Regulation of L-type CaV1.3 channel activity and insulin secretion by the cGMP-PKG signaling pathway |
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| Institution: | 1. FES Iztacala UNAM, Tlalnepantla De Baz, Mexico;2. Departamento de Biología Celular, Cinvestav-IPN, Ciudad de México, Mexico;3. Hotchkiss Brain Institute, University of Calgary, Calgary, AB, Canada;4. Department of Biological Sciences, University of New Hampshire, Durham, NH, USA;5. Dirección de Investigaciones en Neurociencias, Instituto Nacional de Psiquiatría, Ramón de la Fuente Muñiz, Ciudad de México, Mexico;6. Department of Ophtalmology and Molecular Pharmacology & Therapeutics, Loyola University, Chicago, Strich School of Medicine, Maywood, IL, USA;1. Department of Molecular and Translational Oncology, Cancer Center Institute, Warsaw, Poland;2. Center for Infectious Medicine, Department of Medicine, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden;3. Division of Molecular Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden;3. Departments of Chemistry, University of Puerto Rico, Rio Piedras Campus, San Juan, Puerto Rico, 00931;4. Departments of Biology, University of Puerto Rico, Rio Piedras Campus, San Juan, Puerto Rico, 00931;6. Departments of Physical Sciences, University of Puerto Rico, Rio Piedras Campus, San Juan, Puerto Rico, 00931;5. the California State University Dominguez Hills, Carson, California 90747;1. Department of Molecular Biology, Princeton University, Princeton, NJ, United States;2. Department of Biological and Allied Health Sciences, Fairleigh Dickinson University, Madison, NJ, United States;1. Programa de Pós-graduação em Ciências Farmacêuticas, Departamento de Fármacos e Medicamentos, Faculdade de Ciências Farmacêuticas, Universidade Estadual Paulista ?Júlio de Mesquita Filho?, Araraquara, SP, Brazil;2. Escola de Artes, Ciências e Humanidades, Universidade de São Paulo, São Paulo, SP, Brazil;3. Departamento de Química, Faculdade de Ciências, Universidade Estadual Paulista ?Julio de Mesquita Filho?, Bauru, SP, Brazil;4. Universidade Estadual Paulista ?Júlio de Mesquita Filho?, Campus do Litoral Paulista, Unidade São Vicente, SP, Brazil;5. Fundação Herminio Ometto, Centro Universitário Herminio Ometto, Araras, SP, Brazil;3. M. M. Shemyakin and Yu. A. Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;4. Moscow Institute of Physics and Technology (State University), 117303 Moscow, Russia;5. Toxicology and Pharmacology, University of Leuven, 3000 Leuven, Belgium |
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| Abstract: | cGMP is a second messenger widely used in the nervous system and other tissues. One of the major effectors for cGMP is the serine/threonine protein kinase, cGMP-dependent protein kinase (PKG), which catalyzes the phosphorylation of a variety of proteins including ion channels. Previously, it has been shown that the cGMP-PKG signaling pathway inhibits Ca2+ currents in rat vestibular hair cells and chromaffin cells. This current allegedly flow through voltage-gated CaV1.3L-type Ca2+ channels, and is important for controlling vestibular hair cell sensory function and catecholamine secretion, respectively. Here, we show that native L-type channels in the insulin-secreting RIN-m5F cell line, and recombinant CaV1.3 channels heterologously expressed in HEK-293 cells, are regulatory targets of the cGMP-PKG signaling cascade. Our results indicate that the CaVα1 ion-conducting subunit of the CaV1.3 channels is highly expressed in RIN-m5F cells and that the application of 8-Br-cGMP, a membrane-permeable analogue of cGMP, significantly inhibits Ca2+ macroscopic currents and impair insulin release stimulated with high K+. In addition, KT-5823, a specific inhibitor of PKG, prevents the current inhibition generated by 8-Br-cGMP in the heterologous expression system. Interestingly, mutating the putative phosphorylation sites to residues resistant to phosphorylation showed that the relevant PKG sites for CaV1.3 L-type channel regulation centers on two amino acid residues, Ser793 and Ser860, located in the intracellular loop connecting the II and III repeats of the CaVα1 pore-forming subunit of the channel. These findings unveil a novel mechanism for how the cGMP-PKG signaling pathway may regulate CaV1.3 channels and contribute to regulate insulin secretion. |
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| Keywords: | L-type channels Cav channels PKG cGMP Insulin Rin-m5F cells |
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