Regulation and intracellular trafficking pathways of the endothelin receptors
Regulation and intracellular trafficking pathways of the endothelin receptors
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DOI：
10.1074/jbc.m000142200
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发表时间：
2000-06-09
影响因子：
4.8
通讯作者：
Attramadal, H
中科院分区：
文献类型：
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作者：
Bremnes, T;Paasche, JD;Attramadal, H
内皮素（ET）的作用是通过G蛋白偶联受体ETA和ETB介导的。然而，ET受体脱敏，内化和细胞内运输的机制知之甚少。本研究的目的是探讨ET受体调节的分子机制，并表征ET刺激的ETA和ETB受体的细胞内途径。通过分析在存在过表达的β方舟、β-arrestin-1、β-arrestin-2或发动蛋白以及这些调节因子的显性负突变体的情况下，转染的中国仓鼠卵巢细胞中ETA和ETB受体的内化，我们已经证明两种ET受体亚型都遵循抑制蛋白和发动蛋白/网格蛋白依赖性的内化机制。中国仓鼠卵巢和COS细胞表达绿色荧光蛋白（GFP）标记的ET受体的荧光显微镜显示，在ET刺激后，ETA和ETB亚型靶向不同的细胞内途径。虽然ETB-GFP遵循再循环途径并与转铁蛋白共定位在中心粒周围再循环区室中，但ETB-GFP在ET诱导的内化后靶向溶酶体。这两种受体亚型共定位与Rab 5在经典的早期内体，表明该室是一个共同的早期中间体的两个ET受体在细胞内运输。ET刺激的ETA和ETB受体的不同细胞内途径可以解释通过ETA受体的持续信号响应和通过ETB受体的瞬时响应。此外，ETB受体的溶酶体靶向可作为通过该亚型清除血浆内皮素的生化机制。
The effects of endothelin (ET) are mediated via the G protein-coupled receptors ETA and ETB. However, the mechanisms of ET receptor desensitization, internalization, and intracellular trafficking are poorly understood. The aim of the present study was to investigate the molecular mechanisms of ET receptor regulation and to characterize the intracellular pathways of ET-stimulated ETA and ETB receptors, By analysis of ETA and ETB receptor internalization in transfected Chinese hamster ovary cells in the presence of overexpressed beta ARK, beta-arrestin-1, beta-arrestin-2, or dynamin as well as dominant negative mutants of these regulators, we have demonstrated that both ET receptor subtypes follow an arrestin- and dynamin/clathrin-dependent mechanism of internalization. Fluorescence microscopy of Chinese hamster ovary and COS cells expressing green fluorescent protein (GFP)-tagged ET receptors revealed that the ETA and ETB subtypes were targeted to different intracellular routes after ET stimulation. While ETB-GFP followed a recycling pathway and colocalized with transferrin in the pericentriolar recycling compartment, ETB-GFP was targeted to lysosomes after ET-induced internalization. Both receptor subtypes colocalized with Rab5 in classical early endosomes, indicating that this compartment is a common early intermediate for the two ET receptors during intracellular transport. The distinct intracellular routes of ET-stimulated ETA and ETB receptors may explain the persistent signal response through the ETA receptor and the transient response through the ETB receptor. Furthermore, lysosomal targeting of the ETB receptor could serve as a biochemical mechanism for clearance of plasma endothelin via this subtype.