title: Role of G protein-coupled receptor kinase 4 and beta-arrestin 1 in agonist-stimulated metabotropic glutamate receptor 1 internalization and activation of mitogen-activated protein kinases.
abstract: The metabotropic glutamate 1 (mGlu(1)) receptor in cerebellar Purkinje cells plays a key role in motor learning and motor coordination. Here we show that the G protein-coupled receptor kinases (GRK) 2 and 4, which are expressed in these cells, regulate the mGlu(1) receptor by at least in part different mechanisms. Using kinase-dead mutants in HEK293 cells, we found that GRK4, but not GRK2, needs the intact kinase activity to desensitize the mGlu(1) receptor, whereas GRK2, but not GRK4, can interact with and regulate directly the activated Galpha(q). In cells transfected with GRK4 and exposed to agonist, beta-arrestin was first recruited to plasma membranes, where it was co-localized with the mGlu(1) receptor, and then internalized in vesicles. The receptor was also internalized but in different vesicles. The expression of beta-arrestin V53D dominant negative mutant, which did not affect the mGlu(1) receptor internalization, reduced by 70-80% the stimulation of mitogen-activated protein (MAP) kinase activation by the mGlu(1) receptor. The agonist-stimulated differential sorting of the mGlu(1) receptor and beta-arrestin as well as the activation of MAP kinases by mGlu(1) agonist was confirmed in cultured cerebellar Purkinje cells. A major involvement of GRK4 and of beta-arrestin in agonist-dependent receptor internalization and MAP kinase activation, respectively, was documented in cerebellar Purkinje cells using an antisense treatment to knock down GRK4 and expressing beta-arrestin V53D dominant negative mutant by an adenovirus vector. We conclude that GRK2 and GRK4 regulate the mGlu(1) receptor by different mechanisms and that beta-arrestin is directly involved in glutamate-stimulated MAP kinase activation by acting as a signaling molecule.
authors.affiliation: Istituto Neurologico Mediterraneo Neuromed, Istituto di Ricovero e Cura a Carattere Scientifico, 86077 Pozzilli, Italy.
authors.forename: Luisa
authors.initials: L
authors.lastname: Iacovelli
authors.forename: Lorena
authors.initials: L
authors.lastname: Salvatore
authors.forename: Loredana
authors.initials: L
authors.lastname: Capobianco
authors.forename: Antonietta
authors.initials: A
authors.lastname: Picascia
authors.forename: Eliana
authors.initials: E
authors.lastname: Barletta
authors.forename: Marianna
authors.initials: M
authors.lastname: Storto
authors.forename: Stefania
authors.initials: S
authors.lastname: Mariggiò
authors.forename: Michele
authors.initials: M
authors.lastname: Sallese
authors.forename: Antonio
authors.initials: A
authors.lastname: Porcellini
authors.forename: Ferdinando
authors.initials: F
authors.lastname: Nicoletti
authors.forename: Antonio
authors.initials: A
authors.lastname: De Blasi
mesh_terms: D000818:Animals; D019390:Arrestins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D022162:Cytoplasmic Vesicles / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D054771:G-Protein-Coupled Receptor Kinase 4; D006801:Humans; D007668:Kidney / Q000166:cytology; D020935:MAP Kinase Signaling System / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011689:Purkinje Cells / Q000166:cytology / Q000201:enzymology*; D016318:Quisqualic Acid / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D018094:Receptors, Metabotropic Glutamate / Q000819:agonists / Q000378:metabolism*; D014162:Transfection; D051552:beta-Adrenergic Receptor Kinases; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins
chemical_list: C000605253:ARRB1 protein, human; C000605256:Arrb1 protein, rat; D019390:Arrestins; D018690:Excitatory Amino Acid Agonists; D018094:Receptors, Metabotropic Glutamate; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; C104077:metabotropic glutamate receptor type 1; D016318:Quisqualic Acid; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D054771:G-Protein-Coupled Receptor Kinase 4; C517855:GRK4 protein, human; C517854:Grk4 protein, rat; D020928:Mitogen-Activated Protein Kinases