Analysis of Fur Regulated Genes in Staphylococcus aureus

金黄色葡萄球菌毛皮调控基因分析

• 批准号: 7012363
• 负责人: RADHESHYAM K JAYASWAL
• 金额: $ 21.45万
• 依托单位: ILLINOIS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7012363
• 关键词: DNA footprinting; Staphylococcus aureus; Staphylococcus infection; bacterial cytopathogenic effect; bacterial genetics; bacterial proteins; binding sites; disease /disorder model; gel mobility shift assay; gene expression; genetic regulation; host organism interaction; iron; kidney disorder; laboratory mouse; microarray technology; microorganism culture; northern blottings; nucleic acid sequence; phenotype; regulatory gene; site directed mutagenesis; virulence

DESCRIPTION (provided by applicant): Staphylococcus aureus is a major cause of human diseases. The acquisition of broad antibiotic resistance in this bacterium, especially, methicillin and vancomycin-resistant S. aureus strains pose a major threat to public health. Various stress conditions within the host are proposed to stimulate the expression of virulence genes in S. aureus. Iron, which has been shown to be an important virulence determinant and required for many vital cellular processes, is not readily available for bacterial cells in the animal host. To cope with an iron-deficient host environment, the bacteria have evolved several mechanisms for iron acquisition regulation. We have cloned and characterized a ferric-uptake regulatory gene (fur) from S. aureus. This gene codes for a protein, Fur, which in the presence of sufficient intracellular iron binds to a specific DNA sequence located in the promoters of sirA and fhu and regulated their expression (Xiong et al., 2000; Cabrera et al., 2001). Our recent DNA microarray analyses indicated that many genes are differentially expressed by iron. Whether these genes are directly or indirectly under the control of Fur-iron cannot be distinguished by the microarray analysis. Therefore I propose to identify some of those genes, which are directly under the control of Fur regulation and involved in the pathogenesis of S. aureus. The long-term objective of this study is to identify Fur regulated genes, which are essential for the survival of S. aureus in the host. The specific goals of this study are: 1) to identify Fur-regulated genes with the fur mutant and the parent strains using microarray analysis followed by northern blot analysis to confirm microarray data, 2) to determine the sequences of Fur binding site of 5-10 genes which are highly regulated by Fur-iron using EMSA and DNase protection assays, and 3) to construct mutations in at least 5 genes to determine their roles in pathogenic phenotype using a mouse kidney abscess model. It is hoped that the results of these studies may find yet unidentified Fur-regulated genes involved in the iron homeostasis and pathogenesis of S. aureus.

描述（由申请人提供）：金黄色葡萄球菌是人类疾病的主要原因。这种细菌获得广泛的抗生素耐药性，特别是耐甲氧西林和万古霉素的金黄色葡萄球菌菌株，对公众健康构成重大威胁。提出了宿主内的各种应激条件来刺激金黄色葡萄球菌毒力基因的表达。铁已被证明是重要的毒力决定因素，并且是许多重要细胞过程所必需的，但动物宿主中的细菌细胞不容易获得铁。为了应对缺铁的宿主环境，细菌进化出了多种铁获取调节机制。我们从金黄色葡萄球菌中克隆并鉴定了铁吸收调节基因（fur）。该基因编码一种蛋白质 Fur，在存在足够的细胞内铁的情况下，该蛋白质与位于 SirA 和 fhu 启动子中的特定 DNA 序列结合并调节其表达（Xiong 等，2000；Cabrera 等，2001） 。我们最近的 DNA 微阵列分析表明，许多基因因铁而差异表达。微阵列分析无法区分这些基因是否直接或间接受Fur-iron控制。因此，我建议鉴定其中一些基因，它们直接受 Fur 调节控制并参与金黄色葡萄球菌的发病机制。这项研究的长期目标是确定毛皮调控基因，这些基因对于金黄色葡萄球菌在宿主中的生存至关重要。本研究的具体目标是：1) 使用微阵列分析鉴定毛皮突变体和亲本菌株的毛皮调控基因，然后进行 Northern 印迹分析以确认微阵列数据，2) 确定 5- 的毛皮结合位点的序列使用 EMSA 和 DNase 保护测定检测 10 个受 Fur-iron 高度调控的基因，以及 3) 使用小鼠肾脓肿模型在至少 5 个基因中构建突变，以确定它们在致病表型中的作用。希望这些研究的结果可能会发现尚未鉴定的毛皮调节基因，这些基因参与金黄色葡萄球菌的铁稳态和发病机制。