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ARF6 and GASP ‐1 are post‐endocytic sorting proteins selectively involved in the intracellular trafficking of dopamine D 2 receptors mediated by GRK and PKC in transfected cells
Author(s) -
Cho DI,
Zheng M,
Min C,
Kwon KJ,
Shin CY,
Choi HK,
Kim KM
Publication year - 2013
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/bph.12025
Subject(s) - internalization , g protein coupled receptor , microbiology and biotechnology , receptor , endocytic cycle , intracellular , biology , protein kinase c , homologous desensitization , hek 293 cells , endosome , phosphorylation , signal transduction , chemistry , agonist , endocytosis , biochemistry
Background and Purpose GPCRs undergo both homologous and heterologous regulatory processes in which receptor phosphorylation plays a critical role. The protein kinases responsible for each pathway are well established; however, other molecular details that characterize each pathway remain unclear. In this study, the molecular mechanisms that determine the differences in the functional roles and intracellular trafficking between homologous and PKC ‐mediated heterologous internalization pathways for the dopamine D 2 receptor were investigated. Experimental Approach All of the S /T residues located within the intracellular loops of D 2 receptor were mutated, and the residues responsible for GRK ‐ and PKC ‐mediated internalization were determined in HEK ‐293 cells and SH‐SY5Y cells. The functional role of receptor internalization and the cellular components that determine the post‐endocytic fate of internalized D 2 receptors were investigated in the transfected cells. Key Results T 134, T 225/ S 228/ S 229 and S 325 were involved in PKC ‐mediated D 2 receptor desensitization. S 229 and adjacent S/T residues mediated the PKC ‐dependent internalization of D 2 receptors, which induced down‐regulation and desensitization. S/T residues within the second intracellular loop and T 225 were the major residues involved in GRK ‐mediated internalization of D 2 receptors, which induced receptor resensitization. ARF6 mediated the recycling of D 2 receptors internalized in response to agonist stimulation. In contrast, GASP ‐1 mediated the down‐regulation of D 2 receptors internalized in a PKC ‐dependent manner. Conclusions and Implications GRK ‐ and PKC ‐mediated internalizations of D 2 receptors occur through different intracellular trafficking pathways and mediate distinct functional roles. Distinct S/T residues within D 2 receptors and different sorting proteins are involved in the dissimilar regulation of D 2 receptors by GRK 2 and PKC .