The facilitative glucose transport GLUT1 (SLC2A1) is a constitutively expressed membrane protein involved in basal uptake of blood glucose. GLUT1 modification by N-linked glycosylation at a single asparagine residue (N45) appears to play multiple roles in the trafficking, stability and transport activity of this protein. Here we examine the role of complex N-glycosylation on GLUT1 function in renal epithelial cells by arresting this modification at the high-mannose stage with the mannosidase I inhibitor kifunensine. Consistent with prior work in which GLUT1 glycosylation was completely inhibited, we find that kifunensine treatment results in a time-dependent decrease of up to 40% in cellular glucose uptake. We further demonstrate that this effect is primarily a result of deficient GLUT1 trafficking to the cell membrane due to quality control mechanisms that instead direct GLUT1 to the ER-associated degradation (ERAD) pathway. Unlike tunicamycin, which inhibits the first step in N-glycosyl transfer and causes dramatic cell cycle arrest, kifunensine causes only a modest decrease in GLUT1 levels and cell cycle progression in both normal and transformed renal cells. The effect of kifunensine on the cell cycle appears to be independent of its effect on GLUT1, since all renal cell types in this study displayed decreased proliferation regardless of their dependence on glucose uptake for growth and survival. Together these results indicate that proper N-glycan processing plays an important role in directing GLUT1 to the cell surface and that disruption of mannosidase activity results in aberrant degradation of GLUT1 by the ERAD pathway.

中文翻译：

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药理学上抑制用基夫农胺对N联聚糖修饰的作用会破坏GLUT1的运输和葡萄糖的吸收。

促进性葡萄糖转运GLUT1（SLC2A1）是一种组成型表达的膜蛋白，参与基础血糖的吸收。在单个天冬酰胺残基（N45）上通过N-联糖基化修饰GLUT1似乎在该蛋白的运输，稳定性和运输活性中起多种作用。在这里，我们通过用甘露糖苷酶I抑制剂kifunensine阻止这种修饰在高甘露糖阶段的作用，从而研究了复杂的N-糖基化对肾上皮细胞GLUT1功能的作用。与完全抑制GLUT1糖基化的先前工作一致，我们发现kifunensine治疗导致细胞葡萄糖摄取的时间依赖性降低高达40％。我们进一步证明，这种影响主要是由于质量控制机制将GLUT1引导至ER相关降解（ERAD）途径所致，这是由于GLUT1向细胞膜运输不足所致。与衣霉素抑制N-糖基转移的第一步并引起剧烈的细胞周期阻滞不同，其在体内和正常肾细胞中，kifunensine仅引起GLUT1水平的适度降低和细胞周期进程。由于该研究中所有肾细胞类型均显示出增殖减少，而不论其对葡萄糖摄取的生长和存活的依赖性如何，基夫农碱对细胞周期的作用似乎与其对GLUT1的作用无关。