Frizzleds (FZDs) are classified as G-protein-coupling receptors, but how signals are initiated and specified through heterotrimeric G proteins is unknown. FZD6 regulates convergent extension movements, and its C-terminal Arg511Cys mutation causes nail dysplasia in humans. We investigated the functional relationship between FZD6, Disheveled (DVL), and heterotrimeric G proteins. Live cell imaging combined with fluorescence recovery after photobleaching (FRAP) revealed that inactive human FZD6 precouples to Gαi1 and Gαq but not to GαoA,Gαs, and Gα12 proteins. G-protein coupling is measured as a 10-20% reduction in the mobile fraction of fluorescently tagged G proteins on chemical receptor surface cross-linking. The FZD6 Arg511Cys mutation is incapable of G-protein precoupling, even though it still binds DVL. Using both FRAP and Förster resonance energy transfer (FRET) technology, we showed that the FZD6-Gαi1 and FZD-Gαq complexes dissociate on WNT-5A stimulation. Most important, G-protein precoupling of FZD6 and WNT-5A-induced signaling to extracellular signal-regulated kinase1/2 were impaired by DVL knockdown or overexpression, arguing for a strict dependence of FZD6-G-protein coupling on DVL levels and identifying DVL as a master regulator of FZD/G-protein signaling. In summary, we propose a mechanistic connection between DVL and G proteins integrating WNT, FZD, G-protein, and DVL function.
Like most neurotransmitters, serotonin possesses a simple structure. However, the pharmacological consequences are more complex and diverse. Serotonin is involved in numerous functions in the human body including the control of appetite, sleep, memory and learning, temperature regulation, mood, behavior, cardiovascular function, muscle contraction, endocrine regulation, and depression. Low levels of serotonin may be associated with several disorders, namely increase in aggressive and angry behaviors, clinical depression, Parkinson's disease, obsessive-compulsive disorder, eating disorders, migraine, irritable bowel syndrome, tinnitus, and bipolar disease. These effects are mediated via different serotonin (5-HT) receptors. In this review, we will focus on the last discovered member of this serotonin receptor family, the 5-HT7 receptor. This receptor belongs to the G protein-coupled receptor superfamily and was cloned two decades ago. Later, different splice variants were described but no major functional differences have been described so far. All 5-HT7 receptor variants are coupled to Gαs proteins and stimulate cAMP formation. Recently, several interacting proteins have been reported, which can influence receptor signaling and trafficking.
5-HT4 receptor pre-mRNA is alternatively spliced in human (h) tissue to produce several splice variants, called 5-HT4(a) to 5-HT4(h) and 5-HT4(n). Polymerase chain reaction (PCR) with primers designed to amplify both 5-HT4(a) and 5-HT4(b) amplified three additional bands in different tissues, two representing different mRNA species both encoding 5-HT4(g) and one representing mRNA for a novel splice variant named 5-HT4(i), cloned from testis and pancreas respectively. Primary and nested PCR detected both 5-HT4(g) and 5-HT4(i) in multiple tissues. Whereas 5-HT4(i), was found in all cardiovascular tissues analysed, 5-HT4(g) was mainly present in atria. However, quantitative RT-PCR indicated 5-HT4(g) expression also in cardiac ventricle. The pharmacological profiles and ability to activate adenylyl cyclase (AC) were compared between four recombinant h5-HT4 splice variants (a, b, g and i) expressed transiently and stably in HEK293 cells. Displacement of [(3)H]GR113808 with ten ligands revealed identical pharmacological profiles (affinity rank order: GR125487, SB207710, GR113808>SB203186>serotonin, cisapride, tropisetron>renzapride, 5-MeOT>5-CT). In transiently transfected HEK293 cells cisapride was a partial agonist compared to serotonin at 5-HT4(b), 5-HT4(g) and 5-HT4(i) receptors. In membranes from HEK293 cells stably expressing 5-HT4(g) (3,000 fmol/mg protein) or 5-HT4(i) (500 fmol/mg protein), serotonin and 5-MeOT were full agonists while cisapride was full agonist at 5-HT4(g) and partial agonist at 5-HT4(i), probably due to different receptor expression levels. At both 5-HT4(g) and 5-HT4(i), the behaviour of 5-HT4 receptor antagonists was dependent on receptor level. At high receptor levels, tropisetron and SB207710 and to a variable extent SB203186 and GR113808 displayed some partial agonist activity, whereas GR125487 and SB207266 reduced the AC activity below basal, indicating both receptors to be constitutively active. We conclude that the novel 5-HT4(i) receptor splice variant is pharmacologically indistinguishable from other 5-HT4 splice variants and that the 5-HT4(i) C-terminal tail does not influence coupling to AC.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.