Ismail-Beigi F, Vanderburg G
Department of Medicine, Division of Clinical and Molecular Endocrinology, Case Western Reserve University, Euclid Avenue, Cleveland, Ohio, 44106-4951, USA.
Arch Biochem Biophys. 1996 Jul 15;331(2):201-7. doi: 10.1006/abbi.1996.0299.
Glucose transport is markedly stimulated in response to inhibition of oxidative phosphorylation by cyanide or azide in Clone 9 cells, a rat liver cell line in which only the GLUT1 isoform of glucose transporters is expressed. Here, we examine the possibility that the stimulation of glucose transport by azide is similarly observed in cells exhibiting high basal rates of glucose transport. We stably transfected Clone 9 cells with an expression plasmid containing full-length rat GLUT1 cDNA; nontransfected cells and cells transfected with plasmid alone served as controls. Two clones of cells transfected with the GLUT1-cDNA-containing insert, labeled A and B, respectively, expressed 8- and 20-fold higher levels of GLUT1 mRNA, contained 11- and 23-fold higher levels of GLUT1, and manifested 11- and 17-fold higher rates of glucose transport in the basal state. Upon incubation with 5 mM azide for 2 h, the rate of glucose transport was markedly stimulated in both clones. Moreover, the transient fall in cell ATP content following exposure to azide did not correlate with the magnitude of the glucose transport response. We conclude that in GLUT1-overexpressing Clone 9 cells (i) GLUT1 content and glucose transport parallel cellular GLUT1 mRNA content, suggesting no major translational or posttranslational control of GLUT1 expression and function in the basal state, and (ii) the rate of glucose transport in cells overexpressing GLUT1 is markedly stimulated by exposure to azide. These results indicate that the stimulation of glucose transport in response to inhibition of oxidative phosphorylation is maintained in cells with very high basal rates of glucose transport.
在克隆9细胞（一种仅表达葡萄糖转运蛋白GLUT1亚型的大鼠肝细胞系）中，氰化物或叠氮化物抑制氧化磷酸化会显著刺激葡萄糖转运。在此，我们研究了在具有高基础葡萄糖转运速率的细胞中是否同样能观察到叠氮化物对葡萄糖转运的刺激作用。我们用含有全长大鼠GLUT1 cDNA的表达质粒稳定转染克隆9细胞；未转染的细胞和仅转染质粒的细胞作为对照。分别标记为A和B的两个转染了含GLUT1 - cDNA插入片段的细胞克隆，其GLUT1 mRNA水平分别高8倍和20倍，GLUT1水平分别高11倍和23倍，并且在基础状态下葡萄糖转运速率分别高11倍和17倍。用5 mM叠氮化物孵育2小时后，两个克隆的葡萄糖转运速率均受到显著刺激。此外，暴露于叠氮化物后细胞ATP含量的短暂下降与葡萄糖转运反应的幅度无关。我们得出结论，在过表达GLUT1的克隆9细胞中：（i）GLUT1含量和葡萄糖转运与细胞GLUT1 mRNA含量平行，表明在基础状态下GLUT1的表达和功能不存在主要的翻译或翻译后调控；（ii）过表达GLUT1的细胞中，暴露于叠氮化物会显著刺激葡萄糖转运速率。这些结果表明，在具有非常高基础葡萄糖转运速率的细胞中，氧化磷酸化抑制引发的葡萄糖转运刺激作用依然存在。
