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（由申请人提供）：比较基因组学分析与功能测定相结合，揭示了进化上保守的信号通路，并为理解昆虫免疫提供了重要的见解。然而，将这种方法外推到非昆虫节肢动物，例如肩突硬蜱，有时会产生问题，因为缺乏可区分的蛋白质同源物，并且由于序列覆盖率低或特定基因组不完整而导致基因注释不正确。例如，我们观察到肩胛蜱的免疫缺陷（IMD）信号通路对于防御立克次体病原体无形体吞噬细胞至关重要，尽管其基因组上不存在衔接分子imd和基因fadd。此外，我们还了解到，E3 泛素连接酶 x 连锁凋亡抑制蛋白 (XIAP) 对于激活 IMD 途径以对抗肩胛蜱蜱的嗜吞噬细胞 A. 感染至关重要。总的来说，这些发现得出了一个中心假设，即 XIAP 调节非典型蜱 IMD 途径以响应立克次体病原体 A. phagocytophilum。因此，在本提案的目标#1中，我们将描述 XIAP 和 E2 缀合酶 Bendless 之间的功能相互作用。基于结构对接和质谱分析，我们推测这两种蛋白质之间存在直接相互作用。在目标 #2 中，我们将使用细胞培养物中氨基酸的稳定同位素标记 (SILAC) 以及二甘氨酸残留亲和力分析与串联质谱法相结合来识别蜱病原体定植期间的 XIAP 底物。在目标#3中，我们将在体内肩胛蜱蜱的嗜吞噬细胞 A. 感染过程中验证 IMD 网络。我们将确定病原体进入蜱后的免疫反应，并评估 IMD 信号通路在病原体感染过程中的作用。这项研究的成果将有两个：（1）它将提供与蜱病原体相互作用相关的基础知识； (2)它将揭示节肢动物中的一种新的免疫途径，对媒介生物学界具有广泛的影响。