title: G protein-coupled receptor 30 (GPR30) forms a plasma membrane complex with membrane-associated guanylate kinases (MAGUKs) and protein kinase A-anchoring protein 5 (AKAP5) that constitutively inhibits cAMP production.
abstract: GPR30, or G protein-coupled estrogen receptor, is a G protein-coupled receptor reported to bind 17β-estradiol (E2), couple to the G proteins Gs and Gi/o, and mediate non-genomic estrogenic responses. However, controversies exist regarding the receptor pharmacological profile, effector coupling, and subcellular localization. We addressed the role of the type I PDZ motif at the receptor C terminus in receptor trafficking and coupling to cAMP production in HEK293 cells and CHO cells ectopically expressing the receptor and in Madin-Darby canine kidney cells expressing the native receptor. GPR30 was localized both intracellularly and in the plasma membrane and subject to limited basal endocytosis. E2 and G-1, reported GPR30 agonists, neither stimulated nor inhibited cAMP production through GPR30, nor did they influence receptor localization. Instead, GPR30 constitutively inhibited cAMP production stimulated by a heterologous agonist independently of Gi/o. Moreover, siRNA knockdown of native GPR30 increased cAMP production. Deletion of the receptor PDZ motif interfered with inhibition of cAMP production and increased basal receptor endocytosis. GPR30 interacted with membrane-associated guanylate kinases, including SAP97 and PSD-95, and protein kinase A-anchoring protein (AKAP) 5 in the plasma membrane in a PDZ-dependent manner. Knockdown of AKAP5 or St-Ht31 treatment, to disrupt AKAP interaction with the PKA RIIβ regulatory subunit, decreased inhibition of cAMP production, and St-Ht31 increased basal receptor endocytosis. Therefore, GPR30 forms a plasma membrane complex with a membrane-associated guanylate kinase and AKAP5, which constitutively attenuates cAMP production in response to heterologous agonists independently of Gi/o and retains receptors in the plasma membrane.
authors: From the Departments of Experimental Medical Science.
authors: Stefan
authors: S
authors: Broselid
authors: the Department of Pharmacology, University of Texas Health Science Center, San Antonio, Texas 78229.
authors: Kelly A
authors: KA
authors: Berg
authors: the Department of Pharmacology, University of Texas Health Science Center, San Antonio, Texas 78229.
authors: Teresa A
authors: TA
authors: Chavera
authors: Nephrology, and.
authors: Robin
authors: R
authors: Kahn
authors: the Department of Pharmacology, University of Texas Health Science Center, San Antonio, Texas 78229.
authors: William P
authors: WP
authors: Clarke
authors: Cardiology, Lund University, 22184 Lund, Sweden and.
authors: Björn
authors: B
authors: Olde
authors: From the Departments of Experimental Medical Science, fredrik.leeb-lundberg@med.lu.se.
authors: L M Fredrik
authors: LM
authors: Leeb-Lundberg
mesh_terms: D054758:A Kinase Anchor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000096:biosynthesis*; D004285:Dogs; D055785:Gene Knockdown Techniques; D051528:Guanylate Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054731:PDZ Domains; D011960:Receptors, Estrogen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D054758:A Kinase Anchor Proteins; C487775:AKAP5 protein, human; C109584:GPER1 protein, human; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D011960:Receptors, Estrogen; D043562:Receptors, G-Protein-Coupled; D011994:Recombinant Proteins; D000242:Cyclic AMP; D051528:Guanylate Kinases
keywords: A Kinase-anchoring Protein (AKAP); Adenylate Cyclase (Adenylyl Cyclase); Cyclic AMP (cAMP); G Protein-coupled Estrogen Receptor; G Protein-coupled Receptor (GPCR); GPR30; Membrane Trafficking; Membrane-associated Guanylate Kinase; PDZ Domain; Protein Kinase A (PKA)