Résumé
GPCRs are seven-transmembrane proteins that regulate a broad range of cellular responses, making them prime therapeutic targets in many diseases. These proteins trigger complex intracellular responses resulting from their ability to couple to effectors, such as different G protein subtypes and β-arrestins. Current models suggest that GPCRs can adopt multiple distinct conformational states, differentially stabilized by various ligands, leading to varied signaling outcomes, a phenomenon known as functional selectivity. In this context, we aimed to gain insight into the interplay between conformational dynamics and signaling selectivity using the ghrelin receptor GHSR as a model. In addition to being a key receptor in many biological functions (food intake, energy homeostasis, reward-seeking behaviors), it is a particularly adapted model to explore signaling selectivity coupling to multiple G protein pathways (Gq, Gi/O, G13) and β-arrestins, which regulate the various physiological functions of ghrelin. We have explored the conformational features of this receptor using a strategy that combines in vitro reconstituted models and cellular systems. The combination of these systems allowed us to assess what pertains to the intrinsic properties of the receptor protein and what results from the influence of the cellular environment. The in vitro assay was based on the use of isolated receptors assembled into nanodiscs and labeled with fluorophores that allowed the monitoring of conformational changes associated to the binding of a series of original compounds with specific pharmacological profiles (full, partial, inverse agonists, antagonists). The cellular system relied on intramolecular biosensors based on bioluminescence resonance energy transfer (BRET) that monitored receptor conformational rearrangements in real time. These tools have highlighted certain specific states stabilized by these compounds and the impact of the cellular environment on the receptor conformational landscape. Further study of these features in an integrated, spatiotemporally resolved manner may provide a deeper understanding of the molecular processes underlying GPCR-mediated signaling.