ACIDIFICATION TOLERANCE RESPONSE OF SALMONELLA

沙门氏菌的酸化耐受反应

• 批准号: 3307442
• 负责人: JOHN W FOSTER
• 金额: $ 15.67万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-08-15 至 1996-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3307442
• 关键词: DNA damage; Salmonella infections; Salmonella typhimurium; acidity /alkalinity; adenosinetriphosphatase; environmental stressor; gene expression; genetic operator element; genetic promoter element; homeostasis; host organism interaction; lac operon; membrane permeability; microorganism genetics; molecular cloning; molecular genetics; mutant; regulatory gene; sensory signal detection; stress proteins; structural genes; transposon /insertion element; virulence

Salmonella must endure intervals of acid stress in widely diverse environments ranging from pond water to fused phagolysosomes in infected macrophages. We have discovered a unique survival response that effectively protects the cells from extreme low pH. This acid tolerance response (ATR) is composed of two distinct stages. A pre-acid shock stage triggered below pH 6 and a post-acid shock stage induced below pH 4.5. Both stages are required for the cell to survive low pH (pH 3.3) These findings present a challenging biological question. How does a neutralophillic organism protect itself from severe acid stress? Preliminary results indicate preshock produces an inducible pH homeostasis system that augments the housekeeping system as external pH decreases below pH 4 while acid shock appears to generate a set of proteins designed to minimize acid damage to macromolecules. An adapted culture in which both systems are engaged can survive pH 3.3 stress approximately 1000 fold better than unadapted cultures. Biochemical and genetic approaches have revealed integral roles for the Mg+2 dependent proton-translocating ATPase (atp) and the iron-regulatory locus, fur, in the ATR system. This proposal has three aims designed to investigate the ATR phenomenon at the biochemical, genetic and molecular levels. They are to (1) define the genetic components of the ATR system, (2) reveal the protective mechanisms used and (3) assign specific functions to specific atr loci. Genetic characterization will occur through the generation of acid-sensitive atr insertions (including lacZ operon fusion) and the identification of new regulatory loci. These genes will be cloned and the most intriguing subjected to molecular analysis. As part of this aim, the Salmonella fur product will be scrutinized for properties commensurate to its role in acid tolerance. We will examine the preshock inducible pH homeostasis mechanism by measuring buffering capacity, proton influx and proton efflux in adapted and unadapted cells. Insights regarding post shock protection of cellular macromolecules will be gained by using acid-sensitive indicator proteins and observing acid-damaged DNA repair. The unifying theme throughout this project is to correlate specific atr mutations with the loss of a given repair or protection system. This proposal presents a comprehensive research plan designed to reveal the elegant molecular response of salmonella to low pH. On a higher level, the knowledge gained will address fundamental questions concerning general defense strategies used by micro-organisms to survive harsh environments.

沙门氏菌必须在广泛多样的环境中忍受酸胁迫的间隔 环境从池塘水融合吞噬溶酶体在感染 巨噬细胞 我们发现了一种独特的生存反应， 有效地保护细胞免受极低pH值的影响。这种耐酸性 ATR反应由两个不同的阶段组成。 酸休克前 低于pH 6触发的阶段和低于pH诱导的酸休克后阶段 4.5. 这两个阶段都是细胞在低pH（pH 3.3）下存活所必需的。 这些发现提出了一个具有挑战性的生物学问题。 是如何 嗜中性生物保护自己免受严重的酸胁迫？ 初步结果表明，震前产生诱导pH值 增强管家系统作为外部pH值的内稳态系统 降低到pH 4以下，而酸休克似乎会产生一系列 旨在减少酸对大分子的损害的蛋白质。 适配的 两个系统都参与的培养物可以在pH 3.3胁迫下存活 比未适应的培养物好大约1000倍。 生化和 遗传学方法已经揭示了Mg+2依赖性 质子转运ATP酶（atp）和铁调节位点fur， ATR系统。 该建议有三个目的，旨在研究ATR现象 在生物化学、遗传学和分子水平上。 他们是（1） 定义ATR系统的遗传成分，（2）揭示 （3）保护机制的使用和（3）分配特定的功能， ATR基因座。 遗传特征将通过产生 酸敏感性atr插入（包括lacZ操纵子融合）和 鉴定新的调节基因座。 这些基因将被克隆， 最有趣的是进行分子分析。 作为其中的一部分 目的是对沙门氏菌毛皮产品的特性进行审查 与其在酸耐受性中的作用相称。 我们会研究 通过测量缓冲的震前诱导pH稳态机制 适应和不适应中的质子流入和质子流出 细胞 关于细胞电击后保护的见解 利用酸敏感指示蛋白， 观察酸损伤的DNA修复。 贯穿始终的统一主题 这个项目是将特定的atr突变与a 提供维修或保护系统。 该提案提出了一项全面的研究计划，旨在揭示 saltrin对低pH的优雅分子反应。在较高的pH值下， 在这一层面上，所获得的知识将解决基本问题， 关于微生物生存的一般防御策略 恶劣的环境