Identification of binding sites for the group A streptococcal global regulator CovR
Identification of binding sites for the group A streptococcal global regulator CovR
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DOI：
10.1046/j.1365-2958.2002.02810.x
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发表时间：
2002-03-01
影响因子：
3.6
通讯作者：
Scott, JR
中科院分区：
文献类型：
--
作者：
Federle, MJ;Scott, JR
A组链球菌（GAS）的CovRS双组分系统（也称为CsrRS）作为一个全局调节因子，影响至少6个毒力因子的转录。透明质酸胶囊（hasABC操纵子编码的毒力因子）的合成受CovRS负调控。我们证实CovR的磷酸化增加了其与含有hasA启动子的DNA片段的结合。使用DNA酶I足迹法，我们确定了hasA启动子周围从碱基-79到+73的5个结合位点（其中+1是转录的起点）。如尿嘧啶干扰分析所示，每个结合位点内的一对胸腺嘧啶似乎是体外CovR结合所必需的。当通过定点诱变改变这些胸腺嘧啶对中的每一个时，CovR结合在体外减少，证实了每个胸腺嘧啶对在结合中的作用。使用一个转录报告系统与一个单一的染色体拷贝的PhasA-gusA，我们证明了这些结合位点的CovR阻遏的hasA启动子的重要性。基于这些信息，我们提出了CovR与DNA结合的共有序列。
The CovRS two-component system (also called CsrRS) of the group A streptococcus (GAS) acts as a global regulator, influencing the transcription of at least six virulence factors. The synthesis of the hyaluronic acid capsule, a virulence factor encoded by the hasABC operon, is negatively regulated by CovRS. We confirmed that phosphorylation of CovR increases its binding to a DNA fragment containing the hasA promoter. Using DNase I footprinting, we identified five binding sites surrounding the hasA promoter from bases -79 to +73 (where +1 is the start of transcription). One pair of thymines within each binding site appears to be necessary for CovR binding in vitro, as shown by uracil interference analysis. When each of these thymine pairs was altered by site-directed mutagenesis, CovR binding was reduced in vitro, confirming the role of each thymine pair in binding. Using a transcriptional reporter system with a single chromosomal copy of PhasA-gusA, we demonstrated the importance of each of four of these binding sites for CovR repression of the hasA promoter. Based on this information, we propose a consensus sequence for CovR binding to DNA.