title: Regulatory properties of alpha(1B)-adrenergic receptors defective in coupling to phosphoinositide hydrolysis.
abstract: Previous studies have suggested that G protein coupling, phospholipase C activation, phosphoinositide hydrolysis, and protein kinase C activation may be required for alpha(1B)-adrenergic receptor regulation, particularly for their endocytosis into intracellular vesicles. Accordingly, the internalization and down-regulation properties of mutated receptors with defects in G protein coupling and second messenger generation were investigated. The Delta12 and Delta5 receptors, previously shown to be defective in G protein coupling, exhibited greater agonist-induced losses of cell surface accessibility assessed by radioligand binding to intact cells on ice than for the wild-type receptor; however, these receptors were completely defective in endocytosis into intracellular vesicles assessed by sucrose density gradient centrifugation. These receptors also did not undergo down-regulation with long-term agonist exposure as did the wild-type receptor; instead, a prominent up-regulation was observed. The Y348A receptor, previously shown to be defective in phosphoinositide hydrolysis and endocytosis was also defective in down-regulation but did not exhibit significant up-regulation. In contrast, a receptor construct with amino acid residues 246 to 261 deleted (Delta[246-261]) was also defective in stimulation of phosphoinositide hydrolysis but exhibited internalization and down-regulation properties essentially identical to those for the wild-type receptor. Together, these results suggest that stimulation of phosphoinositide hydrolysis by alpha(1B)-adrenergic receptors is not required for their endocytosis or down-regulation but that similar and overlapping receptor structural domains are involved in mediating these processes.
authors.affiliation: Department of Pharmacology, University of Nebraska Medical Center, Omaha, Nebraska 68198-6260, USA.
authors.forename: Jiefa
authors.initials: J
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.forename: Jodi L
authors.initials: JL
authors.lastname: Anderson
authors.forename: Nancy A
authors.initials: NA
authors.lastname: Schulte
authors.forename: Myron L
authors.initials: ML
authors.lastname: Toews
mesh_terms: D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D016466:CHO Cells; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D006224:Cricetinae; D015536:Down-Regulation / Q000187:drug effects; D004705:Endocytosis / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation / Q000235:genetics; D010716:Phosphatidylinositols / Q000378:metabolism*; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism*; D014162:Transfection
chemical_list: D000318:Adrenergic beta-Agonists; D010716:Phosphatidylinositols; D018340:Receptors, Adrenergic, alpha-1; D019204:GTP-Binding Proteins