PDF(Pubmed)
Abstract：
Class-B1 G-protein-coupled receptors (GPCRs) are an important family of clinically relevant drug targets that remain difficult to investigate via high-throughput screening and in animal models. Here, we engineered PAClight1P78A, a novel genetically encoded sensor based on a class-B1 GPCR (the human PAC1 receptor, hmPAC1R) endowed with high dynamic range (ΔF/F0 = 1100%), excellent ligand selectivity, and rapid activation kinetics (τON = 1.15 s). To showcase the utility of this tool for in vitro applications, we thoroughly characterized and compared its expression, brightness and performance between PAClight1P78A-transfected and stably expressing cells. Demonstrating its use in animal models, we show robust expression and fluorescence responses upon exogenous ligand application ex vivo and in vivo in mice, as well as in living zebrafish larvae. Thus, the new GPCR-based sensor can be used for a wide range of applications across the life sciences empowering both basic research and drug development efforts.
摘要：
B1类G蛋白偶联受体（GPCRs）是临床相关药物靶标的重要家族，难以通过高通量筛选和动物模型进行研究。这里,我们设计了PAClight1P78A,一种基于B1类GPCR（人类PAC1受体，hmPAC1R)具有高动态范围(ΔF/F0=1100%)，优异的配体选择性，和快速活化动力学(τON=1.15s)。为了展示该工具在体外应用的实用性，我们彻底表征和比较了它的表达，PAClight1P78A转染和稳定表达细胞之间的亮度和性能。证明其在动物模型中的用途，我们显示了强大的表达和荧光反应的外源配体应用离体和体内小鼠，以及斑马鱼幼虫。因此，这种基于GPCR的新型传感器可用于生命科学领域的广泛应用，为基础研究和药物开发工作提供了动力。
