The human solute carrier gene SLC35B4 encodes a bifunctional nucleotide sugar transporter with specificity for UDP-xylose and UDP-N-acetylglucosamine
The human solute carrier gene SLC35B4 encodes a bifunctional nucleotide sugar transporter with specificity for UDP-xylose and UDP-N-acetylglucosamine
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DOI：
10.1074/jbc.m504783200
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发表时间：
2005-07-22
影响因子：
4.8
通讯作者：
Bakker, H
中科院分区：
文献类型：
--
作者：
Ashikov, A;Routier, F;Bakker, H
核苷酸糖从细胞质转运到高尔基体是由专门的III型蛋白质，核苷酸糖转运蛋白（NST）介导的。在体外与高尔基体囊泡从哺乳动物细胞进行的运输试验表明，具体的摄取共8个核苷酸糖。当这项研究开始时，除了两种核苷酸糖（UDP-Xyl和UDP-Glc）之外，已经克隆了对所有核苷酸糖具有转运活性的NST。为了识别这些难以捉摸的NST，生物信息学方法被用来显示人类基因组中假定的NST序列。10个开放阅读框被鉴定、克隆并在酵母中异源表达。用从转化细胞分离的高尔基体囊泡测定UDP-Glc和UDP-Xyl的转运能力。尽管由于高内源性转运背景而无法鉴定潜在的UDP-Glc转运蛋白，但在零背景下可以测量UDP-Xyl转运。来自表达人类基因SLC 35 B4的酵母细胞的囊泡显示出对UDP-Xyl的特异性摄取，并且随后对其他核苷酸糖的测试揭示了UDP-GlcNAc的第二活性。表位标记的SLC 35 B4在哺乳动物细胞中的表达表现出严格的高尔基体定位。因为已知UDP-GlcA的脱羧直接在内质网和高尔基体腔中产生UDP-Xyl，所以我们的数据证明存在两种方式将UDP-Xyl递送到高尔基体。
The transport of nucleotide sugars from the cytoplasm into the Golgi apparatus is mediated by specialized type III proteins, the nucleotide sugar transporters (NSTs). Transport assays carried out in vitro with Golgi vesicles from mammalian cells showed specific uptake for a total of eight nucleotide sugars. When this study was started, NSTs with transport activities for all but two nucleotide sugars (UDP-Xyl and UDP-Glc) had been cloned. Aiming at identifying these elusive NSTs, bioinformatic methods were used to display putative NST sequences in the human genome. Ten open reading frames were identified, cloned, and heterologously expressed in yeast. Transport capabilities for UDP-Glc and UDP-Xyl were determined with Golgi vesicles isolated from transformed cells. Although a potential UDP-Glc transporter could not be identified due to the high endogenous transport background, the measurement of UDP-Xyl transport was possible on a zero background. Vesicles from yeast cells expressing the human gene SLC35B4 showed specific uptake of UDP-Xyl, and subsequent testing of other nucleotide sugars revealed a second activity for UDP-GlcNAc. Expression of the epitope-tagged SLC35B4 in mammalian cells demonstrated strict Golgi localization. Because decarboxylation of UDP-GlcA is known to produce UDP-Xyl directly in the endoplasmic reticulum and Golgi lumen, our data demonstrate that two ways exist to deliver UDP-Xyl to the Golgi apparatus.