Polymorphic Deletion of Three Intracellular Acidic Residues of the α2B-Adrenergic Receptor Decreases G Protein-coupled Receptor Kinase-mediated Phosphorylation and Desensitization

Abstract

A polymorphic variant of the human ␣ 2B -adrenergic receptor (␣ 2B AR), which consists of a deletion of three glutamic acids (residues 301-303) in the third intracellular loop was found to be common in Caucasians (31%) and to a lesser extent in African-Americans (12%). The consequences of this deletion were assessed by expressing wild-type and the Del301-303 receptors in Chinese hamster ovary and COS cells. Ligand binding was not affected, although a small decrease in coupling efficiency to the inhibition of adenylyl cyclase was observed with the mutant. The deletion occurs within a stretch of acidic residues that is thought to establish the milieu for agonist-promoted phosphorylation and desensitization of the receptor by G protein-coupled receptor kinases (GRKs). Agonist-promoted phosphorylation studies carried out in cells coexpressing the ␣ 2B ARs and GRK2 revealed that the Del301-303 receptor displayed ϳ56% of wild-type phosphorylation. Furthermore, the depressed phosphorylation imposed by the deletion was found to result in a complete loss of short term agonistpromoted receptor desensitization. Thus the major phenotype of the Del301-303 ␣ 2B AR is one of impaired phosphorylation and desensitization by GRKs, and thus the polymorphisms renders the receptor incapable of modulation by this key mechanism of dynamic regulation.␣ 2 -Adrenergic receptors (␣ 2 AR) 1 are cell surface receptors for catecholamines that bind to the G i /G o family of G proteins, coupling to multiple effector systems including inhibition of adenylyl cyclase activity (1). ␣ 2 AR are widely expressed within the central and peripheral nervous system (2-4) and participate in a broad spectrum of physiologic functions such as regulation of blood pressure both centrally and within the vasculature, sedation, analgesia, regulation of insulin release, renal function, and cognitive and behavioral functions (5-12). Three human ␣ 2 AR subtypes have been cloned and characterized (␣ 2A , ␣ 2B , and ␣ 2C ). The ␣ 2B AR has a distinct pattern of expression within the brain, liver, lung, and kidney, and recent studies using gene knockouts in mice have shown that disruption of this receptor effects mouse viability (13), blood pressure responses to ␣ 2 AR agonists (13), and the hypertensive response to salt loading (14).Like the ␣ 2A AR subtype (15, 16), the ␣ 2B AR undergoes short term agonist promoted desensitization (17). This desensitization is due to phosphorylation of the receptor, which evokes a partial uncoupling of the receptor from functional interaction with G i /G o (18,19). Such phosphorylation appears to be due to G protein-coupled receptor kinases (GRKs), a family of serine/ threonine kinases that phosphorylate the agonist-occupied conformations of many G protein-coupled receptors (20). The process serves to finely regulate receptor function providing for rapid adaptation of the cell to its environment. Desensitization may also limit the therapeutic effectiveness of administered agonists. For the ␣ 2B AR, phosphorylation of serines/threonine...

α2B-Adrenergic receptor deletion polymorphism and cardiac autonomic nervous system responses to exercise in obese women

“…28 Study using a in vitro cell culture model indicated that the Glu9 variant of the a 2B -AR resulted in a complete loss of the ability for the receptor to undergo agonist-promoted phosphorylation compared with the Glu12 variant. 5 Our findings extend the knowledge to the alteration of the cardiac sympathetic and vagal functions during sustained handgrip exercise in humans with 12Glu9 deletion polymorphism of the a 2B -AR gene. In fact, the present study provides evidences that normotensive women with Glu 9 /Glu 9 genotype have attenuated increase in sympathetic modulation and attenuated reduction in parasympathetic modulation during sustained handgrip exercise when compared with Glu 12 /Glu 12 and Glu 12 /Glu 9 genotypes.…”

“…[1][2][3][4] The functional significance of the 12Glu9 polymorphism of the a 2B -AR gene investigated in vitro cell culture model indicated that the Glu9 variant was associated with impaired receptor desensitization properties under prolonged agonist activation as compared with the Glu12 variant. 5 Studies in obese humans demonstrated that the homozygote short form of a 2B alleles (Glu 9 /Glu 9 ) were associated with slower heart rate (HR). 1 And, a 10-year follow-up study showed that the decrease in parasympathetic function in nondiabetic men with abdominal obesity and impaired cardiac autonomic nervous system (ANS) modulation who were homozygous for Glu 9 /Glu 9 a 2B -AR was greater than the other genotypes.…”

Complex haplotypes derived from noncoding polymorphisms of the intronless α 2A -adrenergic gene diversify receptor expression

“…Five SNPs (Ϫ2014, Ϫ1903, Ϫ1607, 753, and 849) were found exclusively in African Americans, although the allele frequency of the latter was only 0.01. Compared with the ␤ 1 AR (25,26), ␤ 2 AR (27, 28), ␣ 2B AR (29), and ␣ 2C AR (30), which have high-frequency nonsynonymous polymorphisms, the current data show that the ORF of the ␣ 2A AR is virtually invariant at the encoded protein level except for the uncommon substitution at 753, which was found only in those of African descent.…”

Acute stress modulates genotype effects on amygdala processing in humans

“…The increased amygdala response we observed in deletion carriers as compared with nondeletion carriers may result from changes in noradrenaline availability, because the deletion leads to a perturbation of the relationship between incoming noradrenaline signals and receptor responsiveness (23). The α2b-adrenoreceptor is a presynaptic receptor and has a negative feedback function.…”

“…In vitro data suggest that the deletion exerts both agonistic and antagonistic effects (23). This deletion has been found to be associated with increased reexperiencing of traumatic memories in humans (24), which suggests that the deletion acts as a reduced-function variant.…”