Abstract
Human α1D-adrenoceptors are G protein-coupled receptors that mediate adrenaline/noradrenaline actions. There is a growing interest in identifying regulatory domains in these receptors and determining how they function. In this work, we show that the absence of the human α1D-adrenoceptor carboxyl tail results in altered ERK (extracellular signal-regulated kinase) and p38 phosphorylation states. Amino terminus-truncated and both amino and carboxyl termini-truncated α1D-adrenoceptors were transfected into Rat-1, HEK293, and B103 cells, and changes in the phosphorylation state of extracellular signal-regulated kinase was assessed using biochemical and biophysical approaches. The phosphorylation state of other protein kinases (p38, MEK1, and Raf-1) was also studied. Noradrenaline-induced ERK phosphorylation in Rat-1 fibroblasts expressing amino termini-truncated α1D-adrenoceptors. However, in cells expressing receptors with both amino and carboxyl termini truncations, noradrenaline-induced activation was abrogated. Interestingly, ERK phosphorylation that normally occurs through activation of endogenous G protein-coupled receptors, EGF receptors, and protein kinase C, was also decreased, suggesting that downstream steps in the mitogen-activated protein kinase pathway were affected. A similar effect was observed in B103 cells but not in HEK 293 cells. Phosphorylation of Raf-1 and MEK1 was also diminished in Rat-1 fibroblasts expressing amino- and carboxyl-truncated α1D-adrenoceptors. Our data indicate that expression of carboxyl terminus-truncated α1D-adrenoceptors alters ERK and p38 phosphorylation state.
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Acknowledgments
This research was partially supported by grants from CONACyT [177556-253156] and DGAPA-UNAM [200812-200915]. JC was supported by the NIH (NS084398). Marco Antonio Alfonzo is a student of the Programa de Maestría y Doctorado en Ciencias Bioquímicas-UNAM and the recipient of a fellowship from CONACyT; the present work constitutes a major part of his thesis. We express our gratitude to Dr. Rocío Alcántara-Hernández, Dr. Claudia Rivera, Dr. Héctor Malagón, Juan Barbosa, Aurey Galván, and Manuel Ortínez for technical help and advice.
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Jean A. Castillo-Badillo is currently affiliated at Research Support Network, Instituto Nacional de Ciencias Médicas y Nutrición “Salvador Zubirán”, Universidad Nacional Autónoma deMéxico (CIC-UNAM),Vasco de Quiroga 15, 14000 México, D.F, Mexico
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Alfonzo-Méndez, M.A., Castillo-Badillo, J.A., Romero-Ávila, M.T. et al. Carboxyl terminus-truncated α1D-adrenoceptors inhibit the ERK pathway. Naunyn-Schmiedeberg's Arch Pharmacol 389, 911–920 (2016). https://doi.org/10.1007/s00210-016-1254-2
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DOI: https://doi.org/10.1007/s00210-016-1254-2