Huet J, Cottrelle P, Cool M, Vignais M L, Thiele D, Marck C, Buhler J M, Sentenac A, Fromageot P
EMBO J. 1985 Dec 16;4(13A):3539-47. doi: 10.1002/j.1460-2075.1985.tb04114.x.
Fractionation of yeast extracts on heparin-agarose revealed the presence of a DNA footprinting activity that interacted specifically with the 5'-upstream region of TEF1 and TEF2 genes coding for the protein synthesis elongation factor EF-1 alpha, and of the ribosomal protein gene RP51A. The protected regions encompassed the conserved sequences 'HOMOL1' (AACATC TA CG T A G CA) or RPG-box (ACCCATACATT TA) previously detected 200-400 bp upstream of most of the yeast ribosomal protein genes examined. Two types of protein-DNA complexes were separated by a gel electrophoresis retardation assay. Complex 1, formed on TEF1, TEF2 and RP51A 5'-flanking region, was correlated with the protection of a 25-bp sequence. Complex 2, formed on TEF2 or RP51A probes at higher protein concentrations, corresponded to an extended footprint of 35-40 bp. The migration characteristics of the protein-DNA complexes and competition experiments indicated that the same component(s) interacted with the three different promoters. It is suggested that this DNA factor(s) is required for activation and coordinated regulation of the whole family of genes coding for the translational apparatus.
在肝素-琼脂糖上对酵母提取物进行分级分离,结果显示存在一种DNA足迹活性,该活性与编码蛋白质合成延伸因子EF-1α的TEF1和TEF2基因以及核糖体蛋白基因RP51A的5'-上游区域特异性相互作用。受保护区域包含先前在大多数所检测的酵母核糖体蛋白基因上游200 - 400 bp处检测到的保守序列“HOMOL1”(AACATC TA CG T A G CA)或RPG-box(ACCCATACATT TA)。通过凝胶电泳阻滞试验分离出两种类型的蛋白质-DNA复合物。在TEF1、TEF2和RP51A 5'-侧翼区域形成的复合物1与一个25 bp序列的保护相关。在较高蛋白质浓度下在TEF2或RP51A探针上形成的复合物2对应于一个35 - 40 bp的扩展足迹。蛋白质-DNA复合物的迁移特性和竞争实验表明,相同的成分与这三个不同的启动子相互作用。提示这种DNA因子是编码翻译装置的整个基因家族激活和协调调控所必需的。