Ubiquitylation and Rickettsial Colonization of a Tick Vector

蜱载体的泛素化和立克次体定植

• 批准号: 9188063
• 负责人: Joao Pedra
• 金额: $ 53.97万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-12-01 至 2020-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9188063
• 关键词: Affinity; Amino Acids; Anaplasma phagocytophilum; Area; Arthropod Vectors; Arthropods; Attention; Babesia microti; Babesiosis; Bacteria; Baltimore; Binding; Biological Assay; Biology; Black-legged Tick; Borrelia burgdorferi; Borrelia miyamotoi; Bovine Anaplasmosis; Candidate Disease Gene; Cell Culture Techniques; Communicable Diseases; Communities; Comparative Genomic Analysis; Coupled; Docking; Drosophila genus; Ehrlichia; Ehrlichiosis; Enzymes; FADD gene; Genes; Genome; Glycylglycine; Homologous Protein; Human; Immune; Immune response; Immune signaling; Immune system; Immunity; Infection; Insecta; Ixodes; Knowledge; Laboratories; Lyme Disease; Lysine; Maryland; Mass Spectrum Analysis; Measures; Outcomes Research; Output; Pathway interactions; Pharmacy Schools; Powassan virus; Prevention; Proteins; Proteomics; Publications; RNA Interference; Relapsing Fever; Research; Rickettsia; Rickettsia Infections; Rickettsiales; Signal Pathway; Signal Transduction; Stable Isotope Labeling; Structural Biologist; Testing; Tick-Borne Diseases; Tick-Borne Encephalitis; Ticks; Universities; base; granulocyte; holistic approach; in vivo; infancy; insight; medical schools; microbial; novel; pathogen; public health relevance; response; synergism; tandem mass spectrometry; ubiquitin-protein ligase; vector; x-linked inhibitor of apoptosis protein

 DESCRIPTION (provided by applicant): Comparative genomics analysis coupled to functional assays uncovered evolutionarily conserved signaling pathways and provided important insights towards the understanding of insect immunity. However, extrapolating this approach to non-insect arthropods, such as the tick Ixodes scapularis, sometimes constitutes a problem because of the lack of distinguishable protein homologues and incorrect annotation of genes due to low sequence coverage or incompleteness of a particular genome. For instance, we observed that the immune deficiency (IMD) signaling pathway of the tick I. scapularis is critical for defense against the rickettsial agent Anaplasma phagocytophilum despite the absence of the adaptor molecule imd and the gene fadd on its genome. Moreover, we learned that the E3 ubiquitin ligase x-linked inhibitor of apoptosis protein (XIAP) is critical for activating the IMD pathway against A. phagocytophilum infection of I. scapularis ticks. Collectively, these findings led to ou central hypothesis stating that XIAP regulates the non-canonical tick IMD pathway in response to the rickettsial agent A. phagocytophilum. Accordingly, in Aim #1 of this proposal, we will characterize the functional interaction between XIAP and the E2-conjugating enzyme Bendless. We surmise a direct interaction between these two proteins based on structural docking and mass spectrometry analysis. In Aim #2, we will identify XIAP substrates during pathogen colonization of ticks using stable isotope labeling with amino acids in cell culture (SILAC) and diglycine remnant affinity profiling combined with tandem mass spectrometry. In Aim #3, we will validate the IMD network during A. phagocytophilum infection of I. scapularis ticks in vivo. We will ascertain the immune response following pathogen entry into the tick and evaluate the effect of the IMD signaling pathway during pathogen infection. The outcome of this research will be two-fold: (1) it will provide fundamental knowledge related to tick-pathogen interactions; and (2) it will uncover a novel immune pathway in arthropods with broad implications for the vector biology community.

 描述（由适用提供）：比较基因组学分析与功能测定法相结合的，发现了进化配置的信号通路，并为理解绝缘免疫提供了重要的见解。然而，将这种方法推送到非侵蚀性节肢动物（例如tick ixodes capapularis），有时由于缺乏可区分的蛋白质同源物和由于低序列覆盖率或特定基因组不完整而导致基因的不正确注释，有时构成了问题。例如，我们观察到，tick I的免疫缺陷（IMD）信号通路对于防御抗体剂Anaplasma phapocytophilum的防御至关重要。此外，我们了解到，E3泛素连接酶X连接的凋亡蛋白（XIAP）对于激活抗吞噬菌滴虫的吞噬细胞噬菌体感染的IMD途径至关重要。总的来说，这些发现导致中央假设指出，XIAP响应Rickettsial A. phagocytophilum，调节非典型的tick IMD途径。根据该提案的AIM＃1，我们将表征XIAP与E2偶联酶无弯曲之间的功能相互作用。我们根据结构对接和质谱分析在这两种蛋白质之间进行直接相互作用。在AIM＃2中，我们将使用稳定的同位素标记在细胞培养（SILAC）和Diglycine Remnant亲和力分析中结合串联质谱法的稳定同位素标记在tick虫的病原体定植期间识别XIAP底物。在AIM＃3中，我们将在体内的肩cap虫感染的吞噬细胞感染期间验证IMD网络。我们将确定病原体进入壁虱后的免疫反应，并评估病原体感染期间IMD信号通路的结果。这项研究的结果将是两个方面：（1）它将提供与tick病原体相互作用有关的基本知识； （2）它将在节肢动物中发现一种新颖的免疫途径，对媒介生物学界具有广泛的影响。