Functional Receptor Coupling to G_i Is A Mechanism of Agonist-Promoted Desensitization of the β₂-Adrenergic Receptor

Abstract

The β₂-adrenergic receptor (β₂AR) couples to G_s, activating adenylyl cyclase (AC) and increasing cAMP. Such signaling undergoes desensitization with continued agonist exposure. β₂AR also couple to G_i after receptor phosphorylation by the cAMP dependent protein kinase A, but the efficiency of such coupling is not known. Given the PKA dependence of β₂AR-G_i coupling, we explored whether this may be a mechanism of agonist-promoted desensitization. HEK293 cells were transfected to express β₂AR or β₂AR and G_iα2, and then treated with vehicle or the agonist isoproterenol to evoke agonist-promoted β₂AR desensitization. Membrane AC activities showed that G_iα2 overexpression decreased basal levels, but the fold-stimulation of the AC over basal by agonist was not altered. However, with treatment of the cells with isoproterenol prior to membrane preparation, a marked decrease in agonist-stimulated AC was observed with the cells overexpressing G_iα2. in the absence of such overexpression, β₂AR desensitization was 23 ± 7%, while with 5-fold G_iα2 overexpression desensitization was 58 ± 5% (p<0.01, n=4). the effect of G_i on desensitization was receptor-specific, in that forskolin responses were not altered by G_iα2 overexpression. Thus, acquired β₂AR coupling to G_i is an important mechanism of agonist-promoted desensitization, and pathologic conditions that increase G_i levels contribute to β₂AR dysfunction.