Trafficking of Preassembled Opioid μ−δ Heterooligomer−Gz Signaling Complexes to the Plasma Membrane:  Coregulation by Agonists^†

The cellular site of formation, Gα-coupling preference, and agonist regulation of μ−δ opioid receptor (OR) heterooligomers were studied. Bioluminescence resonance energy transfer (BRET) showed that μ−δOR heterooligomers, composed of preformed μ and δ homooligomers, interacted constitutively in the endoplasmic reticulum (ER) with Gα-proteins forming heteromeric signaling complexes before being targeted to the plasma membrane. Compared to μOR homooligomers, the μ−δ heterooligomers showed higher affinity and efficiency of interaction for Gz over Gi, indicating a switch in G-protein preference. Treatment with DAMGO or deltorphin II led to coregulated internalization of both receptors, whereas DPDPE and DSLET had no effect on μ−δ internalization. Staggered expression resulted in non-interacting μ and δ receptors, even though both receptors were colocalized at the cell surface. Agonists failed to induce BRET between staggered receptors, and resulted in internalization solely of the receptor targeted by agonist. Thus, μ−δOR heterooligomers form and preferentially associate with Gz to generate a signaling complex in the ER, and have a distinct agonist-internalization profile compared to either μ or δ homooligomers.