Defining the pathobiological roles of Orientia tsutsugamushi Ank proteins

确定恙虫病东方体 Ank 蛋白的病理生物学作用

基本信息

批准号：
10455792
负责人：
Jason A Carlyon
金额：
$ 46.57万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-07-12 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10455792
关键词：
ANK1 gene Africa Ankyrin Repeat Antigen-Presenting Cells Asia Bacteria Binding Biological Assay Cell Nucleus Cell physiology Cells Cellular biology Communities Complex Coupled Cowpox Cytoplasm Data Defense Mechanisms Detection Disease Etiology Event F Box Domain F-Box Motifs Fatality rate Funding Gene Expression Genetic Immune Immunity Immunology Immunoprecipitation Impairment Infection Knowledge Lysine Mass Spectrum Analysis Mediating Microbe Middle East Natural Immunity Nature Orientia tsutsugamushi Pathogen detection Pathogenesis Pathway interactions Phosphotransferases Proteins Proteomics Research Role SKP Cullin F-Box Protein Ligases Scrub Typhus South America TNF gene Time Toxic effect Trans-Activators Ubiquitin Ubiquitination Virulence Virulence Factors War Work Yeasts adaptive immunity antimicrobial cell type cohort functional outcomes gain of function global health immunoregulation in vivo inhibitor/antagonist innovation microbial mortality multicatalytic endopeptidase complex neglect novel pathogen pathogenic bacteria prevent protein protein interaction response screening

项目摘要

Orientia tsutsugamushi is an obligate intracellular bacterium and causative agent of scrub typhus, a severe illness with a high fatality rate that results in approximately 1 million new cases each year. Scrub typhus has long been considered endemic to the Asia-Pacific region, but non-travel related cases and detection of the pathogen on multiple continents signify the disease as an emerging global health threat. The ankyrin repeat is one of the most-common protein-protein interaction motifs in nature. O. tsutsugamushi subverts innate and adaptive immunity using its cadre of ankyrin repeat-containing effectors (Anks), many of which also carry a PRANC/F- box motif that co-opts SCF ubiquitin ligases. Ank1 and Ank6 inhibit NF-κB accumulation in the nucleus to impair NF-κB-dependent gene expression in an ankyrin repeat- and PRANC/F-box-dependent manner. Both effectors bind the host cell NF-κB inhibitor, p105, and prevent its TNFα-induced degradation. Conspicuously, Ank1 and Ank6 ubiquitinate Crybg3, a host kinase that influences p105 stability. Further screening revealed that 13 Anks antagonize NF-κB, some of which bind p105 and others of which do not. Thus, multiple O. tsutsugamushi Anks inhibit NF-κB by distinct, overlapping mechanisms. Ank5 promotes ubiquitination and proteasomal degradation of NLRC5, a transactivator of MHC-I gene expression, to potently reduce MHC-I levels. Our findings indicate that Ank immunomodulation enables O. tsutsugamushi to win, at least initially, its tug-of-war with host cells to establish infection. Yet, the detailed mechanisms by which Anks modulate NF-κB, NLRC5, and other targets remain poorly understood. Indeed, while our data establish that O. tsutsugamushi uses Anks to alter the host cell ubiquitome, the extent of this strategy, identity of modified targets, and specific functional outcomes are poorly defined. Due to the bacterium’s genetic intractability, Ank-mediated virulence in vivo has not been pursued. Finally, several additional Anks target unknown host pathways that also likely influence O. tsutsugamushi pathobiology. To fill these knowledge gaps, we propose to (1) decipher the mechanisms by which O. tsutsugamushi Anks inhibit NF-κB and will employ a novel in vivo screen as part of this approach; (2) dissect how Ank5 promotes NLRC5 degradation to block MHC-I expression; and (3) identify new host cell pathways and ubiquitome changes that Anks modulate. The culmination of our studies will define novel and previously unsurmised mechanisms by which intracellular pathogens modulate host immunity. Overall, this work will have a broad and powerful impact.

东方tsutsugamushi是一种强性的细胞内细菌和灾难性的潮湿症状，严重高死亡率的疾病每年大约导致大约100万例新病例。磨砂鼠伤寒很长被认为是亚太地区的内在，但非旅行的病例和病原体的检测在多大洲，这表明该疾病是新兴的全球健康威胁。 Ankyrin重复是自然界中的大多数蛋白质 - 蛋白质相互作用基序。 O. tsutsugamushi颠覆了先天和自适应使用其氨基林重复效应的干部（ANK）的免疫力，其中许多也带有pranc/f- 盒子主题，可以选择SCF泛素连接酶。 ANK1和ANK6抑制核中NF-κB的积累以损害 NF-κB依赖性基因表达以arnkyrin重复和pranc/f-box依赖性方式。两种影响结合宿主细胞NF-κB抑制剂P105，并防止其TNFα诱导的降解。明显地，ank1和 ANK6泛素酸CryBG3，一种影响p105稳定性的宿主激酶。进一步的筛选显示了13个ANK 对抗NF-κB，其中一些结合p105，而另一些则不结合。那是多个O. tsutsugamushi ank 通过不同的重叠机制抑制NF-κB。 ANK5促进泛素化和蛋白酶体降解 NLRC5是MHC-I基因表达的反式激活剂，可潜在地降低MHC-I水平。我们的发现表明 ANK免疫调节使O. tsutsugamushi至少在最初获胜建立感染。然而，ANK调节NF-κB，NLRC5和其他目标的详细机制保持不当理解。确实，当我们的数据建立O. tsutsugamushi使用ANK来改变主机时细胞的泛素，该策略的程度，修改目标的身份以及特定的功能结果是定义不佳。由于细菌的遗传性顽固性，ANK介导的体内病毒尚未追捕。最后，另外几个ANK目标未知的宿主途径也可能影响O。 Tsutsugamushi病理生物学。为了填补这些知识差距，我们建议（1）破译 O. tsutsugamushi Anks抑制NF-κB，并将采用一种新颖的体内筛选作为这种方法的一部分。（2）解剖 ANK5如何促进NLRC5降解以阻止MHC-I表达；（3）确定新的宿主细胞途径和泛素的变化ANKS调制。我们的研究的高潮将定义新颖和以前细胞内病原体调节宿主免疫的无效机制。总的来说，这项工作将有广泛而强大的影响。