Sequestration and recycling of beta 2-adrenergic receptors permit receptor resensitization.

Stimulation of beta 2-adrenergic receptors in intact cells causes, first, rapid functional uncoupling from Gs, which is triggered by receptor phosphorylation, and, second, somewhat slower sequestration of the receptors to an internal compartment. The present study addresses a possible role of sequestration in the resensitization of desensitized beta 2-adrenergic receptors in human A431 cells. Exposure of these cells to isoproterenol caused rapid phosphorylation, desensitization (as assessed in adenylyl cyclase assays), and sequestration of the receptors.

Overexpression of beta-arrestin and beta-adrenergic receptor kinase augment desensitization of beta 2-adrenergic receptors.

Receptor-specific or homologous desensitization of beta 2-adrenergic receptors is thought to be effected via phosphorylation of the receptor by the beta-adrenergic receptor kinase (beta ARK), followed by binding of beta-arrestin. We have generated stably transfected Chinese hamster ovary cell lines overexpressing either of the two regulatory proteins and also expressing low or high levels of beta 2-adrenergic receptors (approximately 80 and approximately 600 fmol/mg of membrane protein). In these cells, we studied the process of desensitization induced by the beta-adrenergic receptor agonist isoproterenol.

Receptor-specific desensitization with purified proteins. Kinase dependence and receptor specificity of beta-arrestin and arrestin in the beta 2-adrenergic receptor and rhodopsin systems.

Homologous desensitization of beta-adrenergic receptors, as well as adaptation of rhodopsin, are thought to be triggered by specific phosphorylation of the receptor proteins. However, phosphorylation alone seems insufficient to inhibit receptor function, and it has been proposed that the inhibition is mediated, following receptor phosphorylation, by the additional proteins beta-arrestin in the case of beta-adrenergic receptors and arrestin in the case of rhodopsin. In order to test this hypothesis with isolated proteins, beta-arrestin and arrestin were produced by transient overexpression of their cDNAs in COS7 cells and purified to apparent homogeneity.