Barrestin1-biased agonism at human δ-opioid receptor by peptidic and alkaloid ligands

We have previously reported on the differential regulation of the human delta-opioid receptor (hDOR) by alkaloid (etorphine) and peptidic (DPDPE and deltorphin 1) ligands, in terms of both receptor desensitization and post-endocytic sorting. Since Barrestins are well known to regulate G protein-coupled receptors (GPCRs) signaling and trafficking, we therefore investigated the role of Barrestin1 (the only isoform expressed in our cellular model) in the context of the hDOR. We established clonal cell lines of SK-N-BE cells over-expressing Barrestin1, its dominant negative mutant (Barrestin1(319-418)), and shRNA directed against endogenous Barrestin1. Interestingly, both binding and confocal microscopy approaches demonstrated that Barrestin1 is required for hDOR endocytosis only when activated by etorphine. Conversely, functional experiments revealed that Barrestin1 is exclusively involved in hDOR desensitization promoted by the peptides. Taken together, these results provide substantial evidence for a Barrestin1-biased agonism at hDOR, where Barrestin1 is differentially involved during receptor desensitization and endocytosis depending on the ligand. (C) 2011 Elsevier Inc. All rights reserved.