Serotonin 5-HT receptor slows down the G protein: a single molecule perspective.

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