Immunologic distribution of an organic anion transport protein in rat liver and kidney
Immunologic distribution of an organic anion transport protein in rat liver and kidney
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DOI：
10.1152/ajpgi.1996.271.2.g231
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发表时间：
1996-08-01
影响因子：
4.5
通讯作者：
Wolkoff, AW
中科院分区：
文献类型：
--
作者：
Bergwerk, AJ;Shi, XY;Wolkoff, AW
最近利用非洲爪哇卵母细胞表达系统从大鼠肝脏克隆了非Na+依赖的有机阴离子转运蛋白[E.Jacqumin，B.Hagenbuch，B.Stieger，A.W.Wolkoff和P.J.Meier，Proc.娜塔莉。阿卡德。SCI。美国91：133-137,1994]。尽管该蛋白的表达足以使细胞转运有机阴离子溴磺化邻苯二甲酸，但对其细胞生物学或生化特性知之甚少。在严格条件下进行的Northern印迹分析表明，仅与来自肝脏和肾脏的RNA杂交；转录本在这两个器官中显示相同。在肾脏内，当使用从外髓提取的RNA时，杂交效果最好。免疫印迹分析表明，在肝脏中，转运蛋白在0.1M Na2CO3提取膜和正弦质膜制剂中得到丰富，与其是一种完整的膜蛋白一致。经N-葡聚糖酶处理后，这一80 kDa的蛋白以65 kDa的形式迁移。肝组织免疫形态学检查显示基底侧质膜定位。在0.1M Na2CO3提取的肾膜中，转运蛋白在非还原十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法(SDS-PAGE)上以83 kDa蛋白的形式迁移。还原时，它分解成33 kDa和37 kDa的多肽。肝蛋白质的SDS-PAGE迁移不受还原影响。肾脏免疫组织学检查显示，顶端质膜位于外髓近端小管的S3段。这种转运蛋白在肝脏和肾脏的不同加工和运输可能会产生重要的功能和调控后果。
Na+-independent organic anion transport protein was recently cloned from rat liver using a Xenopus laevis oocyte expression system [E. Jacquemin, B. Hagenbuch, B. Stieger, A. W. Wolkoff, and P. J. Meier, Proc. Natl. Acad. Sci. USA 91: 133-137, 1994]. Although expression of this protein is sufficient for cells to transport the organic anion bromosulfophthalein, little is known about its cell biology or biochemical characteristics. Northern blot analysis performed under high-stringency conditions revealed hybridization with RNA only from liver and kidney; transcripts appeared the same in these two organs. Within kidney, hybridization was greatest when RNA extracted from the outer medulla was used. Immunoblot analysis revealed that in liver, the transporter was enriched in 0.1 M Na2CO3-extracted membranes and sinusoidal plasma membrane preparations, consistent with its being an integral membrane protein. This 80-kDa protein migrated as a 65-kDa protein after treatment with N-glycanase. Immunomorphological examination of liver revealed basolateral plasma membrane localization. In 0.1 M Na2CO3-extracted membranes of kidney, the transporter migrated as an 83-kDa protein on nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). On reduction, it resolved into peptides of 33 and 37 kDa. SDS-PAGE migration of the liver protein was unaffected by reduction. Immuno-morphological examination of kidney revealed apical plasma membrane localization in the S3 segment of the proximal tubule of the outer medulla. Differential processing and trafficking of this transporter in liver and kidney may have important functional and regulatory consequences.