Lühn Kerstin, Laskowska Anna, Pielage Jan, Klämbt Christian, Ipe Ute, Vestweber Dietmar, Wild Martin K
Max Planck Institute for Molecular Biomedicine/Institute of Cell Biology, ZMBE, University of Münster, Germany.
Exp Cell Res. 2004 Dec 10;301(2):242-50. doi: 10.1016/j.yexcr.2004.08.043.
Nucleotide sugar transporters play a central role in the process of glycosylation. They are responsible for the translocation of nucleotide sugars from the cytosol, their site of synthesis, into the Golgi apparatus where the activated sugars serve as substrates for a variety of glycosyltransferases. We and others have recently identified and cloned the first GDP-fucose transporters of H. sapiens and C. elegans. Based on sequence similarity, we could identify a putative homolog in Drosophila melanogaster showing about 45% identity on protein level. The gene (CG9620) encodes a highly hydrophobic, multi-transmembrane spanning protein of 38.1 kDa that is localized in the Golgi apparatus. In order to test whether this protein serves as a GDP-fucose transporter, we performed complementation studies with fibroblasts from a patient with LADII (leukocyte adhesion deficiency II) which exhibit a strong reduction of fucosylation due to a point mutation in the human GDP-fucose transporter gene. We show that transient transfection of these cells with the Drosophila CG9620 cDNA corrects the GDP-fucose transport defect and reestablishes fucosylation. This study gives experimental proof that the product of the in silico identified Drosophila gene CG9620 serves as a functional GDP-fucose transporter.
核苷酸糖转运体在糖基化过程中起着核心作用。它们负责将核苷酸糖从其合成位点胞质溶胶转运至高尔基体，在高尔基体中，活化的糖作为多种糖基转移酶的底物。我们和其他研究人员最近鉴定并克隆了人类和秀丽隐杆线虫的首个GDP-岩藻糖转运体。基于序列相似性，我们在黑腹果蝇中鉴定出一个推定的同源物，其在蛋白质水平上具有约45%的同一性。该基因（CG9620）编码一个高度疏水的、跨膜的38.1 kDa蛋白质，定位于高尔基体。为了测试该蛋白质是否作为GDP-岩藻糖转运体发挥作用，我们用来自一名LADII（白细胞黏附缺陷II型）患者的成纤维细胞进行了互补研究，这些细胞由于人类GDP-岩藻糖转运体基因中的点突变而表现出岩藻糖基化的强烈减少。我们表明，用果蝇CG9620 cDNA瞬时转染这些细胞可纠正GDP-岩藻糖转运缺陷并恢复岩藻糖基化。这项研究提供了实验证据，证明通过计算机鉴定的果蝇基因CG96,20的产物作为一种功能性GDP-岩藻糖转运体发挥作用。
