Abstract
The sweet taste receptor is rather unique, recognizing a diverse repertoire of natural or synthetic ligands, with a surprisingly large structural diversity, and with potencies stretching over more than six orders of magnitude. Yet, it is not clear if different cell-based assays can faithfully report the relative potencies and efficacies of these molecules. Indeed, up to now, sweet taste receptor agonists have been almost exclusively characterized using cell-based assays developed with overexpressed and promiscuous G proteins. This non-physiological coupling has allowed the quantification of receptor activity via phospholipase C activation and calcium mobilization measurements in heterologous cells on a FLIPR system, for example. Here, we developed a novel assay for the human sweet taste receptor where endogenous G proteins and signaling pathways are recruited by the activated receptor. The effects of several sweet taste receptor agonists and other types of modulators were recorded by measuring changes in dynamic mass redistribution (DMR) using an Epic® reader. Potency and efficacy values obtained in the DMR assay were compared to those results obtained with the classical FLIPR assay. Results demonstrate that for some ligands, the two assay systems provide similar information. However, a clear bias for the FLIPR assay was observed for one third of the agonists evaluated, suggesting that the use of non-physiological coupling may influence the potency and efficacy of sweet taste receptor ligands. Replacing the promiscuous G protein with a chimeric G protein containing the C-terminal tail 25 residues of the physiologically relevant G protein subunit Gαgustducin reduced or abrogated bias.
Highlights
Two families of G protein-coupled receptors (GPCRs), T1Rs and T2Rs, expressed in specific taste receptor cells on the tongue and palate mediate the taste sensation of sweet, savory- and bittertasting substances
The hT1R2 and hT1R3 sweet receptor subunits were introduced in U2OS cells and a stable clone was used to develop a dynamic mass redistribution (DMR) assay
To evaluate functional selectivity in taste receptors we undertook the most comprehensive pharmacological characterization of human sweet taste receptor reported to date
Summary
Two families of G protein-coupled receptors (GPCRs), T1Rs and T2Rs, expressed in specific taste receptor cells on the tongue and palate mediate the taste sensation of sweet-, savory- and bittertasting substances. Sweet taste is triggered at the periphery by a pair of GPCRs called T1R2 and T1R3, which function as an obligate heterodimer (Chandrashekar et al, 2006). Agonists such as fructose, sucrose, glucose, aspartame, neotame and saccharin activate the sweet receptor dimer (T1R2/R3) expressed in heterologous cells (Nelson et al, 2001; Li et al, 2002) and deletion of T1R2 or T1R3 in. Umami taste is triggered by a related pair of heterodimeric GPCRs called T1R1 and T1R3 and amino acids such as glutamate (MSG) and aspartate activate the umami receptor dimer (T1R1/R3) expressed in heterologous cells (Li et al, 2002; Nelson et al, 2002). Activation of Gαgustducin in turn activates other effectors such as PLCβ2 and TRPM5 (Zhang et al, 2003) leading to cell depolarization and taste perception
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