Kanai Y, Lee W S, You G, Brown D, Hediger M A
Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115.
J Clin Invest. 1994 Jan;93(1):397-404. doi: 10.1172/JCI116972.
The major reabsorptive mechanism for D-glucose in the kidney is known to involve a low affinity high capacity Na+/glucose cotransporter, which is located in the early proximal convoluted tubule segment S1, and which has a Na+ to glucose coupling ratio of 1:1. Here we provide the first molecular evidence for this renal D-glucose reabsorptive mechanism. We report the characterization of a previously cloned human kidney cDNA that codes for a protein with 59% identity to the high affinity Na+/glucose cotransporter (SGLT1). Using expression studies with Xenopus laevis oocytes we demonstrate that this protein (termed SGLT2) mediates saturable Na(+)-dependent and phlorizin-sensitive transport of D-glucose and alpha-methyl-D-glucopyranoside (alpha MeGlc) with Km values of 1.6 mM for alpha MeGlc and approximately 250 to 300 mM for Na+, consistent with low affinity Na+/glucose cotransport. In contrast to SGLT1, SGLT2 does not transport D-galactose. By comparing the initial rate of [14C]-alpha MeGlc uptake with the Na(+)-influx calculated from alpha MeGlc-evoked inward currents, we show that the Na+ to glucose coupling ratio of SGLT2 is 1:1. Using combined in situ hybridization and immunocytochemistry with tubule segment specific marker antibodies, we demonstrate an extremely high level of SGLT2 message in proximal tubule S1 segments. This level of expression was also evident on Northern blots and likely confers the high capacity of this glucose transport system. We conclude that SGLT2 has properties characteristic of the renal low affinity high capacity Na+/glucose cotransporter as previously reported for perfused tubule preparations and brush border membrane vesicles. Knowledge of the structural and functional properties of this major renal Na+/glucose reabsorptive mechanism will advance our understanding of the pathophysiology of renal diseases such as familial renal glycosuria and diabetic renal disorders.
已知肾脏中D-葡萄糖的主要重吸收机制涉及一种低亲和力、高容量的Na⁺/葡萄糖共转运体，其位于近端小管起始段S1，且Na⁺与葡萄糖的偶联比为1:1。在此，我们为这种肾脏D-葡萄糖重吸收机制提供了首个分子证据。我们报告了一个先前克隆的人肾cDNA的特征，该cDNA编码一种与高亲和力Na⁺/葡萄糖共转运体（SGLT1）有59%同源性的蛋白质。通过对非洲爪蟾卵母细胞进行表达研究，我们证明这种蛋白质（称为SGLT2）介导D-葡萄糖和α-甲基-D-吡喃葡萄糖苷（αMeGlc）的可饱和的、Na⁺依赖性且根皮苷敏感的转运，αMeGlc的Km值为1.6 mM，Na⁺的Km值约为250至300 mM，这与低亲和力Na⁺/葡萄糖共转运一致。与SGLT1不同，SGLT2不转运D-半乳糖。通过比较[¹⁴C]-αMeGlc摄取的初始速率与根据αMeGlc诱发的内向电流计算出的Na⁺内流，我们表明SGLT2的Na⁺与葡萄糖偶联比为1:1。使用原位杂交和免疫细胞化学结合小管段特异性标记抗体，我们证明近端小管S1段中SGLT2信使核糖核酸水平极高。这种表达水平在Northern印迹上也很明显，可能赋予了这个葡萄糖转运系统高容量。我们得出结论，SGLT2具有肾脏低亲和力、高容量Na⁺/葡萄糖共转运体的特性，这与先前对灌注小管制剂和刷状缘膜囊泡的报道一致。了解这种主要的肾脏Na⁺/葡萄糖重吸收机制的结构和功能特性，将有助于我们进一步理解诸如家族性肾性糖尿和糖尿病性肾脏疾病等肾脏疾病的病理生理学。
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