Akt mediates sequestration of the beta(2)-adrenergic receptor in response to insulin.

The counterregulation of catecholamine action by insulin includes insulin-stimulated sequestration of the beta(2)-adrenergic receptor. Herein we examined the signaling downstream of insulin receptor activation, focusing upon the role of 1-phosphatidylinositol 3-kinase and the serine-threonine protein kinase Akt (also known as protein kinase B) in the internalization of beta(2)-adrenergic receptors. Inhibition of 1-phosphatidylinositol 3-kinase by LY294002 blocks insulin-induced sequestration of the beta(2)-adrenergic receptor, implicating Akt in downstream signaling to the beta(2)-adrenergic receptor. Phosphorylation studies of the C-terminal cytoplasmic domain of the beta(2)-adrenergic receptor by Akt in vitro identified Ser(345) and Ser(346) within a consensus motif for Akt phosphorylation. Double mutation (i.e. S345A/S346A) within this motif abolishes insulin counterregulation of beta-adrenergic stimulation of cyclic AMP accumulation as well as insulin-stimulated sequestration. Furthermore, expression of constitutively activated Akt (T308D/S473D) mimics insulin action on cyclic AMP responses and beta(2)-adrenergic receptor internalization. Expression of the dominant-negative version of Akt (K179A/T308A/S473A), in contrast, abolishes both insulin counterregulation of the cyclic AMP response as well as insulin-stimulated sequestration of the beta(2)-adrenergic receptor. The action of the serine-threonine protein kinase Akt in insulin counterregulation mirrors the central role of protein kinase A in beta-agonist-induced desensitization.