Sequestered A431 cell beta-adrenoceptors remain functional after homologous desensitization.
This study seeks to investigate the functional state of sequestered beta-adrenoceptors in A431 cells which have undergone homologous desensitization. Incubation of cells with isoprenaline under desensitizing conditions caused a reduction (a) in the number of cell surface beta-receptors as measured by 3H-CGP 12,177 binding, and (b) in the extent of agonist-stimulatable adenylate cyclase activity in membranes prepared from desensitized cells. We infer that those receptors have been sequestered to an internal membrane site since they were detectable by the lipophilic ligand 125I-CYP but not by the more hydrophilic 3H-CGP 12,177. We confirmed that sequestration had occurred by fractionation on non-linear sucrose density gradients of membranes prepared from desensitized cells. The lighter density membrane fraction contained up to 50% of the total receptor pool after desensitization, but only 5-7% of membrane (fluoride-stimulatable) adenylate cyclase. Fusion of desensitized cells in the presence of polyethylene glycol caused a reassociation of sequestered beta-receptors with their biochemical effector Gs since agonist-dependent adenylate cyclase stimulation was restored to the levels measurable after fusion of non-desensitized cells. We conclude that sequestered beta-receptors in desensitized A431 cells are fully functional.
Cooney
D
D
Department of Pharmacology, University College, Belfield, Dublin, Republic of Ireland.
Keenan
A K
AK
D000262:Adenylyl Cyclases / Q000032:analysis; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000032:analysis; D010766:Phosphorylation; D011092:Polyethylene Glycols / Q000494:pharmacology; D011943:Receptors, Adrenergic, beta / Q000032:analysis / Q000502:physiology*
D011943:Receptors, Adrenergic, beta; D003208:Concanavalin A; D011092:Polyethylene Glycols; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases