An extended role for the nucleoid structuring protein H-NS in the virulence gene regulatory cascade of Shigella flexneri
An extended role for the nucleoid structuring protein H-NS in the virulence gene regulatory cascade of Shigella flexneri
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DOI：
10.1046/j.1365-2958.2003.03347.x
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发表时间：
2003-02-01
影响因子：
3.6
通讯作者：
Dorman, CJ
中科院分区：
文献类型：
--
作者：
Beloin, C;Dorman, CJ
H-NS类核结构蛋白先前已被证明在非允许生长条件下通过抑制virF和virB调节基因以及VirF依赖性icsA结构基因的转录来控制福氏志贺菌中的毒力基因表达中起负面作用。在这里，我们表明，H-NS也作用于VirB依赖的结构基因的启动子的调节级联。H-NS蛋白在体内和体外与启动子区域结合。物理上和通过计算机分析显示启动子含有DNA弯曲区域，这是H-NS结合位点的特征。通过在icsB和virA启动子处的DNA酶I足迹法确定H-NS结合位点。这些位点的位置与H-NS作为转录抑制因子的作用一致。VirB依赖的结构基因启动子被发现直接响应于H-NS阻遏物，揭示了一个控制水平，这是额外的H-NS依赖的virB激活基因所施加的。此外，启动子对细胞中VirB蛋白的水平敏感，在变得活跃之前需要达到VirB的阈值水平。一个模型进行了讨论，在该模型中，结构基因的表达水平反映了H-NS和VirB蛋白的抵消调节活性之间的竞争的结果。
The H-NS nucleoid structuring protein has been shown previously to play a negative role in controlling virulence gene expression in Shigella flexneri by repressing transcription of the virF and virB regulatory genes and the VirF-dependent icsA structural gene under non-permissive growth conditions. Here, we show that H-NS also acts at the promoters of the VirB-dependent structural genes in the regulatory cascade. H-NS protein binds to the promoter regions in vivo and in vitro. The promoters were shown physically and by in silico analysis to contain regions of DNA curvature, a feature of H-NS binding sites. H-NS binding sites were determined by DNase I footprinting at the icsB and the virA promoters. The locations of these sites were consistent with a role for H-NS as a transcription repressor. The VirB-dependent structural gene promoters were found to respond directly to the H-NS repressor, revealing a level of control that is additional to that exerted by the H-NS-dependent virB activator gene. Moreover, the promoters were sensitive to the level of VirB protein in the cell, requiring a threshold level of VirB to be reached before becoming active. A model is discussed in which the levels of expression of the structural genes reflect the outcome of competition between the countervailing regulatory activities of the H-NS and VirB proteins.