Human cystinuria-related transporter: Localization and functional characterization
Human cystinuria-related transporter: Localization and functional characterization
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DOI：
10.1046/j.1523-1755.2001.0590051821.x
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发表时间：
2001-05-01
影响因子：
19.6
通讯作者：
Kanai, Y
中科院分区：
文献类型：
--
作者：
Mizoguchi, K;Cha, SH;Kanai, Y
背景胱氨酸尿症被认为是肾近曲小管中胱氨酸和碱性氨基酸的顶膜转运系统的遗传缺陷。AT与非I型胱氨酸尿症有关，人BAT 1（hBAT 1）/B（0.+）AT尚未得到很好的表征。从人肾组织中分离出编码hBAT 1的cDNA。应用荧光原位杂交技术将hBAT 1基因定位在人染色体上。用Northern杂交和免疫组化方法检测其组织分布和表达定位。将hBAT 1 cDNA与rBAT cDNA一起转染COS-7细胞，并测量C-14标记的氨基酸的摄取和流出以确定功能特性。使用蛋白激酶抑制剂或激活剂研究蛋白激酶依赖性磷酸化的作用。将hBAT 1基因定位于人染色体19 q12 -13.1，这是非I型胱氨酸尿症的基因座。hBAT 1信使主要在肾脏表达。hBAT 1蛋白定位于人肾近曲小管顶膜。当在COS-7细胞中表达II型膜糖蛋白rBAT（与B（0.+）相关）时hBAT 1具有氨基酸转运系统B（0.+）的转运活性，推测其通过底物交换机制转运胱氨酸以及碱性和中性氨基酸。BAT介导的转运被蛋白激酶A激活剂所减少，并被酪氨酸激酶激活剂所增强。hBAT 1表现出预期的转运子subserving高亲和力胱氨酸转运系统在肾近端小管的属性。将hBAT 1基因定位于非I型胱氨酸尿症的位点，证实hBAT 1参与胱氨酸尿症。
Background. Cystinuria has been proposed to be an inherited defect of apical membrane transport systems for cystine and basic amino acids in renal proximal tubules, Although the mutations of the recently identified transporter BAT1/b(0.+) AT have been related to nontype I cystinuria, the function and localization of human BAT1 (hBAT1)/b(0.+) AT have not been well characterized.Methods. The cDNA encoding hBAT1 was isolated from human kidney. Fluorescence in situ hybridization was performed to map the hBAT1 gene on human chromosomes. Tissue distribution and localization of expression were examined by North ern blot and immunohistochemical analyses. hBAT1 cDNA was transfected to COS-7 cells with rBAT cDNA, and the uptake and efflux of C-14-labeled amino acids were measured to determine the functional properties. The roles of protein kinase-dependent phosphorylation were investigated using inhibitors or activators sf protein kinases.Results. The hBAT1 gene was mapped to 19q12-13.1 on the human chromosome, which is the locus of nontype I cystinuria. hBAT1 message was expressed predominantly in kidney. hBAT1 protein was localized in the apical membrane of proximal tubules in human kidney. When expressed in COS-7 cells with a type II membrane glycoprotein rBAT (related to b(0.+)-amino acid transporter), hBAT1 exhibited the transport activity with he properties of amino acid transport system b(0.+), which transported cystine as well as basic and neutral amino acids presumably via a substrate exchange mechanism. BAT-mediated transport was reduced by the protein kinase A activator and enhanced by the tyrosine kinase inhibitor.Conclusions. hBAT1 exhibited the properties expected for a transporter subserving the high-affinity cystine transport system in renal proximal tubules. The hBAT1 gene was mapped to the locus of nontype I cystinuria, confirming the involvement of hBAT1 in cystinuria.