Function and localization of urate transporter 1 in mouse kidney
Function and localization of urate transporter 1 in mouse kidney
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DOI：
10.1097/01.asn.0000107560.80107.19
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发表时间：
2004-02-01
影响因子：
13.6
通讯作者：
Endou, H
中科院分区：
文献类型：
--
作者：
Hosoyamada, M;Ichida, K;Endou, H
小鼠肾特异性转运蛋白cDNA与人尿酸盐转运蛋白1（hURAT 1）的氨基酸序列有74%的同源性，可能是遗传性肾性低尿酸血症基因hURAT 1的小鼠同源物。本研究的目的是利用非洲爪蟾卵母细胞表达系统，研究尿酸转运蛋白在小鼠肾脏中的定位和特性。C-14-尿酸盐的转运以Michaelis-Menten方式进行。RST依赖性尿酸盐转运的Km和Vmax值分别为1213 +/- 222 μ M和268.8 +/- 38.0 pmol/卵母细胞/hr（n = 3）。1 mM丙磺舒（68.7 +/- 9.4%）、50 μ M苯溴马隆（67.9 +/- 6.4%）和10 mM乳酸盐（50.9 +/- 9.5%）显著抑制RST依赖性尿酸盐转运。然而，1 mM对氨基马尿酸盐（PAH），1 mM黄嘌呤和1 mM oxonate并不抑制RST依赖性尿酸盐转运。在外部溶液中用葡萄糖酸盐取代Cl阴离子增强了RST依赖的尿酸盐转运。预先注射吡嗪酸（PZA）或L-乳酸反式刺激RST依赖性尿酸盐转运。用免疫组织化学方法对小鼠肾脏近端小管刷状缘或细胞内膜进行染色，用亲和纯化的抗体识别爪蟾卵母细胞上表达的小鼠URAT 1（mURAT 1）。Western blotting检测到70 kD和62 kD蛋白条带。70 kD蛋白被N-糖基化，并被确定为Triton X-100不溶性刷状缘膜蛋白。雄鼠肾组织中β-淀粉样蛋白mRNA和蛋白水平均高于雌鼠。与hURAT 1相似，mURAT 1转运尿酸盐，因此似乎是mURAT 1-hURAT 1的小鼠同系物。
Mouse renal-specific transporter (RST) cDNA, the amino acid sequence of which has 74% identity with that of human urate transporter 1 (hURAT1), is potentially the mouse homologue of hURAT1, the gene responsible for hereditary renal hypouricemia. The aim of this study is to determine the location and characteristics of RST molecule in mouse kidney and investigate urate transport by RST using the Xenopus oocyte expression system. RST transported C-14-urate in a Michaelis-Menten manner. The K-m and the V-max values of RST-dependent urate transport were 1213 +/- 222 muM and 268.8 +/- 38.0 pmol/oocyte per hr, respectively (n = 3). RST-dependent urate transport was cis-inhibited significantly by 1 mM probenecid (68.7 +/- 9.4%), 50 muM benzbromarone (67.9 +/- 6.4%), and 10 mM lactate (50.9 +/- 9.5%). However, 1 mM p-aminohippurate (PAH), 1 mM xanthine, and 1 mM oxonate did not inhibit RST-dependent urate transport. Substitution of Cl anion with gluconate in the external solution enhanced RST-dependent urate transport. Pre-injected pyrazinoic' acid (PZA) or L-lactate trans-stimulated RST-dependent urate transport. Using inummohistochemistry for mouse kidney, the brush border or intracellular membrane of proximal tubules was stained by an affinity-purified antibody that recognized mouse URAT1 (mURAT1) expressed on Xenopus oocyte. Using Western blotting, anti-mURAT1 antibody detected 70-kD and 62-kD protein bands. The 70-kD protein was N-glycosylated and was identified as a Triton X-100 insoluble brush border membrane protein. RST mRNA and protein levels were higher in male kidneys than female. RST transported urate similar to hURAT1 and, therefore, appears to be mURAT1-the mouse homologue of hURAT1.