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首页> 外文期刊>Journal of bacteriology >Integration host factor and sequences downstream of the Pseudomonas aeruginosa algD transcription start site are required for expression.
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Integration host factor and sequences downstream of the Pseudomonas aeruginosa algD transcription start site are required for expression.

机译：表达需要铜绿假单胞菌algD转录起始位点下游的整合宿主因子和序列。

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Pseudomonas aeruginosa is an extremely important opportunistic pathogen in immunocompromised individuals. Strains of P. aeruginosa isolated from chronic lung infections in patients with the genetic disease cystic fibrosis have a mucoid colony morphology. This phenotype is due to overproduction of the exopolysaccharide alginate, which is believed to confer a selective advantage on P. aeruginosa in cystic fibrosis lungs. Alginate biosynthesis is controlled by a complex regulatory mechanism. Genes located in the 34-min region of the P. aeruginosa chromosome form an operon which encodes most of the biosynthetic enzymes necessary for alginate production. algD, the first gene in the operon and a critical point for the transcriptional regulation of alginate biosynthesis, is controlled by several trans, cis, and environmental factors. In this study, the involvement of the histone-like protein integration host factor (IHF) in algD expression was examined. Sequences with similarity to consensus IHF-binding sites of Escherichia coli were identified 75 bp upstream (site 1) and 90 bp downstream (site 2) of the start of algD transcription. In gel band mobility shift assays, DNA fragments containing either site bind IHF but site 2 has an approximately 90-fold higher affinity for IHF. Mutations in each of the elements were generated, and they resulted in the reduction or loss of in vitro IHF binding and a three- to fourfold decrease in algD-cat expression. This indicates that IHF binding is necessary for high-level algD transcription. The presence of a high-affinity IHF-binding site located 3' of the algD transcription start site suggested that sequences further downstream of this element are involved in algD expression. When a fragment located downstream of site 2 and upstream of the promoterless cat gene (+110 to +835) was deleted, algD-cat expression was reduced 10-fold supporting the notion that 3' enhancer elements are required for algD transcription. This is the first direct evidence of a 3' element involved in the control of a P. aeruginosa gene. It is postulated that IHF mediates the formation of a higher-order looped structure which is necessary for efficient algD transcription.
机译：铜绿假单胞菌是免疫受损个体中极其重要的机会病原体。从患有遗传性疾病囊性纤维化的患者的慢性肺部感染中分离出的铜绿假单胞菌菌株具有粘液样菌落形态。该表型是由于胞外藻酸盐藻酸盐的过量产生，据信在囊性纤维化肺中赋予铜绿假单胞菌选择性优势。藻酸盐的生物合成受复杂的调节机制控制。位于铜绿假单胞菌染色体34分钟区域的基因形成一个操纵子，该操纵子编码藻酸盐生产所需的大多数生物合成酶。 algD是操纵子中的第一个基因，也是藻酸盐生物合成转录调控的关键点，它受几种反式，顺式和环境因素控制。在这项研究中，检查了组蛋白样蛋白质整合宿主因子（IHF）在algD表达中的参与。在algD转录开始的上游75 bp（位点1）和下游90 bp（位点2）确定了与大肠杆菌的共有IHF结合位点相似的序列。在凝胶带迁移分析中，含有两个位点的DNA片段均与IHF结合，但位点2对IHF的亲和力约为90倍。在每个元素中产生突变，它们导致体外IHF结合减少或丢失，而algD-cat表达减少三到四倍。这表明IHF结合对于高水平algD转录是必需的。位于algD转录起始位点3'处的高亲和力IHF结合位点的存在表明，该元件下游的序列也参与了algD的表达。当缺失位于位点2下游和无启动子cat基因上游（+110至+835）的片段时，algD-cat的表达降低了10倍，从而支持了algD转录需要3'增强子的想法。这是3'元件参与铜绿假单胞菌基因控制的第一个直接证据。据推测，IHF介导了高效algD转录所必需的高级环状结构的形成。

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