Agonist-stimulated and tonic internalization of metabotropic glutamate receptor 1a in human embryonic kidney 293 cells:: Agonist-stimulated endocytosis is β-arrestin1 isoform-specific

Metabotropic glutamate receptors (mGluRs) are G protein-coupled receptors (GPCRs) that contribute to the regulation of integrative brain functions such as cognition, motor control, and neural development. Metabotropic glutamate receptors are members of a unique class of GPCRs (class III) that include the calcium sensing and gamma -aminobutyric acid type B receptors. Although mGluRs bear little sequence homology to well-characterized members of the GPCR superfamily, both second messenger-dependent protein kinases and G protein-coupled receptor kinases (GRKs) contribute to mGluR desensitization. Therefore, in the present study, we examined whether beta -arrestins, regulators of GPCR desensitization and endocytosis, are required for mGIuR1 a desensitization and internalization in human embryonic kidney (HEK) 293 cells. Unlike what has been reported for other GPCRs, we find that in response to agonist stimulation, mGluR1a internalization is selectively mediated by beta -arrestin1 in HEK 293 cells. However, even though beta -arrestin1 binds directly to the carboxyl-terminal tall of mGluR1a and redistributes with mGluR1a to endosomes, neither beta -arrestin1 nor beta -arrestin2 seems to contribute to mGluR1a desensitization in HEK 293 cells. We also observed extensive tonic mGluR1a internalization via clathrin-coated vesicles in the absence of agonist. The tonic internalization of mGluR1a is insensitive to antagonist treatment, dominant-negative mutants of GRK2, beta -arrestin1, and dynamin as well as treatments that disrupt caveolae, but is blocked by hypertonic sucrose and concanavalin A treatment. Internalized mGluR1 a is colocalized with clathrin, transferrin receptor, beta (2)-adrenergic receptor, and Rab5 GTPase in endocytic vesicles. Therefore, although mGluR1a internalizes with beta -arrestin in response to agonist, the agonistin-dependent internalization of mGluR1a involves the beta -arrestinin-dependent targeting of mGluR1a to clathrin-coated vesicles.