Orientia tsutsugamushi modulation of host cell ubiquitination machinery

恙虫病东方体对宿主细胞泛素化机制的调节

• 批准号: 8427914
• 负责人: Jason A Carlyon
• 金额: $ 21.5万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-14 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8427914
• 关键词: Acute; Afghanistan; Ankyrin Repeat; Architecture; Asia; Bacteria; Bacterial Proteins; Binding; Binding Proteins; Blood Vessels; Boxing; Case Study; Cell Line; Cell physiology; Cells; Cessation of life; Chiggers; Communicable Diseases; Complex; Data; Disease; Endothelial Cells; Extravasation; F Box Domain; F-Box Motifs; F-Box Proteins; Fever; Funding; Genes; Immune Sera; Infection; Invaded; Knowledge; Lead; Mediating; Middle East; Mites; Morbidity - disease rate; Nature; Orientia tsutsugamushi; Pakistan; Pathogenesis; Proteins; Risk; Role; SKP Cullin F-Box Protein Ligases; Scrub Typhus; Soldier; Structure; Testing; Ubiquitin; Ubiquitination; Vietnam Conflict; Viral; Virus; Work; World War II; cell type; in vivo; killings; macrophage; mortality; multicatalytic endopeptidase complex; neglect; neutrophil; novel; pathogen; protein expression; protein protein interaction; public health relevance; research study; tissue/cell culture; ubiquitin ligase

DESCRIPTION (provided by applicant): Scrub typhus is a neglected disease that threatens the 1 billion inhabitants of the Asia-Pacific rim and causes 1 million new infections annually. Its mortality rate can be as high as 50%. The disease was a significant cause of morbidity in World War II and the Vietnam conflict and presently threatens U.S. soldiers serving in the Middle East. The causative agent is the chigger-transmitted obligate intracellular bacterium, Orientia tsutsugamushi, which colonizes microvascular endothelial cells, macrophages, and neutrophils. The proposed work will advance understanding of how Orientia facilitates its survival in its diverse host cell types, a subject that has long-remained a metaphoric "black box". The ankyrin repeat is the most common protein-protein interaction motif in nature. Many intracellular bacterial and viral pathogens translocate ankyrin repeat-containing proteins (Anks) into eukaryotic host cells. The Anks interact with target proteins to modulate host cell functions. O. tsutsugamushi encodes 38 Anks, several of which also carry another eukaryotic-like motif, the F-box, in their C-termini. We refer to these as Ank/F-box proteins. Eukaryotic F-boxes bind SKP1, a part of the SCF1 ubiquitin ligase complex, which catalyzes the transfer of ubiquitin onto proteins to target them for degradation in the proteasome. The bipartite structure of Ank/F-box proteins conceivably enables them to bind protein substrates via their Ank domains and, using their F-box motifs, bind SKP1 to direct SCF1-mediated ubiquitination and subsequent proteasomal degradation of the substrates. We discovered that 5 of the 8 ank genes that Orientia expresses as it establishes infection in tissue culture cells encode Ank/F-box proteins. In Aim 1, we will evaluate our hypothesis that the Ank/F-box proteins interact with SKP1 to promote SCF1- mediated ubiquitination of host cell proteins. We will also identify the proteins that become ubiquitinated as a result of interacting with Ank/F-box proteins. Given its extensive Ank repertoire, we hypothesize that Orientia expresses specific Anks to establish infection in endothelial cells, macrophages, and neutrophils. In Aim 2, we will close an important knowledge gap by identifying the Anks that the pathogen expresses as it establishes infection in each of the 3 biologically relevant host cells.

描述（由申请人提供）：Scrub Typhus是一种被忽视的疾病，威胁到亚太地区的10亿居民，每年引起100万个新感染。它是 死亡率可以高达50％。该疾病是第二次世界大战和越南冲突中发病率的重要原因，目前威胁到中东服役的美国士兵。致病剂是Chigert的强制性细胞内细菌，东方Tsutsugamushi，它在微血管内皮细胞，巨噬细胞和嗜中性粒细胞中定居。拟议的工作将促进对东方人如何促进其在多样化的宿主细胞类型中生存的理解，该主题长期以来一直是隐喻的“黑匣子”。 Ankyrin重复是本质上最常见的蛋白质 - 蛋白质相互作用基序。许多细胞内细菌和病毒病原体转移了脚踝重复蛋白（ANK）到真核宿主细胞中。 ANK与靶蛋白相互作用以调节宿主细胞功能。 O. tsutsugamushi编码38个ANK，其中一些也带有另一个类似真核的图案，即F-box。我们将其称为ANK/F-box蛋白。真核F-boxes结合SKP1，SKP1是SCF1泛素连接酶复合酶的一部分，该蛋白会催化泛素转移到蛋白质上，以靶向它们在蛋白酶体中降解。 ANK/F-box蛋白的两分结构可以想象，它们能够通过其ANK结构域结合蛋白质底物，并使用其F-box基序结合SKP1，将SKP1结合到底物的SCF1介导的泛素化和随后的蛋白酶体降解。我们发现，在组织培养细胞中建立感染的8个ANK基因中，有5个代表了ANK/F-box蛋白。在AIM 1中，我们将评估ANK/F-box蛋白与SKP1相互作用以促进SCF1介导的宿主细胞蛋白泛素化的假设。我们还将确定由于与ANK/F-box蛋白质相互作用而导致泛素化的蛋白质。鉴于其广泛的ANK曲目，我们假设Orientia表达了在内皮细胞，巨噬细胞和中性粒细胞中建立感染的特定ANK。在AIM 2中，我们将通过确定病原体在3个生物学相关的宿主细胞中建立感染时所表达的ANK来缩小重要的知识差距。