摘要
β_2-肾上腺素受体(β_2AR)是一种典型的G-蛋白偶联受体,在调节心跳和心脏收缩等方面起着重要的作用。根据其激活G蛋白的效率,β_2AR的靶向药物通常可分为激动剂、拮抗剂和反向激动剂。近来研究发现一些传统意义的β_2AR拮抗剂和反向激动剂能够作为激动剂激活独立于G-蛋白的信号通路如β-arrestin通路,而β-arrestin信号通路具有心脏保护作用。这种偏向激活概念的出现为设计副作用低的功能选择性药物开辟了新途径,因而迫切需要理解偏向激活的分子机制。我们利用活细胞单分子荧光成像技术测定了GFP标记的β_2AR分子的聚集状态,结合常规生化分析手段,比较了三种临床上常用的心血管疾病治疗药物对β_2AR化学计量的影响,包括偏向配体carvedilol,完全激动剂isoproterenol和反向激动剂propranolol。结果表明:β_2AR在静息状态下主要是以单体形式存在;偏向配体carvedilol刺激后,β_2AR发生β-arrestin介导下的二聚,而其他配体并不引起β_2AR二聚。该工作为研究偏向激活的分子机制提供了新的方法。
The β2-adrenergic receptor(β_2AR),a prototypical member of G protein-coupled receptors which plays an important role in the regulation of cardiac rate and contractility,is a major target of drugs for treatment of heart disease.The emerging concept of β_2AR biased signalling offers a promising strategy for the design of function-selective drugs with low side effects,thus understanding the molecule mechanism of biased signalling is demanding.In this study,by imaging individual green fluorescent protein(GFP) tagged β2AR with live-cell single-molecule fluorescence microscopy,we analysed the stoichiometry of β2AR under the stimulation of three commonly used clinical drugs,the biased ligand carvedilol,as well as the full agonist isoproterenol,and inverse agonist propranolol.The result showed that β2AR mainly existed as monomers at the resting state and exhibitedβ-arrestin-dependent dimerization only upon the treatment of pharmacological biased ligand carvedilol.This work provides a new approach for the study of biased agonism by single-molecule fluorescence imaging
引文
