Insights into cannabinoid receptor 2 (CB2) anterograde trafficking and pharmacological chaperoning

Cannabinoid Receptor 2 (CB₂) is a promising therapeutic target for modulating inflammation. Canonical signalling responses to receptor ligands are critically dependent on cell surface receptor expression. However, it is also now appreciated that intracellular G protein-coupled receptors can contribute to signalling responses and influence functional outcomes. Therefore, understanding how the subcellular distribution of receptors is controlled is also highly pertinent. CB₂ is observed to be expressed at the cell surface as well as having a considerable proportion expressed intracellularly. Despite this distribution being well established, little is known about the regulation of CB₂ anterograde trafficking and subcellular distribution. We report that sustained treatment with a range of CB₂ agonists and inverse agonists stimulates a distinct population of CB₂ to be delivered to the cell surface, at various expression levels and despite agonists concurrently internalising cell surface CB₂. We present evidence that this ligand-stimulated anterograde trafficking is a result of CB₂ agonists, as well as inverse agonists, acting as pharmacological chaperones. We also report that a di-lysine (KK) motif in the CB₂ C-terminal tail is required for basal delivery to the cell surface. Corroborating the hypothesis that CB₂ ligands can act as pharmacological chaperones, sustained CB₂ ligand stimulation induces cell surface expression of the mutated receptor and alters maturation states as measured by western blotting. Our finding that prolonged exposure to CB₂ ligands can induce CB₂ cell surface delivery via pharmacological chaperoning may well have important implications for optimal design of CB₂-targeted therapeutics.