title: Salmeterol stimulation dissociates beta2-adrenergic receptor phosphorylation and internalization.
abstract: Salmeterol is a long-acting beta(2)-adrenergic receptor (beta(2)AR) agonist commonly used in the treatment of asthma and chronic obstructive pulmonary disease. It differs from other beta-agonists in that it has a very low intrinisic efficacy, especially when compared with the other available long-acting beta-agonist, formoterol. Receptor desensitization and down-regulation has been described with the chronic use of beta-agonists. This effect may not be the same with all beta-agonists and may be related to their stabilization of altered receptor states. The extreme hydrophobicity and high-affinity quasi-irreversible binding of salmeterol have rendered studies examining the mechanisms by which it mediates receptor desensitization, down-regulation, and internalization difficult. We determined the capacity of salmeterol to induce beta(2)AR endocytosis, G protein-coupled receptor kinase (GRK)-site phosphorylation, degradation, and beta-arrestin2 translocation in HEK293 cells as compared with other agonists of varying intrinsic efficacies. Despite stimulating GRK-mediated phosphorylation of Ser355,356 after 30 min and 18 h to an extent similar to that observed with agonists of high intrinsic efficacy, such as epinephrine and formoterol, salmeterol did not induce significant beta(2)AR internalization or degradation and was incapable of stimulating the translocation of enhanced green fluorescent protein-beta-arrestin2 chimera (EGFP-beta-arrestin2) to the cell surface. Salmeterol-induced receptor endocytosis was rescued, at least in part, by the overexpression of EGFP-beta-arrestin2. Our data indicate that salmeterol binding induces an active receptor state that is unable to recruit beta-arrestin or undergo significant endocytosis or degradation despite stimulating considerable GRK-site phosphorylation. Defects in these components of salmeterol-induced receptor desensitization may be important determinants of its sustained bronchodilation with chronic use.
authors.affiliation: Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. rmoore@bcm.tmc.edu
authors.forename: Robert H
authors.initials: RH
authors.lastname: Moore
authors.forename: Ellen E
authors.initials: EE
authors.lastname: Millman
authors.forename: Veronica
authors.initials: V
authors.lastname: Godines
authors.forename: Nicola A
authors.initials: NA
authors.lastname: Hanania
authors.forename: Tuan M
authors.initials: TM
authors.lastname: Tran
authors.forename: Hui
authors.initials: H
authors.lastname: Peng
authors.forename: Burton F
authors.initials: BF
authors.lastname: Dickey
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Knoll
authors.forename: Richard B
authors.initials: RB
authors.lastname: Clark
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000420:Albuterol / Q000031:analogs & derivatives* / Q000494:pharmacology; D019390:Arrestins / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects*; D004797:Enzyme-Linked Immunosorbent Assay; D004837:Epinephrine / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D010766:Phosphorylation / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D018343:Receptors, Adrenergic, beta-2 / Q000032:analysis / Q000378:metabolism*; D000068299:Salmeterol Xinafoate; D051552:beta-Adrenergic Receptor Kinases / Q000378:metabolism
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D000318:Adrenergic beta-Agonists; D019390:Arrestins; D018343:Receptors, Adrenergic, beta-2; D000068299:Salmeterol Xinafoate; D051552:beta-Adrenergic Receptor Kinases; D000420:Albuterol; D004837:Epinephrine