Structure and Function of Drosophila NF-kappaB Signaling Pathways

果蝇 NF-κB 信号通路的结构和功能

• 批准号: 9312820
• 负责人: Steven Alexander Wasserman
• 金额: $ 33.7万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9312820
• 关键词: Accelerometer; Address; Animals; Attention; Bacteria; Biochemical; Bioinformatics; Biological Assay; Biological Response Modifiers; Biology; CRISPR/Cas technology; Cluster Analysis; Communicable Diseases; DNA cassette; Defect; Disease Resistance; Drosophila genus; Drosophila melanogaster; Engineering; Epitopes; Family; Family member; Gene Dosage; Gene Expression Profiling; Gene Family; Gene Structure; Gene Transfer Techniques; Genes; Genetic; Genomics; Goals; Gram-Positive Bacteria; Human; Immune; Immune signaling; Immunity; In Vitro; Individual; Infection; Innate Immune Response; Insecta; Invertebrates; Investigation; Light; Link; Maps; Mediating; Microbe; Molecular; Mutagenesis; Mutation; Mycoses; NF-kappa B; Names; Organism; Parasites; Pathway interactions; Pattern; Peptides; Phenotype; Plants; Resistance; Role; Signal Pathway; Signal Transduction; Site-Directed Mutagenesis; Specificity; Structure; Structure-Activity Relationship; Toll-like receptors; Transgenic Organisms; Vertebrates; Virus; antimicrobial; antimicrobial peptide; assault; comparative; design; experimental study; fighting; fungus; gene function; immune function; in vitro activity; in vivo; innovation; insight; interest; loss of function; microbial; mutant; novel; pancreatic secretory trypsin inhibitor I; pathogen; programs; public health relevance; receptor; response; synergism; tool

 DESCRIPTION (provided by applicant): Toll and Toll-like receptors (TLRs) have a central role in innate immune responses in animals ranging from insects to humans. Whereas we know a great deal about the pathogen recognition that initiates Toll signaling and the signaling mechanism itself, we know considerably less about the effectors that mediate innate immune defenses. To address this need, we carried out a cross- species comparative analysis of gene expression to identify conserved features of the Toll- regulated immune repertoire in Drosophila. Focusing our attention on a novel family of immune effectors identified in this manner, we used a state-of-the-art genomic engineering approach to eliminate function in ten of twelve family members. Remarkably, inactivating these ten genes decreased survival upon microbial infection to the same extent and with the same specificity as a complete block in Toll signaling. These findings establish the essential immune function of what we have named the Bom gene family and provide the basis for an original and innovative analysis of effector peptide function. By working in Drosophila, we can readily generate mutations that disrupt pathway activity, monitor and manipulate gene activity, and map out networks of gene function via molecular, biochemical, and bioinformatic approaches. Using a combination of transgenesis and mutagenesis, we will match Bom-mediated defenses to particular pathogens and delineate the structure-function relationships among the Bom peptides. We will then apply these findings to the design and interpretation of in vitro studies of peptide function, focusing on antimicrobial activity. Next, we will explore Bom gene sufficiency in vivo and delineate the extent to which the function of Bom and previously described antimicrobial peptides intersect or overlap. Finally, we will begin mapping out the broader network of Toll mediated defenses by genetic and phenotypic analysis of clusters of other novel effector genes.

 描述（由申请人提供）：Toll 和 Toll 样受体 (TLR) 在从昆虫到人类的动物先天免疫反应中发挥着核心作用。尽管我们对启动 Toll 信号传导的病原体识别以及信号传导机制本身了解很多，但我们对介导先天免疫防御的效应器知之甚少。为了满足这一需求，我们对基因表达进行了跨物种比较分析，以确定果蝇中 Toll 调节的免疫库的保守特征。我们将注意力集中在以这种方式鉴定的新型免疫效应子家族上，使用最先进的基因组工程方法消除了 12 个家族成员中的 10 个成员的功能。值得注意的是，灭活这十个基因会降低微生物感染后的存活率，其程度和特异性与完全阻断 Toll 信号传导相同。这些发现确立了我们称之为 Bom 基因家族的基本免疫功能，并为效应肽功能的原创性和创新性分析提供了基础。通过在果蝇中的研究，我们可以很容易地产生破坏通路活性的突变，监测和操纵基因活性，并通过分子、生物化学和生物信息学方法绘制出基因功能网络。通过结合转基因和诱变，我们将 Bom 介导的防御与特定病原体相匹配，并描绘 Bom 肽之间的结构-功能关系。然后，我们将应用这些发现来设计和解释肽功能的体外研究，重点关注抗菌活性。接下来，我们将探索体内 Bom 基因的充分性，并描绘 Bom 与之前描述的抗菌肽的功能交叉或重叠的程度。最后，我们将开始通过对其他新效应基因簇的遗传和表型分析来绘制更广泛的 Toll 介导的防御网络。