Odorant response assays for a heterologously expressed olfactory receptor
Odorant response assays for a heterologously expressed olfactory receptor
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DOI：
10.1016/s0006-291x(03)00863-5
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发表时间：
2003-06-13
影响因子：
3.1
通讯作者：
Touhara, K
中科院分区：
文献类型：
--
作者：
Katada, S;Nakagawa, T;Touhara, K
用多种检测方法研究了小鼠嗅觉感受器MOR-EG在不同异种细胞中的嗅觉反应。在共表达MOR-EG和混杂G蛋白Galpha15的HEK293细胞中观察到气味诱导的钙反应。在没有Galpha15的情况下，在各种哺乳动物细胞中观察到气味刺激后cAMP水平的强劲增加。采用Zif268启动子的荧光素酶报告基因分析方法扩增cAMP信号。在非洲爪哇卵母细胞中，当MOR-EG cRNA与Galpha15或cAMP依赖的通道共同注射时，记录到气味刺激电流。这些结果表明，在异种细胞系统中，气味响应性可以通过内源性GalpHas或转基因Galpha15介导的信号通路进行监测。本研究中报道的异源表达的嗅觉受体的各种功能分析，对于高通量的配体筛选和数百种嗅觉受体的功能分析具有潜在的实用价值。(C)2003年埃尔塞维尔科学公司(美国)。版权所有。
Odorant responsiveness of a mouse olfactory receptor, mOR-EG, was investigated in various heterologous cells using a variety of detection methods. Odorant-induced Ca2+ response was observed in HEK293 cells that coexpressed mOR-EG and the promiscuous G protein, Galpha15. Without Galpha15, a robust increase in cAMP level was observed upon odorant-stimulation in various mammalian cells. A luciferase reporter gene assay using zif268 promoter was adopted to amplify the cAMP signals. In Xenopus laeVis oocytes, odorant-stimulated currents were recorded when mOR-EG cRNA was co-injected with either Galpha15 or cAMP-dependent channel. These results suggest that odorant responsiveness can be monitored via a signaling pathway mediated by endogenous Galphas or transfected Galpha15 in heterologous cell systems. Various functional assays for a heterologously expressed olfactory receptor reported in this study, are potentially useful for high-throughput ligand screening and functional analyses of hundreds of olfactory receptors. (C) 2003 Elsevier Science (USA). All rights reserved.