The T1R receptors, a family of taste-specific class C G proteincoupled receptors, mediate mammalian sweet and umami tastes. The structure-function relationships of T1R receptors remain largely unknown. In this study, we demonstrate the different functional roles of T1R extracellular and transmembrane domains in ligand recognition and G protein coupling. Similar to other family C G protein-coupled receptors, the N-terminal Venus flytrap domain of T1R2 is required for recognizing sweeteners, such as aspartame and neotame. The G protein coupling requires the transmembrane domain of T1R2. Surprisingly, the C-terminal transmembrane domain of T1R3 is required for recognizing sweetener cyclamate and sweet taste inhibitor lactisole. Because T1R3 is the common subunit in the sweet taste receptor and the umami taste receptor, we tested the interaction of lactisole and cyclamate with the umami taste receptor. A family of class C G protein-coupled receptors (GPCRs), T1Rs, is selectively expressed in the taste buds (1-6). Functional expression of T1Rs in human embryonic kidney (HEK)-293 cells revealed that different combinations of T1Rs respond to sweet and umami taste stimuli (6, 7). T1R2 and T1R3, when coexpressed in HEK-293 cells, recognize diverse natural and synthetic sweeteners. Similarly, T1R1 and T1R3, when coexpressed in HEK-293 cells, respond to the umami taste stimulus L-glutamate. This response is enhanced by 5Ј ribonucleotides, a hallmark of umami taste. Recent experiments with knockout mice confirmed that T1Rs indeed mediate mouse sweet and umami tastes (8, 9).The class C GPCRs possess a large N-terminal extracellular domain, often referred to as the Venus flytrap domain (10), and are known to function as either homodimers, in the cases of metabotropic glutamate receptors (mGluRs) and calciumsensing receptor, or heterodimers, in the case of ␥-aminobutyric acid type B receptor (GABA B R) (10). The functional expression data suggest a heterodimer mechanism for T1Rs: both T1R1 and T1R2 need to be coexpressed with T1R3 to be functional, which is supported by the overlapping expression patterns of T1Rs in rodent tongue. Nonetheless, there has been no direct evidence that T1Rs function as heteromeric complexes. It is possible that T1R3 is not a functional component of sweet and umami taste receptors, but merely a chaperone protein, which facilitates the proper folding or intracellular translocation of T1R1 and T1R2. The distinct ligand specificities of T1R1͞T1R3 and T1R2͞T1R3 receptors suggest that T1R1 and T1R2 play more important roles in ligand binding in sweet and umami taste receptors than T1R3. Support for this hypothesis was provided recently by results from mouse genetics where human T1R2 transgenic mice, generated on the T1R2 knockout background, displayed sweetener taste preferences similar to those of humans (9). However, the functional role of T1R3 and the overall structure͞function relationship of T1R taste receptors remain largely unknown.Another intriguing observation about the T1R2͞T1R3 receptor is th...

Variation in human taste is a well-known phenomenon. However, little is known about the molecular basis for it. Bitter taste in humans is believed to be mediated by a family of 25 G protein-coupled receptors (hT2Rs, or TAS2Rs). Despite recent progress in the functional expression of hT2Rs in vitro, up until now, hT2R38, a receptor for phenylthiocarbamide (PTC), was the only gene directly linked to variations in human bitter taste. Here we report that polymorphism in two hT2R genes results in different receptor activities and different taste sensitivities to three bitter molecules. The hT2R43 gene allele, which encodes a protein with tryptophan in position 35, makes people very sensitive to the bitterness of the natural plant compounds aloin and aristolochic acid. People who do not possess this allele do not taste these compounds at low concentrations. The same hT2R43 gene allele makes people more sensitive to the bitterness of an artificial sweetener, saccharin. In addition, a closely related gene's (hT2R44's) allele also makes people more sensitive to the bitterness of saccharin. We also demonstrated that some people do not possess certain hT2R genes, contributing to taste variation between individuals. Our findings thus reveal new examples of variations in human taste and provide a molecular basis for them.

Earlier, a family of G protein-coupled receptors, termed T2Rs, was identified in the rodent and human genomes through data mining. It was suggested that these receptors mediate bitter taste perception. Analysis of the human genome revealed that the hT2R family is composed of 25 members. However, bitter ligands have been identified for only three human receptors so far. Here we report identification of two novel ligand-receptor pairs. hT2R61 is activated by 6-nitrosaccharin, a bitter derivative of saccharin. hT2R44 is activated by denatonium and 6-nitrosaccharin. Activation profiles for these receptors correlate with psychophysical data determined for the bitter compounds in human studies. Functional analysis of hT2R chimeras allowed us to identify residues in extracellular loops critical for receptor activation by ligands. The discovery of two novel bitter ligand-receptor pairs provides additional support for the hypothesis that hT2Rs mediate a bitter taste response in humans.

Scheme 1. Schematic illustration of the photothermal genome-editing strategy for cancer immunotherapy. a) Process of preparation of ANP/HSP-Cas9 plasmid complex. b) Illustration of photothermal activation for PD-L1 genome editing in tumor cells. c) Photoactivable CRISPR/Cas9 strategy reprograms immunosuppressive tumor environment.