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Título

M3 muscarinic acetylcholine receptor facilitates the endocytosis of mu opioid receptor mediated by morphine independently of the formation of heteromeric complexes

AutorLópez-Giménez, Juan F. ; Alvarez-Curto, Elisa; Milligan, Graeme
Palabras claveHeteromerization
M3 receptor
MOP receptor
GPCR
Morphine
Endocytosis
Fecha de publicación2017
EditorElsevier
CitaciónCellular Signalling 35: 208-222 (2017)
ResumenMorphine inefficiency to induce the internalization of mu opioid (MOP) receptors observed in numerous experimental models constitutes a paradigm of G-protein coupled receptor (GPCR) functional selectivity. We recently described that activation of G proteins through 5-HT serotonin receptors co-expressed in the same cells facilitates MOP receptor endocytosis promoted by morphine. In order to explore whether a different G coupled GPCR would emulate this effect, a double stable Flp-In T-REx HEK293 cell line permanently expressing MOP-YFP receptors along with FLAG-M3-Cerulean receptors expressed in an inducible manner was generated. Fluorescence microscopy examination of these cells revealed a co-distribution of both receptors mainly compartmentalized in plasma membrane. Concurrent stimulation with carbachol and morphine promoted MOP receptor internalization, desensitization and down-regulation and this facilitation was not dependent on PKC activation. Co-immunoprecipitation experiments demonstrated that FLAG-M3-Cerulean/MOP-YFP receptors interact forming heteromeric complexes in a time depending manner, i.e. the strongest interaction was detected after 96 h of FLAG-M3-Cerulean induced expression. Under these experimental conditions, treatment of cells with carbachol plus morphine resulted in the internalization of both receptors within separated endocytic vesicles as visualized by confocal microscopy. This trafficking segregation observed for FLAG-M3-Cerulean and MOP-YFP receptors upon agonist stimulation suggests that this protein-protein interaction presents temporal and dynamic properties. Moreover, MOP-YFP receptor internalization facilitated by FLAG-M3-Cerulean receptors is independent of the constitution of heteromeric complexes.
Versión del editorhttps://doi.org/10.1016/j.cellsig.2017.04.006
URIhttp://hdl.handle.net/10261/164564
Identificadoresdoi: 10.1016/j.cellsig.2017.04.006
e-issn: 1873-3913
issn: 0898-6568
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