Department of Biochemistry & Cell Biology, Chobanian & Avedisian School of Medicine (R.J., M.G.-M.) and Department of Biology, College of Arts & Sciences (M.G.-M.), Boston University, Boston, Massachusetts.
Department of Biochemistry & Cell Biology, Chobanian & Avedisian School of Medicine (R.J., M.G.-M.) and Department of Biology, College of Arts & Sciences (M.G.-M.), Boston University, Boston, Massachusetts
Mol Pharmacol. 2024 Aug 16;106(3):129-144. doi: 10.1124/molpharm.124.000949.
G protein-coupled receptors (GPCRs) are the largest class of transmembrane receptors encoded in the human genome, and they initiate cellular responses triggered by a plethora of extracellular stimuli ranging from neurotransmitters and hormones to photons. Upon stimulation, GPCRs activate heterotrimeric G proteins (Gγ) in the cytoplasm, which then convey signals to their effectors to elicit cellular responses. Given the broad biological and biomedical relevance of GPCRs and G proteins in physiology and disease, there is great interest in developing and optimizing approaches to measure their signaling activity with high accuracy and across experimental systems pertinent to their functions in cellular communication. This review provides a historical perspective on approaches to measure GPCR-G protein signaling, from quantification of second messengers and other indirect readouts of activity to biosensors that directly detect the activity of G proteins. The latter is the focus of a more detailed overview of the evolution of design principles for various optical biosensors of G protein activity with different experimental capabilities. We will highlight advantages and limitations of biosensors that detect different G protein activation hallmarks, like dissociation of G and Gγ or nucleotide exchange on G, as well as their suitability to detect signaling mediated by endogenous versus exogenous signaling components or in physiologically relevant systems like primary cells. Overall, this review intends to provide an assessment of the state-of-the-art for biosensors that directly measure G protein activity to allow readers to make informed decisions on the selection and implementation of currently available tools. SIGNIFICANCE STATEMENT: G protein activity biosensors have become essential and widespread tools to assess GPCR signaling and pharmacology. Yet, investigators face the challenge of choosing from a growing list of G protein activity biosensors. This review provides an overview of the features and capabilities of different optical biosensor designs for the direct detection of G protein activity in cells, with the aim of facilitating the rational selection of systems that align with the specific scientific questions and needs of investigators.
G 蛋白偶联受体（GPCRs）是人类基因组中编码的最大的跨膜受体家族，它们可以启动细胞对各种细胞外刺激（包括神经递质、激素和光子等）的反应。在受到刺激后，GPCR 会在细胞质中激活异三聚体 G 蛋白（Gγ），然后将信号传递给效应器以引发细胞反应。鉴于 GPCRs 和 G 蛋白在生理学和疾病中的广泛生物学和生物医学相关性，人们非常有兴趣开发和优化方法，以高精度和跨与其在细胞通讯中功能相关的实验系统来测量它们的信号活性。
本综述提供了测量 GPCR-G 蛋白信号的方法的历史视角，从第二信使的定量和其他活性的间接读数到直接检测 G 蛋白活性的生物传感器。后者是对各种 G 蛋白活性生物传感器设计原理的详细概述的重点，这些生物传感器具有不同的实验能力。
我们将重点介绍检测不同 G 蛋白激活特征（如 G 和 Gγ的解离或 G 上的核苷酸交换）的生物传感器的优缺点，以及它们在检测内源性和外源性信号成分介导的信号或在生理相关系统（如原代细胞）中的信号的适用性。
总的来说，本综述旨在评估直接测量 G 蛋白活性的生物传感器的最新技术，以使读者能够在选择和实施当前可用工具时做出明智的决策。
