title: Agonist-induced cell surface trafficking of an intracellularly sequestered D1 dopamine receptor homo-oligomer.
abstract: The role of oligomerization in D1 dopamine receptor trafficking to the cell surface was examined using conformationally distinct variants of this receptor. Substitution of the highly conserved aspartic acid (Asp103) in transmembrane domain 3 resulted in a constitutively active receptor, D103A, that did not bind agonists or antagonists but trafficked to the cell surface as oligomers. Coexpression of D103A with the wild-type D1 receptor in human embryonic kidney 293t cells resulted in inhibition of cell surface expression of the D1 receptor because of receptor oligomerization, causing intracellular retention of both proteins. Rescue of the intracellularly retained oligomer could be achieved only by membrane-permeable full and partial agonists, which resulted in cell surface expression of the D1 receptor, whereas cell-permeable antagonists and cell impermeable agonists had no effect. Cell surface fluorescence resonance energy transfer studies of cells coexpressing D103A and D1 revealed no signal before agonist treatment but a robust signal after agonist treatment, indicating that the intact D1/D103A oligomer reached the cell surface only after agonist treatment but not under basal conditions. This suggests that rescue of the retained D1/D103A oligomer to the cell surface was a result of an agonist-induced change in the conformation of D1, permitting cell surface trafficking of the D1/D103A receptor oligomeric complex from the endoplasmic reticulum.
authors.affiliation: Department of Pharmacology, University of Toronto; Medical Sciences Building Rm 4358, Toronto, ON, Canada M5S 1A8.
authors.forename: Michael M C
authors.initials: MM
authors.lastname: Kong
authors.forename: Theresa
authors.initials: T
authors.lastname: Fan
authors.forename: George
authors.initials: G
authors.lastname: Varghese
authors.forename: Brian F
authors.initials: BF
authors.lastname: O'dowd
authors.forename: Susan R
authors.initials: SR
authors.lastname: George
mesh_terms: D015647:2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine / Q000494:pharmacology; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001058:Apomorphine / Q000494:pharmacology; D001552:Benzazepines / Q000378:metabolism / Q000494:pharmacology; D001667:Binding, Competitive / Q000187:drug effects; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002463:Cell Membrane Permeability; D002522:Chlorocebus aethiops; D000242:Cyclic AMP / Q000378:metabolism; D019281:Dimerization; D004298:Dopamine / Q000494:pharmacology; D018491:Dopamine Agonists / Q000494:pharmacology*; D015870:Gene Expression; D006801:Humans; D008856:Microscopy, Fluorescence; D009154:Mutation / Q000235:genetics; D021381:Protein Transport / Q000187:drug effects; D011869:Radioligand Assay; D017447:Receptors, Dopamine D1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D001552:Benzazepines; D018491:Dopamine Agonists; D017447:Receptors, Dopamine D1; D015647:2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; C067113:SK&F 81297; D000242:Cyclic AMP; D001058:Apomorphine; D004298:Dopamine