Monitoring ligand-mediated internalization of G protein-coupled receptor as a novel pharmacological approach

被引:38
作者
Fukunaga, Shin'ichi [1 ]
Setoguchi, Shingo [1 ]
Hirasawa, Akira [1 ]
Tsujimoto, Gozoh [1 ]
机构
[1] Kyoto Univ, Grad Sch Pharmaceut Sci, Dept Genom Drug Discovery Sci, Sakyo Ku, Kyoto 6068501, Japan
关键词
G protein-coupled receptor; internalization; fluorescence imaging; orphan receptors;
D O I
10.1016/j.lfs.2006.08.022
中图分类号
R-3 [医学研究方法]; R3 [基础医学];
学科分类号
1001 ;
摘要
Agonist activation of a G protein-coupled receptor (GPCR) results in the redistribution of the receptor protein away from the cell surface into internal cellular compartments through a process of endocytosis known as internalization. Visualization of receptor internalization has become experimentally practicable by using fluorescent reagents such as green fluorescent protein (GFP). In this study, we examined whether the ligand-mediated internalization of a GPCR can be exploited for pharmacological evaluations. We acquired fluorescent images of cells expressing GFP-labeled GPCRs and evaluated the ligand-mediated internalization quantitatively by image processing. Using beta(2)-adrenoceptor and vasopressin V-1a receptor as model GPCRs that couple to Gs and Gq, respectively, we first examined whether these GFP-tagged GPCRs exhibited appropriate pharmacology. The rank order of receptor internalization potency for a variety of agonists and antagonists specific to each receptor corresponded well with that previously observed in ligand binding studies. In addition to chemical ligand-induced internalization, this cell-based fluorescence imaging system successfully monitored the internalization of the proton-sensing GPCR TDAG8, and that of the free fatty acid-sensitive GPCR GPR120. The results show that monitoring receptor internalization can be a useful approach for pharmacological characterization of GPCRs and in fishing for ligands of orphan GPCRs. (c) 2006 Elsevier Inc. All rights reserved.
引用
收藏
页码:17 / 23
页数:7
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