α_2C-Adrenergic Receptors Exhibit Enhanced Surface Expression and Signaling upon Association with β₂-Adrenergic Receptors

Abstract

The ␣ 2C -adrenergic receptor (␣ 2C AR) is known to be poorly trafficked to the cell surface when expressed in a variety of cell types. We tested the hypothesis that the surface expression and signaling of ␣ 2C AR might be enhanced by heterodimerization with other G protein-coupled receptors (GPCRs). Cotransfection of ␣ 2C AR with more than 25 related GPCRs revealed that only coexpression with the ␤ 2 -adrenergic receptor (␤ 2 AR) increased the surface localization of ␣ 2C AR in human embryonic kidney-293 cells. Coimmunoprecipitation of ␣ 2C AR with ␤ 2 AR confirmed a physical interaction between the two receptors. Confocal microscopy studies demonstrated that ␣ 2C AR expressed alone was mainly intracellular, whereas ␣ 2C AR coexpressed with ␤ 2 AR was predominantly localized to the plasma membrane. Ligand binding studies revealed a significant increase in ␣ 2C AR binding sites upon coexpression with ␤ 2 AR, with no apparent change in affinity for ␣ 2 AR ligands. Functional assays with the ␣ 2 AR-specific agonist brimonidine (UK 14,304) revealed that coexpression of ␤ 2 AR with ␣ 2C AR enhanced ␣ 2C AR-mediated activation of extracellular signalregulated kinase 1/2. Furthermore, analyses of agonist-promoted receptor endocytosis demonstrated enhanced ␣ 2C AR internalization in response to ␣ 2 AR agonists when ␣ 2C AR and ␤ 2 AR were coexpressed. In addition, substantial cointernalization of ␣ 2C AR in response to ␤AR agonists was observed when ␣ 2C AR was coexpressed with ␤ 2 AR. These data reveal that ␣ 2C AR can interact with ␤ 2 AR in cells in a manner that regulates ␣ 2C AR surface expression, internalization, and functionality.The adrenergic receptors are a family of cell-surface G protein-coupled receptors (GPCRs) that mediate the actions of the hormone epinephrine and the neurotransmitter norepinephrine. The three main adrenergic receptor (AR) classes (␣ 1 , ␣ 2 , and ␤ 2 ) can be further divided into three subtypes each, and all of these subtypes are excellent targets for therapeutic pharmaceuticals. The specific roles of the various adrenergic receptor subtypes is becoming increasingly clear through studies on knock-out mice (Philipp and Hein, 2004), and novel therapies making use of these insights await the development of more subtype-specific drugs. However, two of the adrenergic receptor subtypes, ␣ 2C AR and ␣ 1D

Endothelin receptors: what's new and what do we need to know?

“…Heterodimer pairs studied in such systems include the ANG II AT 1 receptor with bradykinin B 2 receptor (106), AT 2 with B 2 receptor (1), thromboxane ␣-and ␤-receptor (120), dopamine D 1 and D 2 receptors (27), ␣ 2C -and ␤ 2 -adrenergic receptors (92), ␣ 1B -and ␣ 1D -adrenergic receptors (40), and vasopressin V 1a and V 2 receptors (111). This list is far from complete.…”

How and why do GPCRs dimerize?

“…For example, the delivery of two subtypes of α-adrenoceptors to the plasma membrane is facilitated by the coexpressed β 2 -adrenoceptor [57,58]; a mutant rhodopsin lacking the targeting sequence is transported to the outer segment only with wild-type rhodopsin [59]. Dimerization could underlie phosphorylation of two coexpressed GPCR subtypes when only one is activated [60,61].…”

Extracellular Signal-Regulated Kinase 1/2-Mediated Transcriptional Regulation of G-Protein-Coupled Receptor Kinase 3 Expression in Neuronal Cells

“…Furthermore, it has been demonstrated that this unique signaling pathway is the result of heterodimerization of the D1 and D2 receptors (Lee et al, 2004). Heterodimerization of ␣ 2C -AR and ␤ 2 -AR has been reported recently in human embryonic kidney 293 cells (Prinster et al, 2006). However, the relevance of this observation to the present study may be limited due to the significant differences in signaling/trafficking between the ␣ 2C -AR and the ␣ 2A -and ␣ 2B -AR present in our neuronal cells.…”