Integration of three signals at the Escherichia coli nrf promoter:: a role for Fis protein in catabolite repression
Integration of three signals at the Escherichia coli nrf promoter:: a role for Fis protein in catabolite repression
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DOI：
10.1111/j.1365-2958.2005.04701.x
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发表时间：
2005-07-01
影响因子：
3.6
通讯作者：
Busby, SJW
中科院分区：
文献类型：
--
作者：
Browning, DF;Grainger, DC;Busby, SJW
大肠杆菌nrf操纵子启动子的表达被厌氧触发的转录因子FNR和硝酸盐/亚硝酸盐离子控制的反应调节剂NarL或NarP激活，但被IHF和Fis蛋白抑制。在这里，我们提出了在体外研究的nrf启动子，使用高锰酸盐足迹来测量开放的复合物的形成，和DNA酶I足迹来监测不同的监管机构和它们之间的相互作用的结合。我们的研究结果表明，开放复合物的形成是完全依赖于FNR和增强NarL，但抑制IHF或Fis。NarL对抗IHF的阻遏，但不能改变Fis的阻遏。这些结果表明，nrf启动子活性是由不同的因素调制的机制。已知nrf启动子的表达在丰富培养基中被抑制，特别是在葡萄糖存在下，但其分子基础尚不清楚。在这里，我们表明，这种分解代谢物的抑制是减轻突变，削弱了Fis的DNA位点，提高FNR的DNA位点或提高启动子-10或-35元件。因此，Fis蛋白是负责nrf启动子处的分解代谢物阻遏的主要因子，并且Fis可以覆盖FNR和NarL或NarP的激活。
Expression from the Escherichia coli nrf operon promoter is activated by the anaerobically triggered transcription factor, FNR, and by the nitrate/nitrite ion-controlled response regulators, NarL or NarP, but is repressed by the IHF and Fis proteins. Here, we present in vitro studies on the nrf promoter, using permanganate footprinting to measure open complex formation, and DNase I footprinting to monitor binding of the different regulators and the interactions between them. Our results show that open complex formation is completely dependent on FNR and is enhanced by NarL, but is repressed by IHF or Fis. NarL counteracts repression by IHF but is unable to alter repression by Fis. These results suggest mechanisms by which nrf promoter activity is modulated by the different factors. Expression from the nrf promoter is known to be repressed in rich media, especially in the presence of glucose, but the molecular basis of this is not understood. Here, we show that this catabolite repression is relieved by mutations that weaken the DNA site for Fis, improve the DNA site for FNR or improve the promoter -10 or -35 elements. Hence, Fis protein is a major factor responsible for catabolite repression at the nrf promoter, and Fis can override activation by FNR and NarL or NarP.