The N-terminal region of the dopamine D2 receptor, a rhodopsin-like GPCR, regulates correct integration into the plasma membrane and endocytic routes |
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Authors: | Cho D I Min C Jung K S Cheong S Y Zheng M Cheong S J Oak M H Cheong J H Lee B K Kim K M |
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Affiliation: | Department of Pharmacology, College of Pharmacy, Research Institute of Drug Development, Chonnam National University, Gwang-Ju, 500-757 Korea. |
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Abstract: | BACKGROUND AND PURPOSEFunctional roles of the N-terminal region of rhodopsin-like GPCR family remain unclear. Using dopamine D2 and D3 receptors as a model system, we probed the roles of the N-terminal region in the signalling, intracellular trafficking of receptor proteins, and explored the critical factors that determine the functionality of the N-terminal region.EXPERIMENTAL APPROACHThe N-terminal region of the D2 receptor was gradually shortened or switched with that of the D3 receptor or a non-specific sequence (FLAG), or potential N-terminal glycosylation sites were mutated. Effects of these manipulations on surface expression, internalization, post-endocytic behaviours and signalling were determined.KEY RESULTSShortening the N-terminal region of the D2 receptor enhanced receptor internalization and impaired surface expression and signalling; ligand binding, desensitization and down-regulation were not affected but their association with a particular microdomain, caveolae, was disrupted. Replacement of critical residues within the N-terminal region with the FLAG epitope failed to restore surface expression but partially restored the altered internalization and signalling. When the N-terminal regions were switched between D2 and D3 receptors, cell surface expression pattern of each receptor was switched. Mutations of potential N-terminal glycosylation sites inhibited surface expression but enhanced internalization of D2 receptors.CONCLUSIONS AND IMPLICATIONSShortening of N-terminus or mutation of glycosylation sites located within the N-terminus enhanced receptor internalization but impaired the surface expression of D2 receptors. The N-terminal region of the D2 receptor, in a sequence-specific manner, controls the receptor''s conformation and integration into the plasma membrane, which determine its subcellular localization, intracellular trafficking and signalling properties. |
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Keywords: | GPCR dopamine receptors N-terminal region intracellular trafficking cell surface expression glycosylation |
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