Activation of the inhibitory GTP-binding protein of adenylate cyclase, Gi, by beta-adrenergic receptors in reconstituted phospholipid vesicles.

T Asano,T Katada,A G Gilman,E M Ross

doi:10.1016/s0021-9258(17)42705-0

Abstract

beta-Adrenergic receptors and the inhibitory GTP-binding protein, Gi of the adenylate cyclase system were reconstituted into phospholipid vesicles by the method described previously for reconstituting receptors and the stimulatory GTP-binding protein, Gs (Brandt, D. R., Asano, T., Pedersen, S. E., and Ross, E. M. (1983) Biochemistry 22, 4357-4362). In the receptor-Gi vesicles, beta-adrenergic agonists stimulated both the high-affinity binding of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) to Gi and GTPase activity to an extent similar to that observed in vesicles containing beta-adrenergic receptors and Gs. Stimulation required receptors and displayed appropriate beta-adrenergic specificity. The prior treatment of receptor-Gi vesicles with islet-activating protein (pertussis toxin) plus NAD markedly inhibited both the isoproterenol-stimulated binding of GTP gamma S and the isoproterenol-stimulated GTPase activity. No contamination of Gi by Gs was apparent. These data suggest that receptors that typically stimulate adenylate cyclase activity may also activate the inhibitory system, perhaps as one mechanism of desensitization.

Highlights

Adenylate cyclase activity in animal cells is controlled by receptor-mediated hormonal stimulation as well as receptormediated inhibition
The data presented indicate that P-adrenergic receptors, which generally stimulate adenylate cyclase via the GTPbinding protein, G, can stimulate the activation of the inhibitory GTP-binding protein, Gi
Stimulation was demonstrated as an increased rate of GTPyS binding and a n increased GTPase activity caused by p-adrenergic agonists

Summary

Introduction

Adenylate cyclase activity in animal cells is controlled by receptor-mediated hormonal stimulation as well as receptormediated inhibition (see Ref. 1 for review). The mechanisms of these events are homologous. The activation of Gi causes the cy and By subunits to dissociate This has suggested that Gi-mediated inhibition primarily reflects an increased concentration of free by that promotes the reassociation of fir with the 01 subunit of G, t o cause its deactivation [7, 8]. It is likely that the 01 subunit of Gi causes inhibition of adenylate cyclase, not necessarily directly [8,9,10]

Methods

Results

Conclusion