Homodimerization of the β2-Adrenergic Receptor as a Prerequisite for Cell Surface Targeting

Abstract

Although homodimerization has been demonstrated for a large number of G protein-coupled receptors (GPCRs), no general role has been attributed to this process. Because it is known that oligomerization plays a key role in the quality control and endoplasmic reticulum (ER) export of many proteins, we sought to determine if homodimerization could play such a role in GPCR biogenesis. Using the ␤2-adrenergic receptor (␤2AR) as a model, cell fractionation studies revealed that receptor homodimerization is an event occurring as early as the ER. Supporting the hypothesis that receptor homodimerization is involved in ER processing, ␤2AR mutants lacking an ER-export motif or harboring a heterologous ER-retention signal dimerized with the wild-type receptor and inhibited its trafficking to the cell surface. Finally, in addition to inhibiting receptor dimerization, disruption of the putative dimerization motif, 276 GXXXGXXXL 284, prevented normal trafficking of the receptor to the plasma membrane. Taken together, these data indicate that ␤2AR homodimerization plays an important role in ER export and cell surface targeting.Recent studies have established that G protein-coupled receptors (GPCRs), 1 much like other membrane receptors, form both homodimers and heterodimers (1, 2). Various functions have been attributed to the dimerization between different receptor subtypes. For instance, heterodimerization has been proposed to promote the formation of receptors with unique pharmacological properties, contributing to the pharmacological diversity of GPCRs (3-5). Changes in G protein coupling specificity (6) as well as altered receptor endocytosis (3, 4, 6 -9) have also been proposed to result from receptor heterodimerization. For the metabotropic GABAb receptor, heterodimerization between GABAb-R1 and GABAb-R2 is an obligatory step for cell surface expression of a functional receptor (10,11). This was shown to result from the masking of an endoplasmic reticulum (ER) retention signal found in the C terminus of GABAb-R1, presumably through the formation of a coiled-coil domain between the C termini of the two receptor subtypes (12). The obligatory nature of this heterodimerization for cell surface trafficking and signaling was considered for a long time as a particularity of the GABAb receptor, which belongs to a GPCR subfamily (family 3) containing only a few members (13). However, a recent study indicated that the heterodimerization between two closely related members of family 1 GPCRs, the adrenergic ␣ 1D and ␣ 1B receptors, may be a prerequisite for the proper cell surface expression of the ␣ 1D subtype in both experimental models and real tissues (14).So far, despite the numerous examples of receptor homodimerization reported in the literature (15, 16), the biological function of this phenomenon has remained elusive, probably because it is extremely difficult to experimentally differentiate the dimer properties of identical receptor subtypes from those of monomers. However, a growing number of observations indicates that GPC...

G protein-coupled receptor-receptor interactions give integrative dynamics to intercellular communication

“…Indeed, receptor dimerization can occur in the ER (Smith and Milligan, 2010;Dupré et al, 2012). Mutagenesis studies targeting the putative interaction interface of β 2 -adrenergic receptor showed that inhibiting β 2 dimerization caused its retention in the ER, thus supporting the hypothesis of receptor oligomerization in this cellular compartment (Salahpour et al, 2004). Other examples encompass the D 4 receptors (Van Craenenbroeck et al, 2011), GABA B receptors (MargetaMitrovic et al, 2000), and 5-HT 2C receptors (Herrick-Davis et al, 2006).…”

Cell-surface targeting of α2-adrenergic receptors — Inhibition by a transport deficient mutant through dimerization

“…Therefore, dimers are unable to export from the ER. Dimerization has been well described for a variety of G protein-coupled receptors [9][10][11][12][13][14][15][34][35][36][37][38][39][40][41]. However, whether α 2B -AR is capable of forming homodimers has not been reported.…”

“…In contrast, AT1R-GFP was not found in the anti-HA immunoprecipitate when co-expressed with HA-α 2B -AR. These data indicate that HA-and GFP-tagged α 2B -AR or α 2B -ARm were co-immunoprecipitated as a complex and suggest that α 2B -AR as well as α 2B -ARm underwent homodimerization, similar to many other G protein-coupled receptors [9][10][11][12][13][14][15].…”

“…Third, dimerization of the receptors (homo-and/or hetero-dimerization) in the ER may be required for ER export and further cell-surface targeting. Recently, Bouvier's group has demonstrated that mutation of the putative dimerization motif GxxxGxxxL in the β 2 -adrenergic receptor (AR) prevents normal trafficking of the receptor to the plasma membrane [9]. Although dimerization has been demonstrated for a variety of G protein-coupled receptors [10][11][12][13][14][15], whether α 2 -AR is able to homodimerize and the role of dimerization in the trafficking of α 2 -AR remain unknown.…”

Secretin Receptor Oligomers Form Intracellularly during Maturation through Receptor Core Domains

“…With respect to the latter, a series of studies that have asked when and where in the cell newly-synthesized receptor monomers associate has led to the conclusion that oligomerization is a prerequisite for receptor maturation and plasma membrane sorting (14,22,23). These findings pertain mostly to the more well-studied family A GPCRs, and even in those cases only the most interesting or functionally relevant receptor domains have been evaluated for their effects on oligomerization and sorting.…”