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G-protein-coupled receptors (GPCRs) have recently joined the list of cell surface receptors that dimerize. Dimerization has been shown to alter the ligand-binding, signaling, and trafficking properties of these receptors. Recent studies have shown that GPCRs heterodimerize with closely related members, resulting in the modulation of their function. In this study, we have attempted to determine whether members of GPCR superfamilies that couple to different families of G-proteins can associate and form oligomers. We chose the ␤2 adrenergic receptor that couples to stimulatory G-proteins and ␦ & opioid receptors that couple to inhibitory G-proteins. ␤2 and ␦ receptors undergo robust agonist-mediated endocytosis, whereas receptors do not. We find that when coexpressed, ␤2 receptors can form heteromeric complexes with both ␦ and receptors. This heterooligomerization does not significantly alter the ligand binding or coupling properties of the receptors. However, it affects the trafficking properties of the receptors. For example, we find that ␦ receptors, when coexpressed with ␤2 receptors, undergo isoproterenol-mediated endocytosis. Conversely, ␤2 receptors in these cells undergo etorphine-mediated endocytosis. However, ␤2 receptors, when coexpressed with receptors, undergo neither opioid-nor isoproterenol-mediated endocytosis. Moreover, these cells exhibit a substantial decrease in the isoproterenol-induced phosphorylation of mitogen-activated protein kinases. Taken together, these results provide direct evidence of heteromerization of GPCRs that couple to different types of G-proteins, which results in the modulation of receptor trafficking and signal transduction.receptor subtypes ͉ G-protein-coupled receptor ͉ endocytosis ͉ dimers ͉ heterodimerization G -protein-coupled receptors (GPCRs), the largest and most diverse family of transmembrane receptors, are involved in the transduction of signals in response to a wide variety of stimuli. Until recently they were thought to function as monomers (1). However, a growing number of biochemical, biophysical, and functional studies suggest that GPCRs form functional, SDS-stable dimers (2). Heteromeric assembly of the nonfunctional ␥-aminobutyric acid receptor B R1a and ␥-aminobutyric acid receptor B R2 is necessary for ␥-aminobutyric acid receptor-mediated signaling in cultured cells and in rat superior ganglion neurons (3-8). In contrast, fully functional opioid receptors associate with each other, resulting in the generation of heterodimeric receptors with unique properties (9-11). Heterodimerization of fully functional somatostatin receptor 5 with somatostatin receptor 1 has also been found to alter the pharmacology and signaling of both receptors (12). Although there is increasing evidence for receptor dimerization between members of the same subfamily of GPCRs, not much is known about dimerization between members of different subfamilies.␤ 2 -Adrenergic receptors are prototypical receptors of the rhodopsin family that undergo rapid and robust agonistmediated endocyt...

Protein-protein interactions are involved in the regulation of a large number of biological processes. It is well established that a variety of cell surface receptors interact with each other to form dimers, and that this is essential for their activation. Although the existence of G protein coupled receptor dimers was predicted from early pharmacological and biochemical analysis, solid evidence supporting dimerization has come within the past few years following the cloning of G protein coupled receptor cDNAs. Using differential epitope tagging and selective immunoisolation of receptor complexes, dimerization of a number of G protein coupled receptors including members of the rhodopsin, secretin, and metabotropic glutamate receptor families have been reported. More recently fluorescence or bioluminescence resonance energy transfer techniques have been used to examine dimerization of these receptors in live cells. These studies have found that whereas in some cases there is an agonist induced increase in the level of dimers, in others there is a decrease or no change in dimer levels. Several recent studies have also reported the ability of related members of G protein coupled receptors to heterodimerize. These heterodimers exhibit distinct physical and functional properties. Examination of possible sites of interactions between receptors has implicated a role for extracellular, transmembrane and/or C-terminal region in dimerization. The functional consequences of dimerization, explored mainly using mutant receptors, have demonstrated a role in modulating agonist affinity, efficacy, and/or trafficking properties. Thus dimerization appears to be a universal phenomenon that provides an additional mechanism for modulation of receptor function as well as cross-talk between G protein coupled receptors.

G-protein-coupled receptors (GPCRs) have recently joined the list of cell surface receptors that dimerize. Dimerization has been shown to alter the ligand-binding, signaling, and trafficking properties of these receptors. Recent studies have shown that GPCRs heterodimerize with closely related members, resulting in the modulation of their function. In this study, we have attempted to determine whether members of GPCR superfamilies that couple to different families of G-proteins can associate and form oligomers. We chose the beta2 adrenergic receptor that couples to stimulatory G-proteins and delta & kappa opioid receptors that couple to inhibitory G-proteins. beta2 and delta receptors undergo robust agonist-mediated endocytosis, whereas kappa receptors do not. We find that when coexpressed, beta2 receptors can form heteromeric complexes with both delta and kappa receptors. This heterooligomerization does not significantly alter the ligand binding or coupling properties of the receptors. However, it affects the trafficking properties of the receptors. For example, we find that delta receptors, when coexpressed with beta2 receptors, undergo isoproterenol-mediated endocytosis. Conversely, beta2 receptors in these cells undergo etorphine-mediated endocytosis. However, beta2 receptors, when coexpressed with kappa receptors, undergo neither opioid- nor isoproterenol-mediated endocytosis. Moreover, these cells exhibit a substantial decrease in the isoproterenol-induced phosphorylation of mitogen-activated protein kinases. Taken together, these results provide direct evidence of heteromerization of GPCRs that couple to different types of G-proteins, which results in the modulation of receptor trafficking and signal transduction.