of the IGF-1R's classification from an RTK, to an RTK/GPCR functional hybrid, which integrates both canonical kinase signalling with many functions characteristic of a GPCR. Recognition of the shortcomings of IGF-1R inhibitor drug development programs and the models used not only allows us to reignite the initial interest in the IGF-1R as an anti-cancer therapeutic target, but also points to the possibility of biased ligand therapeutics, which together may hold a very powerful key to unlocking the true potential of IGF-1R modulation.
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Affiliation(s)
- Caitrin Crudden
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden
| | - Marina Ilic
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden
| | - Naida Suleymanova
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden
| | - Claire Worrall
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden
| | - Ada Girnita
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden; Dermatology Department, Karolinska University Hospital, Stockholm, Sweden
| | - Leonard Girnita
- Department of Oncology and Pathology, Cancer Center Karolinska, Karolinska Institutet and Karolinska University Hospital, 17176 Stockholm, Sweden.
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46
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Gurevich VV, Gurevich EV. Analyzing the roles of multi-functional proteins in cells: The case of arrestins and GRKs. Crit Rev Biochem Mol Biol 2015; 50:440-452. [PMID: 26453028 PMCID: PMC4852696 DOI: 10.3109/10409238.2015.1067185] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
Abstract
Most proteins have multiple functions. Obviously, conventional methods of manipulating the level of the protein of interest in the cell, such as over-expression, knockout or knockdown, affect all of its functions simultaneously. The key advantage of these methods is that over-expression, knockout or knockdown does not require any knowledge of the molecular mechanisms of the function(s) of the protein of interest. The disadvantage is that these approaches are inadequate to elucidate the role of an individual function of the protein in a particular cellular process. An alternative is the use of re-engineered proteins, in which a single function is eliminated or enhanced. The use of mono-functional elements of a multi-functional protein can also yield cleaner answers. This approach requires detailed knowledge of the structural basis of each function of the protein in question. Thus, a lot of preliminary structure-function work is necessary to make it possible. However, when this information is available, replacing the protein of interest with a mutant in which individual functions are modified can shed light on the biological role of those particular functions. Here, we illustrate this point using the example of protein kinases, most of which have additional non-enzymatic functions, as well as arrestins, known multi-functional signaling regulators in the cell.
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Affiliation(s)
| | - Eugenia V Gurevich
- a Department of Pharmacology , Vanderbilt University , Nashville , TN , USA
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47
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Khoury E, Nikolajev L, Simaan M, Namkung Y, Laporte SA. Differential regulation of endosomal GPCR/β-arrestin complexes and trafficking by MAPK. J Biol Chem 2014; 289:23302-17. [PMID: 25016018 DOI: 10.1074/jbc.m114.568147] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
β-Arrestins are signaling adaptors that bind to agonist-occupied G protein-coupled receptors (GPCRs) and target them for endocytosis; however, the mechanisms regulating receptor/β-arrestin complexes and trafficking in endosomes, remain ill defined. Here we show, in live cells, differential dynamic regulation of endosomal bradykinin B2 receptor (B2R) complexes with either β-arrestin-1 or -2. We find a novel role for MAPK in the B2R/β-arrestin-2 complex formation, receptor trafficking and signaling mediated by an ERK1/2 regulatory motif in the hinge domain of the rat β-arrestin-2 (PET(178)P), but not rat β-arrestin-1 (PER(177)P). While the ERK1/2 regulatory motif is conserved between rat and mouse β-arrestin-2, it is surprisingly not conserved in human β-arrestin-2 (PEK(178)P). However, mutation of lysine 178 to threonine is sufficient to confer MAPK sensitivity to the human β-arrestin-2. Furthermore, substitution for a phosphomimetic residue in both the rat and the human β-arrestin-2 (T/K178D) significantly stabilizes B2R/β-arrestin complexes in endosomes, delays receptor recycling to the plasma membrane and maintains intracellular MAPK signaling. Similarly, the endosomal trafficking of β2-adrenergic, angiotensin II type 1 and vasopressin V2 receptors was altered by the β-arrestin-2 T178D mutant. Our findings unveil a novel subtype specific mode of MAPK-dependent regulation of β-arrestins in intracellular trafficking and signaling of GPCRs, and suggest differential endosomal receptor/β-arrestin-2 signaling roles among species.
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Affiliation(s)
| | | | | | | | - Stéphane A Laporte
- From the Departments of Medicine, Pharmacology and Therapeutics, and Anatomy and Cell Biology, McGill University Health Center Research Institute, McGill University, Strathcona Anatomy & Dentistry Bldg., Quebec H3A 2B2, Canada
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48
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Abstract
The four members of the mammalian arrestin family, two visual and two nonvisual, share the property of stimulus-dependent docking to G protein-coupled receptors. This conformational selectivity permits them to function in receptor desensitization, as arrestin binding sterically inhibits G protein coupling. The two nonvisual arrestins further act as adapter proteins, linking receptors to the clathrin-dependent endocytic machinery and regulating receptor sequestration, intracellular trafficking, recycling, and degradation. Arrestins also function as ligand-regulated scaffolds, recruiting catalytically active proteins into receptor-based multiprotein "signalsome" complexes. Arrestin binding thus marks the transition from a transient G protein-coupled state on the plasma membrane to a persistent arrestin-coupled state that continues to signal as the receptor internalizes. Two of the earliest discovered and most studied arrestin-dependent signaling pathways involve regulation of Src family nonreceptor tyrosine kinases and the ERK1/2 mitogen-activated kinase cascade. In each case, arrestin scaffolding imposes constraints on kinase activity that dictate signal duration and substrate specificity. Evidence suggests that arrestin-bound ERK1/2 and Src not only play regulatory roles in receptor desensitization and trafficking but also mediate longer term effects on cell growth, migration, proliferation, and survival.
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Affiliation(s)
- Erik G Strungs
- Department of Medicine, Medical University of South Carolina, Charleston, SC, 29425, USA
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49
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Kang DS, Tian X, Benovic JL. Role of β-arrestins and arrestin domain-containing proteins in G protein-coupled receptor trafficking. Curr Opin Cell Biol 2013; 27:63-71. [PMID: 24680432 DOI: 10.1016/j.ceb.2013.11.005] [Citation(s) in RCA: 152] [Impact Index Per Article: 12.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2013] [Accepted: 11/14/2013] [Indexed: 12/12/2022]
Abstract
The arrestin clan can now be broadly divided into three structurally similar subgroups: the originally identified arrestins (visual and β-arrestins), the α-arrestins and a group of Vps26-related proteins. The visual and β-arrestins selectively bind to agonist-occupied phosphorylated G protein-coupled receptors (GPCRs) and inhibit GPCR coupling to heterotrimeric G proteins while the β-arrestins also function as adaptor proteins to regulate GPCR trafficking and G protein-independent signaling. The α-arrestins have also recently been implicated in regulating GPCR trafficking while Vps26 regulates retrograde trafficking. In this review, we provide an overview of the α-arrestins and β-arrestins with a focus on our current understanding of how these adaptor proteins regulate GPCR trafficking.
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Affiliation(s)
- Dong Soo Kang
- Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA
| | - Xufan Tian
- Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA
| | - Jeffrey L Benovic
- Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA.
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50
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Habourdin C, Klein G, Araki T, Williams JG, Aubry L. The arrestin-domain containing protein AdcA is a response element to stress. Cell Commun Signal 2013; 11:91. [PMID: 24267687 PMCID: PMC3879092 DOI: 10.1186/1478-811x-11-91] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2013] [Accepted: 11/06/2013] [Indexed: 01/01/2023] Open
Abstract
Background Cell behaviour is tightly determined by sensing and integration of extracellular changes through membrane detectors such as receptors and transporters and activation of downstream signalling cascades. Arrestin proteins act as scaffolds at the plasma membrane and along the endocytic pathway, where they regulate the activity and the fate of some of these detectors. Members of the arrestin clan are widely present from unicellular to metazoa, with roles in signal transduction and metabolism. As a soil amoeba, Dictyostelium is frequently confronted with environmental changes likely to compromise survival. Here, we investigated whether the recently described arrestin-related protein AdcA is part of the cell response to stresses. Results Our data provide evidence that AdcA responds to a variety of stresses including hyperosmolarity by a transient phosphorylation. Analysis in different mutant backgrounds revealed that AdcA phosphorylation involves pathways other than the DokA and cGMP-dependent osmostress pathways, respectively known to regulate PKA and STATc, key actors in the cellular response to conditions of hyperosmolarity. Interestingly, however, both AdcA and STATc are sensitive to changes in the F-actin polymerization status, suggesting a common primary sensor/trigger and linking the stress-sensitive kinase responsive for AdcA phosphorylation to the actin cytoskeleton. We also show that STATc-dependent transcriptional activity is involved for the timely dephosphorylation of AdcA in cells under stress. Conclusion Under osmotic stress, AdcA undergoes a phosphorylation-dephosphorylation cycle involving a stress-sensitive kinase and the transcription regulator STATc. This transient post-transcriptional modification may allow a regulation of AdcA function possibly to optimize the cellular stress response.
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Affiliation(s)
| | | | | | | | - Laurence Aubry
- CEA, iRTSV, Laboratoire Biologie à Grande Echelle, F-38054 Grenoble, France.
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