Serotonin 5-HT(7) receptor slows down the G(s) protein: a single molecule perspective

The 5-hydroxytryptamine (serotonin) receptor type 7 (5-HT(7)R) is a G protein–coupled receptor present primarily in the nervous system and gastrointestinal tract, where it regulates mood, cognition, digestion, and vasoconstriction. 5-HT(7)R has previously been shown to bind to its cognate stimulatory G(s) protein in the inactive state. This phenomenon, termed “inverse coupling,” is thought to counteract the atypically high intrinsic activity of 5-HT(7)R. However, it is not clear how active and inactive 5-HT(7) receptors affect the mobility of the G(s) protein in the plasma membrane. Here, we used single-molecule imaging of the G(s) protein and 5-HT(7)R to evaluate G(s) mobility in the membrane in the presence of 5-HT(7)R and its mutants. We show that expression of 5-HT(7)R dramatically reduces the diffusion rate of G(s). Expression of the constitutively active mutant 5-HT(7)R (L173A) is less effective at slowing G(s) diffusion presumably due to the reduced ability to form long-lasting inactive complexes. An inactive 5-HT(7)R (N380K) mutant slows down G(s) to the same extent as the wild-type receptor. We conclude that inactive 5-HT(7)R profoundly affects G(s) mobility, which could lead to G(s) redistribution in the plasma membrane and alter its availability to other G protein–coupled receptors and effectors.