An Asn to Lys polymorphism in the third intracellular loop of the human α2A-adrenergic receptor imparts enhanced agonist-promoted Gi coupling

alpha (2A)-Adrenergic receptors (alpha (2A)AR) are presynaptic autoinhibitory receptors of noradrenergic neurons in the central and peripheral sympathetic nervous systems, which act to dynamically regulate neurotransmitter release. Signaling through the G(i)/G(o) family of G-proteins, the receptor subserves numerous homeostatic and central nervous system functions. A single nucleotide polymorphism of this receptor, which results in an Asn to Lys substitution at amino acid 251 of the third intracellular loop, was identified in the human population. The frequency of Lys-251 was 10-fold greater in African-Americans than in Caucasians, but was not associated with essential hypertension. To determine the consequences of this substitution, wild-type and Lys-251 receptors were expressed in CHO and COS-7 cells. Expression, ligand binding, and basal receptor function were unaffected by the substitution. However, agonist-promoted [S-35]GTP gammaS binding was similar to 40% greater with the Lys-251 receptor. This enhanced agonist function was observed with catecholamines, azepines, and imidazolines albeit to different degrees. In studies of agonist-promoted functional coupling to G(i), the polymorphic receptor displayed enhanced inhibition of adenylyl cyclase (60 +/- 4.4 versus 46 +/- 4.1% inhibition) and markedly enhanced stimulation of MAP kinase (57 +/- versus 15- +/- a-fold increase over basal) compared with wild-type alpha (2A)AR. The potency of epinephrine in stimulating inositol phosphate accumulation was increased similar to4 fold with the Lys-251 receptor. Unlike previously described variants of G-protein-coupled receptors, where the minor species causes either a loss of function or increased non-agonist function, Lys-251 alpha (2A)AR represents a new class of polymorphism whose phenotype is a gain of agonist-promoted function.