Preassociation between the 5‐HT7 serotonin receptor and G protein Gs: molecular determinants and association with low potency activation of adenylyl cyclase

ABSTRACT According to early models of GPCR signaling, G proteins only interact with activated receptors. However, some GPCRs were shown to assemble with G proteins before receptor activation, in accordance with more recent models. Previously, we found that the 5‐HT7 receptor, as opposed to the 5‐HT4 receptor, was preassociated with Gs, but the molecular determinants for this interaction are still elusive. In a series of chimeric 5‐HT7 receptors with intracellular segments from 5‐HT4, we determined the receptor–G protein interaction by performing antibody‐immobilized fluorescence recovery after photobleaching and fluorescence resonance energy transfer. We identified the intracellular loop 3 and C‐tail of the 5‐HT7 receptor to be responsible for the preassociation with Gs, and we further delineated the TM5 extension in the intracellular loop 3 and helix 8 in the C‐tail as the molecular determinants. These chimeric exchanges converted the 5‐HT7 receptor into a collision‐coupled receptor that recruited G proteins only upon agonist activation, whereas reciprocal exchanges converted 5‐HT4 to a preassociated receptor. The 5‐HT7 receptor displayed 2‐component agonist‐induced Gs signaling with high and low potency. In addition, the same segments were involved in low‐potency signaling and preassociation. The correspondence between Gs preassociation and low‐potency Gs signaling is a novel aspect of GPCR pharmacology.—Ulsund, A. H., Dahl, M., Frimurer, T. M., Manfra, O., Schwartz, T. W., Levy, F. O., Andressen, K. W. Preassociation between the 5‐HT7 serotonin receptor and G protein Gs: molecular determinants and association with low potency activation of adenylyl cyclase. FASEB J. 33, 3870–3886 (2019). www.fasebj.org