Microtransplantation of membranes from cultured cells to Xenopus oocytes:: A method to study neurotransmitter receptors embedded in native lipids
Microtransplantation of membranes from cultured cells to Xenopus oocytes:: A method to study neurotransmitter receptors embedded in native lipids
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DOI：
10.1073/pnas.0438006100
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发表时间：
2003-03-04
影响因子：
11.1
通讯作者：
Eusebi, F
中科院分区：
文献类型：
--
作者：
Palma, E;Trettel, F;Eusebi, F
非洲爪蟾卵母细胞是研究异源性递质受体和离子通道结构和功能的一个方便的细胞表达系统。最近，我们介绍了一种方法，微移植已经组装的神经递质受体从人脑到非洲爪蟾卵母细胞的质膜。同样的方法也用于将培养细胞表达的神经递质受体移植到卵母细胞中。将从稳定表达大鼠谷氨酸受体1的人胚肾细胞系（HEK 293）制备的膜囊泡注射到卵母细胞中，并且在几小时内，卵母细胞质膜获得α-氨基-3-羟基-5-甲基-4-异恶唑丙酸型谷氨酸受体，其具有与原始HEK细胞中表达的谷氨酸受体相同的性质。类似地，用大鼠垂体GH（4）C1细胞制备的膜注射的卵母细胞，稳定表达同型人神经元0型烟碱乙酰胆碱受体（α 7-AcChoR），掺入其质膜AcChoR，表现为GH（4）C1细胞中表达的AcChoR。用稳定表达异聚体人神经元α 4 β 2-AcChoR的HEK细胞获得了类似的结果。所有这些使得非洲爪蟾卵母细胞成为详细研究培养细胞膜中表达的受体和其他蛋白质的有力工具。
The Xenopus oocyte is used as a convenient cell expression system to study the structure and function of heterogenic transmitter receptors and ion channels. Recently, we introduced a method to microtransplant already assembled neurotransmitter receptors from the human brain to the plasma membrane of Xenopus oocytes. The same approach was used here to transplant neurotransmitter receptors expressed from cultured cells to the oocytes. Membrane vesicles prepared from a human embryonic kidney cell line (HEK293) stably expressing the rat glutamate receptor 1 were injected into oocytes, and, within a few hours, the oocyte plasma membrane acquired alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid-type glutamate receptors, which had the same properties as those expressed in the original HEK cells. Analogously, oocytes injected with membranes prepared from rat pituitary GH(4)C1 cells, stably expressing homomeric human neuronal 0 nicotinic acetylcholine receptors (alpha7-AcChoRs), incorporated in their plasma membrane AcChoRs that behaved as those expressed in GH(4)C1 cells. Similar results were obtained with HEK cells stably expressing heteromeric human neuronal alpha4beta2-AcChoRs. All this makes the Xenopus oocyte a powerful tool for detailed investigations of receptors and other proteins expressed in the membrane of cultured cells.