DegU-P represses expression of the motility fla-che operon in Bacillus subtilis
DegU-P represses expression of the motility fla-che operon in Bacillus subtilis
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DOI：
10.1128/jb.186.18.6003-6014.2004
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发表时间：
2004-09-01
影响因子：
3.2
通讯作者：
Galizzi, A
中科院分区：
文献类型：
--
作者：
Amati, G;Bisicchia, P;Galizzi, A
枯草芽孢杆菌实施了几种适应性策略来科普指数生长结束时经历的营养限制。DegS-DegU双组分系统是参与指数后反应调节的网络的一部分，例如感受态发育、胞外酶的产生和运动性。degU 32（Hy）突变延长了磷酸化形式的DegU（DegU-P）的半衰期;这反过来又增加了碱性蛋白酶、果聚糖蔗糖酶和其他胞外酶的产生，并抑制鞭毛的运动和产生。在degU 32（Hy）遗传背景下，鞭毛特异性σ因子Sigl）、鞭毛蛋白基因hag和fla-che操纵子的表达强烈降低。为了研究DegU-P对运动性的作用机制，我们分离了完全抑制运动性缺陷的degU 32（Hy）突变体。通过PCR和测序对突变进行遗传作图和表征。大多数突变被发现删除的主要鞭毛操纵子，fla-che上游的转录终止信号，从而允许从鳕鱼操纵子的转录通读。另外两个突变改善了fla-che操纵子的o（-A）依赖性启动子序列。利用电泳迁移率变动分析，我们已经证明，纯化的DegU特异性结合的Fla-che操纵子的P，启动子区域。这些数据表明DegU抑制鞭毛操纵子的转录，并且它们表明操纵子在调控鞭毛的合成和装配中起中心作用。
Bacillus subtilis implements several adaptive strategies to cope with nutrient limitation experienced at the end of exponential growth. The DegS-DegU two-component system is part of the network involved in the regulation of postexponential responses, such as competence development, the production of exoenzymes, and motility. The degU32(Hy) mutation extends the half-life of the phosphorylated form of DegU (DegU-P); this in turn increases the production of alkaline protease, levan-sucrase, and other exoenzymes and inhibits motility and the production of flagella. The expression of the flagellum-specific sigma factor Sigl), of the flagellin gene hag, and of the fla-che operon is strongly reduced in a degU32(Hy) genetic background. To investigate the mechanism of action of DegU-P on motility, we isolated mutants of degU32(Hy) that completely suppressed the motility deficiency. The mutations were genetically mapped and characterized by PCR and sequencing. Most of the mutations were found to delete a transcriptional termination signal upstream of the main flagellar operon, fla-che, thus allowing transcriptional readthrough from the cod operon. Two additional mutations improved the o(-A)-dependent promoter sequence of the fla-che operon. Using an electrophoretic mobility shift assay, we have demonstrated that purified DegU binds specifically to the P, promoter region of the fla-che operon. The data suggest that DegU represses transcription of thefla-che operon, and they indicate a central role of the operon in regulating the synthesis and assembly of flagella.