Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, K1H8M5, Canada.
Brain and Mind Research Institute, University of Ottawa, Ottawa, ON, K1H8M5, Canada.
Nat Commun. 2023 Jul 5;14(1):3684. doi: 10.1038/s41467-023-39132-x.
Representing the most attractive and successful druggable receptors of the proteome, GPCRs regulate a myriad of physiological and pathophysiological functions. Although over half of present pharmaceuticals target GPCRs, the advancement of drug discovery is hampered by a lack of adequate screening tools, the majority of which are limited to probing agonist-induced G-protein and β-arrestin-2-mediated events as a measure of receptor activation. Here, we develop Tango-Trio, a comprehensive cell-based high-throughput platform comprising cumate-inducible expression of transducers, capable of the parallelized profiling of both basal and agonist-dependent GPCR activities. We capture the functional diversity of GPCRs, reporting β-arrestin-1/2 couplings, selectivities, and receptor internalization signatures across the GPCRome. Moreover, we present the construction of cumate-induced basal activation curves at approximately 200 receptors, including over 50 orphans. Overall, Tango-Trio's robustness is well-suited for the functional characterization and screening of GPCRs, especially for parallel interrogation, and is a valuable addition to the pharmacological toolbox.
作为蛋白质组中最具吸引力和最成功的可成药受体，G 蛋白偶联受体（GPCRs）调节着无数的生理和病理生理功能。尽管目前超过一半的药物都以 GPCR 为靶点，但药物发现的进展受到缺乏足够的筛选工具的阻碍，其中大多数工具仅限于探测激动剂诱导的 G 蛋白和β-arrestin-2 介导的事件，作为受体激活的衡量标准。在这里，我们开发了 Tango-Trio，这是一个包含 cumate 诱导表达的转导器的综合性细胞高通量平台，能够并行分析基础和激动剂依赖性 GPCR 活性。我们捕捉到了 GPCR 的功能多样性，报告了β-arrestin-1/2 偶联、选择性和受体内化特征，涵盖了整个 GPCR 组。此外，我们还构建了大约 200 个受体的 cumate 诱导的基础激活曲线，包括 50 多个孤儿受体。总的来说，Tango-Trio 的稳健性非常适合 GPCR 的功能表征和筛选，特别是用于并行检测，是药理学工具箱的一个有价值的补充。
