Optimized vector for functional expression of the human bitter taste receptor TAS2R14 in HEK293 cells.

Abstract

Bitter is one of the five basic taste qualities, along with salty, sour, sweet and umami, used by mammals to access the quality of their food and orient their eating behaviour. Bitter taste detection prevents the ingestion of food potentially contaminated by bitter-tasting toxins. Bitter taste perception is mediated by a family of G protein-coupled receptors (GPCRs) called TAS2Rs. Humans possess 25 TAS2Rs (human type II taste receptors), enabling the detection of thousands of chemically diverse bitter compounds. The identification of agonists/antagonists and molecular mechanisms that govern receptor-ligand interaction has been primarily achieved through functional expression of TAS2Rs in heterologous cells. However, TAS2R receptors, like many other GPCRs, suffer from marginal cell surface expression. In this study, we compared the functionality of 9 engineered chimeric receptors, focusing our experiments on TAS2R14, a broadly tuned receptor that recognizes over 151 identified compounds. Among the different tested signal peptides, rat somatostatin receptor subtype 3 results in higher potency of aristolochic acid-induced calcium signalling than other tested export tags, such as bovine rhodopsin, murine Igκ-chain or human mGluR5. The addition of a MAX sequence enhances both TAS2R14 potency and efficacy. We also confirm that the FLAG epitope, when located at the C-terminal, interferes less with the TAS2R14 functionality, enabling reliable evaluation of this receptor at the cell surface using immunohistochemistry. Finally, these observations are also confirmed for TAS2R14 and TAS1R2/TAS1R3 (the sweet taste receptor) stimulated by 12 bitter compounds and by sucralose and neotame, respectively.