Ligand-induced caveolae-mediated internalization of A1 adenosine receptors:: morphological evidence of endosomal sorting and receptor recycling
Ligand-induced caveolae-mediated internalization of A1 adenosine receptors:: morphological evidence of endosomal sorting and receptor recycling
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DOI：
10.1016/s0014-4827(02)00090-3
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发表时间：
2003-04-15
影响因子：
3.7
通讯作者：
Franco, R
中科院分区：
文献类型：
--
作者：
Escriche, M;Burgueño, J;Franco, R
在平滑肌细胞系DDT 1 MF-2中，通过结合分析、共聚焦显微镜和结构水平研究了小窝在A（1）腺苷受体（A（1）R）内化和受体分选和再循环中的参与。用吸附在金上的霍乱毒素结合亚单位作为特异性探针标记神经节苷脂GM（1），免疫电镜技术表明，激动剂刺激可使腺苷受体在小窝中聚集和隔离。pull-down实验表明，A（1）R的C端结构域与caveolin-1之间存在直接的相互作用。加入外源性腺苷脱氨酶（ADA）（一种与A（1）R结合并作为受体活性修饰蛋白（RAMP）的蛋白质）刺激R-PIA诱导的A（1）受体内化。最后分析了A（1）R/ADA复合物的分选和回收。详细的电子显微镜显示，A（1）R/ADA复合物通过小窝内化在一起，在核内体中进行差异分选，并通过不同组的再循环核内体再循环回细胞表面。这些结果提供了洞察的时空调控和交通的A（1）R和RAMPs。(C)2003 Elsevier Science（美国）。All rights reserved.
The involvement of caveolae in the internalization of A(1) adenosine receptors (A(1)R) and the receptor sorting and recycling was studied in the smooth muscle cell line DDT1MF-2, by binding assays, by confocal microscopy, and at the structural level. The use of cholera toxin-binding subunit adsorbed to gold as a specific probe for labeling the ganglioside GM(1) and immunoelectron microscopy techniques showed that agonist stimulation produced a clustering and sequestration of adenosine receptors in caveolae. Furthermore, pull-down experiments showed there to be a direct interaction between the C-terminal domain of A(1)R and caveolin-1. Addition of exogenous adenosine deaminase (ADA), a protein that binds to A(1)R and acts as a receptor activity modifying protein (RAMP) stimulated R-PIA-induced A(1) receptor internalization. Finally, the sorting and recycling of A(1)R/ADA complexes was analyzed. Detailed electron microscopy revealed that A(1)R/ADA complexes internalize together through caveolae, are differentially sorted in endosomes, and are recycled back to the cell surface by different groups of recycling endosomes. These results give insight into the spatiotemporal regulation and traffic of A(1)R and RAMPs. (C) 2003 Elsevier Science (USA). All rights reserved.