Department of Pharmacy, College of Pharmacy, Pusan National University, Busan 609-735, South Korea.
Gene. 2012 Aug 15;505(1):146-52. doi: 10.1016/j.gene.2012.05.004. Epub 2012 May 15.
Bidirectional activation of transcription is a peculiar regulation mode of gene expression. In this study, we show that genes involved in the metabolism of anthranilate, a precursor of biosynthesis of tryptophan and Pseudomonas quinolone signal (PQS) are regulated by this bidirectional activation of transcription. Anthranilate is degraded by anthranilate dioxygenase complex encoded by antABC operon, and AntR, a LysR-type regulator encoded by antR activates the transcription of antABC operon in the presence of anthranilate. In P. aeruginosa, antABC and antR are divergently located and AntR binds to the intergenic region between antA and antR to activate the antABC transcription. In this study, we determined the transcriptional start site of the antA promoter (antA(p)) and AntR-responsive elements (AREs) in P. aeruginosa. The upstream deletion analysis of antA(p) and in vitro gel shift assay with purified AntR showed that there are two AREs at -194 to -148 and -88 to -47 regions. We also found that AntR activates antR promoter (antR(p)) in the opposite direction and both AREs are important in the bidirectional activation of antA(p) and antR(p). Two AREs have different binding affinities to AntR and the strength of transcriptional activation was dramatically asymmetric depending on the direction. We suggest that the different affinities of two AREs may explain the asymmetry of the bidirectional activation by AntR.
双向转录激活是基因表达的一种特殊调控模式。在这项研究中，我们表明，参与色氨酸生物合成前体邻氨基苯甲酸代谢以及假单胞菌喹诺酮信号（PQS）的基因受到这种双向转录激活的调节。邻氨基苯甲酸由编码 antABC 操纵子的邻氨基苯甲酸双加氧酶复合物降解，而 LysR 型调节蛋白 AntR 在邻氨基苯甲酸存在的情况下激活 antABC 操纵子的转录。在铜绿假单胞菌中，antABC 和 antR 是反向定位的，AntR 结合在 antA 和 antR 之间的基因间区域以激活 antABC 转录。在这项研究中，我们确定了铜绿假单胞菌中 antA 启动子（antA(p)）和 AntR 反应元件（AREs）的转录起始位点。antA(p)的上游缺失分析和用纯化的 AntR 进行的体外凝胶迁移实验表明，-194 到-148 和-88 到-47 区域存在两个 AREs。我们还发现 AntR 以相反的方向激活 antR 启动子（antR(p)），并且两个 AREs在 antA(p)和 antR(p)的双向激活中都很重要。两个 AREs 与 AntR 的结合亲和力不同，转录激活的强度取决于方向的不同而存在显著的不对称性。我们认为，两个 AREs 的不同亲和力可能解释了 AntR 双向激活的不对称性。
