OAT2 catalyses efflux of glutamate and uptake of orotic acid
OAT2 catalyses efflux of glutamate and uptake of orotic acid
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DOI：
10.1042/bj20101904
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发表时间：
2011-06-01
影响因子：
4.1
通讯作者：
Gruendemann, Dirk
中科院分区：
文献类型：
--
作者：
Fork, Christian;Bauer, Tim;Gruendemann, Dirk
OAT(有机阴离子转运体)2[人类基因符号SLC22A7(SLC是溶质载体)]是SLC22家族转运蛋白的成员。在大鼠中，OAT2的主要表达部位是肝细胞的肝窦膜域。到目前为止，OAT2在肝脏中的特殊生理功能尚未解决。在本论文中，我们使用LC(LC)-MS差异阴影策略来寻找OAT2的特定底物和跨物种底物。人和大鼠OAT2在人胚胎肾HEK-293细胞中的异源表达刺激两性离子胡芦巴碱的积累；随后，从大鼠(清除量为106mU·min(-1).mg蛋白(-1))和人(46mU·min(-1).mg蛋白(-1))的OAT2被鉴定为OAT2的良好和特异的底物。谷氨酸的摄取驱动力被确认为谷氨酸逆向转运。通过摄取[H-3]谷氨酸，直接证明了OAT2对谷氨酸的有效转运。然而，由于细胞内谷氨酸含量较高，OAT2作为谷氨酸外排转运体发挥作用。因此，即使在没有细胞外交换底物的情况下，OAT2的表达也显著增加了细胞中谷氨酸的释放(LC-MS测量)。乳果酸强烈反式刺激谷氨酸的外流。因此，我们认为OAT2在生理上具有谷氨酸外排转运体的功能。在激光捕获显微解剖大鼠肝脏切片后，同样在门脉周围和中心周围区域检测到OAT2 mRNA；以前关于肝脏向血液中释放谷氨酸的报道现在可以用OAT2活性来解释。一种特异性的OAT2抑制剂可以通过降低血浆谷氨酸，从而促进谷氨酸从脑到血的外流，减轻急性脑状况下的谷氨酸外毒性。
OAT (organic anion transporter) 2 [human gene symbol SLC22A7 (SLC is solute carrier)] is a member of the SLC22 family of transport proteins. In the rat, the principal site of expression of OAT2 is the sinusoidal membrane domain of hepatocytes. The particular physiological function of OAT2 in liver has been unresolved so far. In the present paper, we have used the strategy of LC (liquid chromatography)-MS difference shading to search for specific and cross-species substrates of OAT2. Heterologous expression of human and rat OAT2 in HEK (human embryonic kidney)-293 cells stimulated accumulation of the zwitterion trigonelline; subsequently, orotic acid was identified as an excellent and specific substrate of OAT2 from the rat (clearance = 106 mu l.min(-1).mg of protein(-1)) and human (46 mu l.min(-1).mg of protein(-1)). The force driving uptake of orotic acid was identified as glutamate antiport. Efficient transport of glutamate by OAT2 was directly demonstrated by uptake of [H-3]glutamate. However, because of high intracellular glutamate, OAT2 operates as glutamate efflux transporter. Thus expression of OAT2 markedly increased the release of glutamate (measured by LC-MS) from cells, even without extracellular exchange substrate. Orotic acid strongly trans-stimulated efflux of glutamate. We thus propose that OAT2 physiologically functions as glutamate efflux transporter. OAT2 mRNA was detected, after laser capture microdissection of rat liver slices, equally in periportal and pericentral regions; previous reports of hepatic release of glutamate into blood can now be explained by OAT2 activity. A specific OAT2 inhibitor could, by lowering plasma glutamate and thus promoting brain-to-blood efflux of glutamate, alleviate glutamate exotoxicity in acute brain conditions.