Definition of host genes required for intracellular pathogen growth

细胞内病原体生长所需宿主基因的定义

• 批准号: 8019012
• 负责人: Stephen Keith Chapes
• 金额: $ 18.32万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8019012
• 关键词: Address; Adult; Affect; Amblyomma; Anaplasma phagocytophilum; Anthrax disease; Arthropods; Bacillus anthracis; Bacteria; Biochemical; Biochemical Pathway; Categories; Cells; Communicable Diseases; Coxiella burnetii; Data; Disease; Double-Stranded RNA; Down-Regulation; Drosophila genus; Drosophila melanogaster; Ehrlichia; Ehrlichia chaffeensis; Ehrlichiosis; Emerging Communicable Diseases; Gene Expression; Gene Silencing; Genes; Genetic; Goals; Growth; Head; Hemocytes; Human; Immunologist; In Vitro; Infection; Invertebrates; Lead; Libraries; Modeling; Mutation; Organism; Outcome; Pathogenesis; Pathway interactions; Pharmaceutical Preparations; Plague; Principal Investigator; Q Fever; RNA Interference; Rickettsia Infections; Rickettsia rickettsii; Rocky Mountain Spotted Fever; Scientist; Screening procedure; Sorting - Cell Movement; System; Testing; Ticks; United States; Vector-transmitted infectious disease; Vertebrates; Work; Yersinia pestis; fly; fundamental research; gene repression; human granulocytic ehrlichiosis; in vivo; innovation; intracellular parasitism; macrophage; meetings; multidisciplinary; mutant; pathogen; prevent; public health relevance; research study

DESCRIPTION (provided by applicant): The major goal of this project is to define host genes necessary for the replication of an arthropod (tick)-transmitted pathogen, Ehrlichia chaffeensis, in a model arthropod (Drosophila) system. The principal investigators completed a gene expression array analysis of Drosophila S2 cells infected with E. chaffeensis under permissive and non-permissive conditions. This analysis revealed 528 genes that were significantly upregulated (>1.5 fold higher than uninfected control) exclusively during permissive bacterial growth conditions. The working hypothesis is that one or more of these genes are necessary for optimal growth of E. chaffeensis in a host and disruption of those genes will affect bacterial growth. The first specific aim will be to infect adult flies carrying mutations in genes identified by our array. 110 fly lines that carry mutations in genes specifically upregulated during permissive conditions can be obtained as viable-fertile adults. Should the principal investigators identify any genes in this screen, they will use double stranded RNA to silence the gene to validate that silencing of that Drosophila gene will affect the replication of bacteria in Drosophila S2 hemocyte-like cells. The second specific aim will be use an RNAi library to identify host genes that are not expressed in mutant form in adult flies that are also essential for intracellular growth of E. chaffeensis. The availability of comprehensive RNAi screen at the The Drosophila RNAi Screening Center (DRSC) will allow us to investigate how down regulation of genes in S2 cells affects infection. The principal investigators anticipate that they will identify multiple genes that when disrupted will cause lower levels of infection by E. chaffeensis in vivo (in flies) and in vitro (in S2 cells). That outcome will allow the principal investigators to accept their experimental hypothesis and allow them to sort whether the required genes are necessary in a single or multiple biochemical/cellular pathways. The data obtained from this work will allow the principal investigators to focus on the identified genes to determine how manipulation of biochemical pathways with drugs in vertebrate or invertebrate (e.g. tick) systems affect bacterial growth. Therefore, this project presents a highly innovative way to identify host-dependent factors necessary for E. chaffeensis replication and will be useful for identifying translational applications to prevent human monocytic ehrlichiosis and perhaps other Rickettsial diseases. PUBLIC HEALTH RELEVANCE: This is a biomedically related project that addresses which host genes are necessary for the successful replication of an intracellular bacteria that is a human category C pathogen.

描述（由申请人提供）：该项目的主要目标是定义复制节肢动物（tick）传播病原体Ehrllichia chaffeensis所必需的宿主基因，在模型节肢动物（果蝇）系统中。主要研究人员完成了在允许和非耐药性条件下感染了Chaffeensis的果蝇S2细胞的基因表达阵列分析。该分析表明，在允许的细菌生长条件下，仅明显上调的528个基因（比未感染的对照高1.5倍）。有效的假设是，这些基因中的一个或多个对于宿主中的大肠杆菌的最佳生长是必需的，而这些基因的破坏将影响细菌生长。第一个具体目的是感染在我们阵列鉴定的基因中携带突变的成年苍蝇。在允许条件下特异性上调的基因中携带突变的110个飞行线可以作为生存的毛皮成年人获得。如果主要研究人员在此筛选中识别任何基因，他们将使用双链RNA沉默基因来验证该果蝇基因的沉默将影响果蝇S2半细胞样细胞中细菌的复制。第二个具体目的是使用RNAi文库来识别未在成年果蝇中表达的宿主基因，这些基因也对木叶大肠杆菌的细胞内生长也是必不可少的。果蝇RNAi筛查中心（DRSC）的全面RNAi筛选的可用性将使我们能够调查S2细胞中基因的下调如何影响感染。主要研究人员预计，他们将确定多个基因，当受到干扰会导致木叶大肠杆菌在体内（蝇中）和体外（在S2细胞中）的感染水平较低。该结果将使主要研究者能够接受其实验假设，并允许他们对单个或多个生化/细胞途径中所需的基因进行分类。从这项工作中获得的数据将使主要研究人员能够专注于确定的基因，以确定用脊椎动物或无脊椎动物（例如TICK）系统在生化途径中如何操纵生化途径。因此，该项目提出了一种高度创新的方法，可以识别大肠杆菌复制所必需的宿主依赖性因素，并将有助于识别转化应用以防止人类单核细胞性eHrllichiicois和其他立克疾病。 公共卫生相关性：这是一个与生物医学相关的项目，该项目解决了哪些宿主基因成功复制的细胞内细菌是人类病原体所必需的。