Arrestin-dependent pathways are a central component of G protein-coupled receptor (GPCRs) signaling. However, the molecular processes regulating arrestin binding are to be further illuminated, in particular with regard to the structural impact of GPCR C-terminal disordered regions. Here, we used an integrated biophysical strategy to describe the basal conformations of the C-terminal domains of three class A GPCRs, the vasopressin V2 receptor (V2R), the growth hormone secretagogue or ghrelin receptor type 1a (GHSR) and the β2-adernergic receptor (β2AR). By doing so, we revealed the presence of transient secondary structures in these regions that are potentially involved in the interaction with arrestin. These secondary structure elements differ from those described in the literature in interaction with arrestin. This suggests a mechanism where the secondary structure conformational preferences in the C-terminal regions of GPCRs could be a central feature for optimizing arrestins recognition.

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Arrestin dependent G protein-coupled receptor (GPCR) signaling pathway is regulated by the phosphorylation state of GPCR′s C-terminal domain, but the molecular bases of arrestin:receptor interaction are to be further illuminated. Here we investigated the impact of phosphorylation on the conformational features of the C-terminal region from three Rhodopsin-like GPCRs, the vasopressin V2 Receptor (V2R), the Growth Hormone Secretagogue or ghrelin Receptor type 1a (GHSR) and the β2-Adernergic Receptor (β2AR). Using phosphomimetic variants, we identified pre-formed secondary structure elements, or short linear motif (SLiMs), that undergo specific conformational transitions upon phosphorylation. Of importance, such conformational transition favors arrestin-2 binding. Hence, our results suggest a model in which the cellular signaling specificity of GPCRs is encoded in the phosphorylation-dependent structuration of the C-terminal regions, which will subsequently modulate arrestin conformation and therefore GPCR:arrestin signaling outcomes.