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具有胆小管特征的大鼠肝脏还原型谷胱甘肽转运体的表达克隆

Expression cloning of a rat hepatic reduced glutathione transporter with canalicular characteristics.

作者信息

Yi J R, Lu S, Fernandez-Checa J, Kaplowitz N

机构信息

Department of Medicine, University of Southern California School of Medicine, Los Angeles 90033.

出版信息

J Clin Invest. 1994 Apr;93(4):1841-5. doi: 10.1172/JCI117170.

DOI:10.1172/JCI117170
PMID:8163683
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC294260/
Abstract

Using the Xenopus oocyte expression system, we have previously identified an approximately 4-kb fraction of mRNA from rat liver that expresses sulfobromophthalein-glutathione (BSP-GSH)-insensitive reduced glutathione (GSH) transport (Fernandez-Checa, J., J. R. Yi, C. Garcia-Ruiz, Z. Knezic, S. Tahara, and N. Kaplowitz. 1993. J. Biol. Chem. 268:2324-2328). Starting with a cDNA library constructed from this fraction, we have now isolated a single clone that expresses GSH transporter activity. The cDNA for the rat canalicular GSH transporter (RcGshT) is 4.05 kb with an open reading frame of 2,505 nucleotides encoding for a polypeptide of 835 amino acids (95,785 daltons). No identifiable homologies were found in searching various databases. An approximately 96-kD protein is generated in in vitro translation of cRNA for RcGshT. Northern blot analysis reveals a single 4-kb transcript in liver, kidney, intestine, lung, and brain. The abundance of mRNA for RcGshT in rat liver increased 3, 6, and 12 h after a single dose of phenobarbital. Insensitivity to BSP-GSH and induction by phenobarbital, unique characteristics of canalicular GSH secretion, suggest that RcGshT encodes for the canalicular GSH transporter.

摘要

利用非洲爪蟾卵母细胞表达系统，我们先前已从大鼠肝脏中鉴定出约4kb的mRNA片段，该片段表达对磺溴酞谷胱甘肽（BSP-GSH）不敏感的还原型谷胱甘肽（GSH）转运（费尔南德斯-切卡，J.，J.R.易，C.加西亚-鲁伊斯，Z.克内齐克，S.田原，和N.卡普洛维茨。1993年。《生物化学杂志》268:2324 - 2328）。从该片段构建的cDNA文库开始，我们现在分离出了一个表达GSH转运蛋白活性的单一克隆。大鼠胆小管GSH转运蛋白（RcGshT）的cDNA为4.05kb，有一个2505个核苷酸的开放阅读框，编码一个835个氨基酸（95,785道尔顿）的多肽。在搜索各种数据库时未发现可识别的同源性。RcGshT的cRNA体外翻译产生一种约96kD的蛋白质。Northern印迹分析显示在肝脏、肾脏、肠道、肺和大脑中有一个单一的4kb转录本。单次给予苯巴比妥后3、6和12小时，大鼠肝脏中RcGshT的mRNA丰度增加。对BSP-GSH不敏感以及受苯巴比妥诱导，这是胆小管GSH分泌的独特特征，表明RcGshT编码胆小管GSH转运蛋白。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/70271ebd9d14/jcinvest00033-0506-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/5c76e939f306/jcinvest00033-0506-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/d2e4dd43b1c7/jcinvest00033-0506-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/70271ebd9d14/jcinvest00033-0506-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/5c76e939f306/jcinvest00033-0506-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/d2e4dd43b1c7/jcinvest00033-0506-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/38d2/294260/70271ebd9d14/jcinvest00033-0506-c.jpg

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本文引用的文献

1
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J Biol Chem. 1993 May 25;268(15):10836-41.
2
Expression of rat liver reduced glutathione transport in Xenopus laevis oocytes.大鼠肝脏还原型谷胱甘肽转运体在非洲爪蟾卵母细胞中的表达
J Biol Chem. 1993 Feb 5;268(4):2324-8.
3
A simple method for displaying the hydropathic character of a protein.一种展示蛋白质亲水性特征的简单方法。
酵母分泌囊泡中还原型谷胱甘肽的ATP依赖性转运。
Biochem J. 1998 Sep 15;334 ( Pt 3)(Pt 3):723-9. doi: 10.1042/bj3340723.
4
Molecular alterations of canalicular transport systems in experimental models of cholestasis: possible functional correlations.胆汁淤积实验模型中小管转运系统的分子改变：可能的功能相关性。
Yale J Biol Med. 1997 Jul-Aug;70(4):365-78.
5
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Yale J Biol Med. 1997 Jul-Aug;70(4):301-10.
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Hepatic glutathione and glutathione S-conjugate transport mechanisms.肝脏谷胱甘肽和谷胱甘肽S-共轭物转运机制。
Yale J Biol Med. 1997 Jul-Aug;70(4):287-300.
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