regulation of gene expression in Mannheimia haemolytica A1.

溶血曼海姆菌 A1 基因表达的调控。

• 批准号: 246-2007
• 负责人: Lo, Reggie
• 金额: $ 3.5万
• 依托单位: University of Guelph
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2008
• 资助国家: 加拿大
• 起止时间: 2008-01-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=388225
• 关键词: regulation; gene; expression; Mannheimia; haemolytica

Mannheimia haemolytica A1 (Mh) is an important bacterial pathogen that causes bovine pneumonic pasteurellosis. This respitatory disease results in millions of dollars in economic loss annually to the feedlot cattle industry. Hence, there are continuing interest in understanding the pathogenesis of the bacterium as well as to develop effecive measures to protect animals from the infection. My laboratory has been successful on the characterization of genes that code for many of the virulence factors of the bacterium for more than 20 years. Some of these virulence factors have been produced as recombinant antigens as vaccine components or expressed from transgenic plants in the development of an oral delivery system. On the other hand, the mechanisms that regulate expressin of these virulence genes are not understood. Very little is known about when these virulence genes are expressed during different stages of the infection process. In addition, nothing is known about the enviromental signals that regulate expression of these virulence genes. The objective of this proposal are: i) investigate expression profiles of these genes during an infection in vivo in the animals; and ii) to investigate the mechanisms that regulate expression of the virulence genes. Time course infection experiments will be carried out in which several animals will be infected with Mh and monitored for pneumonic signs. At daily intervals, either nasal samples will be collected or a calf will be sacrificed. Lung washings and lung tissue samples will be collected for extraction of total RNA. The RNA will be used in real-time RT-PCR using Mh specific primers to examine expression of specific genes. It is anticipated that some genes will be expressed early during the infection and some later, depending on the activity of their encoded products in pathogenesis. A second objective is examine the contribution of two-component regulatory systems in regulation of the virulence genes. We have identified several pairs of there systems from the Mh genome sequence. Mutants will be constructed in each of the two cognate proteins to examine the effects on protein expression and the regulatory pathway in response to the two-component system.

溶血曼海姆氏菌 A1 (Mh) 是引起牛肺炎性巴氏杆菌病的重要细菌病原体。这种呼吸道疾病每年给饲养场养牛业造成数百万美元的经济损失。因此，人们对了解这种细菌的发病机制以及制定有效措施来保护动物免受感染一直感兴趣。 20 多年来，我的实验室已经成功地鉴定了编码该细菌许多毒力因子的基因。其中一些毒力因子已作为疫苗成分的重组抗原产生，或在口服递送系统的开发中从转基因植物中表达。另一方面，调节这些毒力基因表达的机制尚不清楚。人们对这些毒力基因在感染过程的不同阶段何时表达知之甚少。此外，对于调节这些毒力基因表达的环境信号一无所知。该提案的目的是：i）研究动物体内感染期间这些基因的表达谱； ii) 研究调节毒力基因表达的机制。将进行时程感染实验，其中几只动物将被 Mh 感染并监测肺炎症状。每隔一天，要么采集鼻腔样本，要么处死小牛。将收集肺洗液和肺组织样本以提取总RNA。 RNA 将用于实时 RT-PCR，使用 Mh 特异性引物来检查特定基因的表达。预计一些基因将在感染早期表达，另一些则稍后表达，具体取决于其编码产物在发病机制中的活性。第二个目标是检查双组分调控系统在毒力基因调控中的贡献。我们已经从 Mh 基因组序列中鉴定出几对系统。将在两种同源蛋白中构建突变体，以检查对蛋白表达的影响以及响应双组分系统的调节途径。