THE ROLE OF CELL SHAPE AND CELL WALL IN HELICOBACTER PYLORI PATHOGENESIS

细胞形状和细胞壁在幽门螺杆菌发病机制中的作用

• 批准号: 7638893
• 负责人: Nina Salama
• 金额: $ 21.22万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-15 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7638893
• 关键词: Acidity; Affect; Agar; Animals; Applications Grants; Bacteria; Biochemical; Biochemical Genetics; Campylobacter jejuni; Cell Line; Cell Shape; Cell Wall; Cell Wall Alteration; Cellular Stress; Cellular biology; Chemotaxis; Clinical; Coculture Techniques; Complement; Complex; Cytoskeleton; Defect; Detection; Disease; Elements; Enzymes; Epithelial; Epithelial Cells; Exposure to; Face; Frequencies; Gastritis; Genes; Genetic Epistasis; Goals; Helicobacter; Helicobacter pylori; Human; Immune system; Immunoprecipitation; Infection; Inflammatory; Intestines; Investigation; Learning; Libraries; Life; Link; Liver diseases; Measures; Mediating; Metabolism; Modification; Molecular; Mucous body substance; Mus; Names; Organism; Osmolar Concentration; Outcome; Pathogenesis; Pathway interactions; Peptic Ulcer; Peptidoglycan; Phenotype; Proteins; Role; Shapes; Signal Transduction; Stomach; Stress; Structure; Swimming; Testing; Tropism; Vibrio cholerae; Virulence; Visual; Work; antimicrobial peptide; cellular imaging; cytokine; gene discovery; genetic analysis; insight; malignant stomach neoplasm; microbial; mutant; novel; pathogen; protein function; public health relevance; research study; retinal rods; theories

DESCRIPTION (provided by applicant): Evident in its name, Helicobacter pylori is a helical bacterium that colonizes the human stomach causing clinical outcomes ranging from mild gastritis to peptic ulcer and gastric cancer. Descriptions of H. pylori virulence include a widely accepted hypothesis that the bacterium's spiral shape enhances its ability to swim through the viscous epithelial mucus layer and successfully colonize the stomach. This theory, however, has never been tested and the molecular determinants of H. pylori's shape remain unknown. Using a visual screen of a transposon mutant library, we identified clones displaying two classes of altered cell shape: straight rod and curved vibrioid. Genetic analyses revealed three chromosomal loci, one with at least three genes all affecting cell shape. Biochemical analyses indicate that some of these proteins modify the cell wall peptidoglycan (PG) structure. All mutants tested show slight, but reproducible defects in halo formation in soft agar suggesting compromised swimming or chemotaxis. One vibrioid and two rod-shaped mutants were tested for their ability to colonize the mouse stomach and all showed an altered phenotype compared to wild type or complemented strains. Interestingly the rod-shaped mutants colonized better than their spiral counter parts while the vibrioid mutant showed lower colonization. Thus the identified spiral shape-determining genes influence stomach colonization, but how and why remains to be elucidated. We have two goals with the current grant proposal. The goal of Aim 1 is to understand how the gene products identified in our loss of spirality screen alter cell shape. What is the role of the cell wall? Do the gene products identified within or between shape classes interact? Do they interact with known PG biosynthetic and degrading enzymes or with bacterial cytoskeletal elements? The goal of Aim 2 is to understand how these proteins function during stomach colonization. Are the colonization phenotypes due to altered swimming as previously hypothesized, or is chemotaxis altered? Are the identified cell wall modifications important for the bacteria's ability to survive stresses encountered in the stomach such as fluctuations in osmolarity, acidity and exposure to antimicrobial peptides? Do the cell wall alterations influence PG signaling and/or detection by the host immune system? We believe this work will yield significant advancements in both cell biology and pathogenesis. An outstanding question in microbial cell biology is how and why spiral shape is maintained. The discovery that some of these proteins modify PG is also significant from both cell biology and pathogenesis perspectives. H. pylori's PG activates a major pro-inflammatory pathway via the intracellular pathogen recognition molecule Nod1. Recognition of specific PG components by epithelial cells has been shown to be important for both symbiotic and pathogenic interactions of bacteria and their host. Altogether the experimental aims will probe how and why H. pylori maintains its spiral shape and its role in pathogenesis. PUBLIC HEALTH RELEVANCE: Evident in its name, Helicobacter pylori is a helical bacterium that colonizes the human stomach causing clinical outcomes that range from mild gastritis to peptic ulcer and gastric cancer. We discovered genes that promote spiral shape by this bacterium can alter its cell wall. Study of how these cell wall modifications promote spiral shape and bacterial survival in its unique niche will help us better understand how this bacteria causes disease and discover new ways to inhibit infection.

描述（由申请人提供）：以其名称明显，幽门螺杆菌是一种螺旋细菌，可在人类胃中定居，导致临床结局，从轻度胃炎到消化性溃疡和胃癌。幽门螺杆菌毒力的描述包括一个广泛接受的假设，即细菌的螺旋形状增强了其穿过粘性上皮粘液层游泳并成功地定居胃的能力。然而，该理论从未进行过测试，幽门螺杆菌形状的分子决定因素仍然未知。使用转座子突变库的视觉屏幕，我们确定了显示两类改变的细胞形状的克隆：直杆和弯曲的颤音。遗传分析揭示了三个染色体基因座，一个至少有三个影响细胞形状的基因。生化分析表明，其中一些蛋白质修饰了细胞壁肽聚糖（PG）结构。所有经过测试的突变体均显示出轻微的，但在软琼脂中的光晕形成中可重复的缺陷，暗示游泳或趋化性受损。测试了一种颤动的和两个棒状突变体，其能力定居小鼠胃，并且与野生型或互补菌株相比，所有颤动的突变体都显示出改变的表型。有趣的是，棒状突变体比螺旋柜台植入更好，而纤维化突变体则表现出较低的定殖。因此，鉴定出的螺旋形定基因会影响胃定植，但如何以及为什么要阐明。当前的赠款提案我们有两个目标。 AIM 1的目的是了解如何在我们失去螺旋屏幕筛选中鉴定的基因产物改变细胞形状。细胞壁的作用是什么？形状类内或之间识别的基因产物是否相互作用？它们是否与已知的PG生物合成和降解酶相互作用或与细菌细胞骨架元素相互作用？ AIM 2的目的是了解这些蛋白质在胃定植过程中的功能。以前假设的游泳改变引起的定殖表型是否改变了，还是改变了趋化性？鉴定出的细胞壁修饰对于细菌在胃中遇到的应激能力（例如渗透压，酸度和暴露于抗菌肽的波动）是否重要？细胞壁的改变会影响宿主免疫系统的PG信号传导和/或检测吗？我们认为，这项工作将在细胞生物学和发病机理中取得重大进步。微生物细胞生物学中的一个杰出问题是如何以及为什么保持螺旋形状。从细胞生物学和发病机理的角度来看，这些蛋白质中的某些修饰PG的发现也很重要。幽门螺杆菌的PG通过细胞内病原体识别分子NOD1激活了主要的促炎途径。已证明对细菌及其宿主的共生和致病相互作用对特定的PG成分的识别非常重要。实验目的总共将探测幽门螺杆菌如何以及为什么保持其螺旋形状及其在发病机理中的作用。公共卫生相关性：以其名义显而易见的是，幽门螺杆菌是一种螺旋细菌，可在人类胃中定居，导致临床结局，从轻度胃炎到消化性溃疡和胃癌。我们发现通过这种细菌促进螺旋形状的基因可以改变其细胞壁。研究这些细胞壁修饰如何在其独特的利基市场中促进螺旋形状和细菌存活，将有助于我们更好地了解这种细菌如何引起疾病并发现抑制感染的新方法。