Control of Pseudomonas aeruginosa algD transcription

描述(申请人提供)：囊性纤维化(CF)患者容易被铜绿假单胞菌细菌反复呼吸道感染。这些感染引起的并发症是发病率的主要来源，也是患者死亡的主要原因。最初在肺部定植的铜绿假单胞菌是非粘液性的，但随着时间的推移，粘液性变异体出现，这与CF患者的临床病情恶化有关。粘液表型是由于一种名为海藻酸盐的胶囊多糖的高水平合成，这种毒力因子的过量生产为CF肺中的铜绿假单胞菌提供了一种选择性优势。因此，这项建议的长期目标是了解导致定植于CF患者的铜绿假单胞菌菌株产生藻酸盐的分子机制。 生产藻酸盐的大部分基因都在一个大的操纵子中，该操纵子由一个严格控制的启动子(PalgD)转录。PalgD在粘液型铜绿假单胞菌中被激活，但在非粘液型菌株中检测不到该启动子的转录。这项建议将重点放在海藻D激活所必需的两种蛋白质--海藻Z和海藻B上。此外，还提出了旨在定义导致粘液转化和随后的藻D激活的事件的实验。生物化学和遗传学方法将被用来解决构成这一建议的基础的三个核心问题：(1)AIgZ介导的藻D启动子转录激活的机制是什么？(2)反应调节因子algB如何控制藻D的表达和藻酸盐的合成？(3)控制铜绿假单胞菌粘液转化的分子事件是什么？由于藻酸盐的过量产生与粘液型铜绿假单胞菌定植的CF患者的不良临床结局相关，而且由于藻D激活是藻酸盐合成的先决条件，因此对藻D转录的基本了解对于了解铜绿假单胞菌的发病机制至关重要。这将导致新的治疗方法，并改善居住在粘液性铜绿假单胞菌的CF患者的生活质量。

DESCRIPTION (provided by applicant): Cystic fibrosis (CF) patients are predisposed to recurrent respiratory tract infections by the bacterium Pseudomonas aeruginosa. Complications arising from these infections are a major source of morbidity and the leading cause of death in those afflicted. P. aeruginosa strains that initially colonize the lungs are nonmucoid, but over time mucoid variants emerge and this is correlated with a worsening clinical condition for the CF patient. The mucoid phenotype is due to high-level synthesis of a capsular polysaccharide called alginate and overproduction of this virulence factor confers a selective advantage for P. aeruginosa in the CF lung. Thus, the long-term objective of this proposal is to understand the molecular mechanisms responsible for the production of alginate by strains of P. aeruginosa which colonize CF patients. Most of the genes for alginate production are in a large operon that is transcribed by a tightly controlled promoter (palgD). palgD is activated in mucoid P. aeruginosa isolates but no transcription is detectable from this promoter in nonmucoid strains. This proposal will focus on AlgZ and AlgB, two proteins that are essential for algD activation. In addition, experiments aimed at defining the events leading to mucoid conversion and subsequent algD activation are proposed. Biochemical and genetic approaches will be utilized to address three central questions which constitute the basis of this proposal: (1) What is the mechanism of AIgZ-mediated transcriptional activation of the algD promoter? (2) How does the response regulator AlgB controls algD expression and alginate synthesis? (3) What are the molecular events controlling mucoid conversion in P. aeruginosa? Since the overproduction of alginate correlates with a poor clinical outcome for CF patients colonized with mucoid P. aeruginosa, and since algD activation is a prerequisite for alginate synthesis, a basic understanding of algD transcription is essential for understanding the pathogenesis of P. aeruginosa. This will lead to novel therapies and improve the quality of life for CF patients colonized with mucoid P. aeruginosa.