title: Dynamic Regulation of Quaternary Organization of the M1 Muscarinic Receptor by Subtype-selective Antagonist Drugs.
abstract: Although rhodopsin-like G protein-coupled receptors can exist as both monomers and non-covalently associated dimers/oligomers, the steady-state proportion of each form and whether this is regulated by receptor ligands are unknown. Herein we address these topics for the M1 muscarinic acetylcholine receptor, a key molecular target for novel cognition enhancers, by using spatial intensity distribution analysis. This method can measure fluorescent particle concentration and assess oligomerization states of proteins within defined regions of living cells. Imaging and analysis of the basolateral surface of cells expressing some 50 molecules·μm(-2) human muscarinic M1 receptor identified a ∼75:25 mixture of receptor monomers and dimers/oligomers. Both sustained and shorter term treatment with the selective M1 antagonist pirenzepine resulted in a large shift in the distribution of receptor species to favor the dimeric/oligomeric state. Although sustained treatment with pirenzepine also resulted in marked up-regulation of the receptor, simple mass action effects were not the basis for ligand-induced stabilization of receptor dimers/oligomers. The related antagonist telenzepine also produced stabilization and enrichment of the M1 receptor dimer population, but the receptor subtype non-selective antagonists atropine and N-methylscopolamine did not. In contrast, neither pirenzepine nor telenzepine altered the quaternary organization of the related M3 muscarinic receptor. These data provide unique insights into the selective capacity of receptor ligands to promote and/or stabilize receptor dimers/oligomers and demonstrate that the dynamics of ligand regulation of the quaternary organization of G protein-coupled receptors is markedly more complex than previously appreciated. This may have major implications for receptor function and behavior.
authors.affiliation: From the Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, Scotland, United Kingdom and.
authors.forename: John D
authors.initials: JD
authors.lastname: Pediani
authors.affiliation: From the Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, Scotland, United Kingdom and.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Ward
authors.affiliation: Institut d'Optique and CNRS, Laboratoire Photonique, Numérique et Nanosciences (LP2N) and Université de Bordeaux, LP2N, F-33405, UMR 5298, 33405 Talence Cedex, France.
authors.forename: Antoine G
authors.initials: AG
authors.lastname: Godin
authors.affiliation: From the Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, Scotland, United Kingdom and.
authors.forename: Sara
authors.initials: S
authors.lastname: Marsango
authors.affiliation: From the Molecular Pharmacology Group, Institute of Molecular, Cell and Systems Biology, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, Scotland, United Kingdom and Graeme.Milligan@glasgow.ac.uk.
authors.forename: Graeme
authors.identifier: 0000-0002-6946-3519
authors.initials: G
authors.lastname: Milligan
mesh_terms: D001285:Atropine / Q000494:pharmacology*; D002460:Cell Line; D006801:Humans; D018727:Muscarinic Antagonists / Q000494:pharmacology*; D010890:Pirenzepine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D055503:Protein Multimerization / Q000187:drug effects*; D043643:Receptor, Muscarinic M1 / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism
chemical_list: D018727:Muscarinic Antagonists; D043643:Receptor, Muscarinic M1; C046338:telenzepine; D010890:Pirenzepine; D001285:Atropine
keywords: 7-helix receptor; G protein-coupled receptor (GPCR); molecular pharmacology; oligomerization; receptor; signal transduction; transmembrane domain