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Abstract：
OrbS and PvdS are extracytoplasmic function (ECF) σ factors that regulate transcription of operons required for the biosynthesis of the siderophores ornibactin and pyoverdine in the Burkholderia cepacia complex and Pseudomonas spp., respectively. Here we show that promoter recognition by OrbS requires specific tetrameric -35 and -10 element sequences that are strikingly similar to those of the consensus PvdS-dependent promoter. However, whereas Pseudomonas aeruginosa PvdS can serve OrbS-dependent promoters, OrbS cannot utilize PvdS-dependent promoters. To identify features present at OrbS-dependent promoters that facilitate recognition by OrbS, we carried out a detailed analysis of the nucleotide sequence requirements for promoter recognition by both OrbS and PvdS. This revealed that DNA sequence features located outside the sigma binding elements are required for efficient promoter utilization by OrbS. In particular, the presence of an A-tract extending downstream from the -35 element at OrbS-dependent promoters was shown to be an important contributor to OrbS specificity. Our observations demonstrate that the nature of the spacer sequence can have a major impact on promoter recognition by some ECF σ factors through modulation of the local DNA architecture.IMPORTANCE ECF σ factors regulate subsets of bacterial genes in response to environmental stress signals by directing RNA polymerase to promoter sequences known as the -35 and -10 elements. In this work, we identify the -10 and -35 elements that are recognized by the ECF σ factor OrbS. Furthermore, we demonstrate that efficient promoter utilization by this σ factor also requires a polyadenine tract located downstream of the -35 region. We propose that the unique architecture of A-tract DNA imposes conformational features on the -35 element that facilitates efficient recognition by OrbS. Our results show that sequences located between the core promoter elements can make major contributions to promoter recognition by some ECF σ factors.
摘要：
OrbS和PvdS是胞质外功能（ECF）σ因子，可调节伯克霍尔德氏菌洋葱复合体和假单胞菌属中铁载体ornibactin和pyoverdine生物合成所需的操纵子的转录。,分别。在这里，我们表明OrbS的启动子识别需要特定的四聚体-35和-10元件序列，这些序列与共有PvdS依赖性启动子的序列非常相似。然而,而铜绿假单胞菌PvdS可以提供OrbS依赖性启动子，OrbS不能利用PvdS依赖性启动子。为了识别存在于OrbS依赖性启动子上的促进OrbS识别的特征，我们对OrbS和PvdS识别启动子的核苷酸序列要求进行了详细分析。这表明，OrbS有效利用启动子需要位于σ结合元件外部的DNA序列特征。特别是,在OrbS依赖性启动子处从-35元件向下游延伸的A-tract的存在被证明是OrbS特异性的重要贡献者。我们的观察结果表明，间隔序列的性质可以通过调节局部DNA结构对某些ECFσ因子的启动子识别产生重大影响。重要性ECFσ因子通过将RNA聚合酶引导至称为-35和-10元件的启动子序列来调节细菌基因的子集以响应环境胁迫信号。在这项工作中,我们确定了由ECFσ因子OrbS识别的-10和-35元素。此外，我们证明，通过该σ因子有效利用启动子还需要位于-35区域下游的聚腺嘌呤束。我们认为，A-tractDNA的独特结构在-35元件上施加了构象特征，从而促进了OrbS的有效识别。我们的结果表明，位于核心启动子元件之间的序列可以通过某些ECFσ因子对启动子识别做出重大贡献。
