Role of cysteine residues in the carboxyl-terminus of the follicle-stimulating hormone receptor in intracellular traffic and postendocytic processing - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Frontiers in Cell and Developmental Biology Année : 2016

Role of cysteine residues in the carboxyl-terminus of the follicle-stimulating hormone receptor in intracellular traffic and postendocytic processing

Résumé

Posttranslational modifications occurring during the biosynthesis of G protein-coupled receptors include glycosylation and palmitoylation at conserved cysteine residues located in the carboxyl-terminus of the receptor. In a number of these receptors, these modifications play an important role in receptor function and particularly, in intracellular trafficking. In the present study, the three cysteine residues present in the carboxyl-terminus of the human FSHR were replaced with glycine by site-directed mutagenesis. Wild-type and mutant (Cys627/629/655Gly) FSHRs were then transiently expressed in HEK-293 cells and analyzed for cell-surface plasma membrane expression, agonist-stimulated signaling and internalization, and postendocytic processing in the absence and presence of lysosome and/or proteasome inhibitors. Compared with the wild-type FSHR, the triple mutant FSHR exhibited ~70% reduction in plasma membrane expression as well as a profound attenuation in agonist-stimulated cAMP production and ERK1/2 phosphorylation. Incubation of HEK-293 cells expressing the wild-type FSHR with 2-bromopalmitate (palmitoylation inhibitor) for 6 h, decreased plasma membrane expression of the receptor by ~30%. The internalization kinetics and β-arrestin 1 and 2 recruitment were similar between the wild-type and triple mutant FSHR as disclosed by assays performed in non-equilibrium binding conditions and by confocal microscopy. Cells expressing the mutant FSHR recycled the internalized FSHR back to the plasma membrane less efficiently than those expressing the wild-type FSHR, an effect that was counteracted by proteasome but not by lysosome inhibition. These results indicate that replacement of the cysteine residues present in the carboxyl-terminus of the FSHR, impairs receptor trafficking from the endoplasmic reticulum/Golgi apparatus to the plasma membrane and its recycling from endosomes back to the cell surface following agonist-induced internalization. Since in the FSHR these cysteine residues are S-palmitoylated, the data presented emphasize on this posttranslational modification as an important factor for both upward and downward trafficking of this receptor.
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Dates et versions

hal-01595693 , version 1 (26-09-2017)

Identifiants

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Brenda Melo-Nava, Patricia Casas-González, Marco A Pérez-Solís, Jean Castillo-Badillo, José L. Maravillas-Montero, et al.. Role of cysteine residues in the carboxyl-terminus of the follicle-stimulating hormone receptor in intracellular traffic and postendocytic processing. Frontiers in Cell and Developmental Biology, 2016, 4, pp.1-13. ⟨10.3389/fcell.2016.00076⟩. ⟨hal-01595693⟩
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