Ferrer-Martínez Andreu, Riera Alberto, Jiménez-Chillarón Josep Carles, Herrero Pilar, Moreno Fernando, Gómez-Foix Anna Maria
Departament de Bioquímica i Biologia Molecular, Universitat de Barcelona, Martí i Franquès, 1. E-08028-Barcelona, Spain.
J Mol Biol. 2004 May 7;338(4):657-67. doi: 10.1016/j.jmb.2004.03.036.
Glucose is an essential nutrient, and a regulator of gene expression in eukaryotic cells. Here, a comparative, function-based genomic approach has been used to identify glucose regulatory elements and transduction pathways common to both yeast and mammalian cells. We have isolated a region in the promoter of the Saccharomyces cerevisiae hexose transporter gene HXT1 that conferred glucose sensitivity in yeast, when located upstream of the minimal CYC1 promoter. This element contained binding motifs for Rgt1, a transcriptional modulator involved in the yeast glucose-induction pathway, that were sufficient to elicit glucose responsiveness. The HXT1 regulatory element was then fused to the minimal cytomegalovirus promoter (HXT1-MIN) and inserted into an adenovirus for delivery to human fibroblasts, where it exhibited glucose-dependent transcriptional activation. Glucose action was mimicked by fructose and unrelated to glucose 6-P content, whilst non-metabolizable glucose analogues showed no effect. Activation of AMP kinase by 5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranosanide blocked glucose induction, revealing parallels with the yeast glucose-repressing pathway. In contrast, delivery of Rgt1 to fibroblasts did not modify HXT1-MIN responsiveness. Thus, elements of the S.cerevisiae HXT1 gene conserve glucose regulation in human fibroblasts equivalent to the metabolism-dependent, glucose-repressing pathway in yeast. These data suggest that the instructions carried within gene regulatory elements controlling nutrient regulation of gene expression have been conserved throughout evolution.
葡萄糖是一种必需营养素，也是真核细胞中基因表达的调节因子。在此，我们采用了一种基于功能的比较基因组学方法，以鉴定酵母和哺乳动物细胞共有的葡萄糖调节元件和转导途径。我们在酿酒酵母己糖转运蛋白基因HXT1的启动子中分离出一个区域，当该区域位于最小CYC1启动子上游时，可赋予酵母葡萄糖敏感性。该元件包含Rgt1的结合基序，Rgt1是参与酵母葡萄糖诱导途径的转录调节因子，这些基序足以引发葡萄糖反应性。然后将HXT1调节元件与最小巨细胞病毒启动子（HXT1-MIN）融合，并插入腺病毒中，用于递送至人成纤维细胞，在该细胞中它表现出葡萄糖依赖性转录激活。果糖可模拟葡萄糖的作用，且与6-磷酸葡萄糖含量无关，而不可代谢的葡萄糖类似物则无作用。5-氨基咪唑-4-甲酰胺-1-β-D-呋喃核糖苷激活AMP激酶可阻断葡萄糖诱导，揭示了与酵母葡萄糖抑制途径的相似之处。相反，将Rgt1递送至成纤维细胞并不会改变HXT1-MIN的反应性。因此，酿酒酵母HXT1基因的元件在人成纤维细胞中保留了葡萄糖调节功能，等同于酵母中依赖代谢的葡萄糖抑制途径。这些数据表明，在整个进化过程中，控制基因表达营养调节的基因调节元件所携带的指令一直得以保留。
