Anaplasma phagocytophilum hijacking of host cell monoubiquitination

嗜吞噬细胞无形体劫持宿主细胞单泛素化

• 批准号: 8637532
• 负责人: Jason A Carlyon
• 金额: $ 22.88万
• 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: 2013-12-10 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8637532
• 关键词: Anaplasma phagocytophilum; Bacteria; Bacterial Proteins; Binding; Bovine Anaplasmosis; C-terminal; Cell Line; Cell physiology; Cells; Complex; Data; Development; Endosomes; Eukaryota; Eukaryotic Cell; F Box Domain; F-Box Motifs; F-Box Proteins; Face; Foundations; Goals; Growth; Human; Infection; Intercept; Invaded; Investigation; Knowledge; Lysine; Mediating; Membrane; Microbe; Monoubiquitination; Mutate; Organelles; Pathway interactions; Polyubiquitin; Polyubiquitination; Post-Translational Protein Processing; Process; Protein Sorting Signals; Proteins; Protocols documentation; Recruitment Activity; Recycling; Research; SKP Cullin F-Box Protein Ligases; Sorting - Cell Movement; Testing; Ubiquitin; Ubiquitination; Vacuole; Work; human disease; microbial; multicatalytic endopeptidase complex; neutrophil; novel; pathogen; polypeptide; prevent; public health relevance

DESCRIPTION (provided by applicant): Anaplasma phagocytophilum is an obligate intracellular bacterium that invades neutrophils to cause the emerging and potentially fatal infection, human granulocytic anaplasmosis. The host cell-derived vacuole in which A. phagocytophilum resides hijacks recycling endosomes, a process that is essential for the bacterium's survival. How this unique pathogen-occupied organelle commandeers endocytic sorting is poorly understood. Monoubiquitination has recently emerged as a posttranslational modification that regulates endocytic sorting, particularly the recycling endosome pathway. Monoubiquitin is itself a sorting signal, and proteins decorated with the moiety make extensive contacts with endocytic machinery. We discovered that the A. phagocytophilum vacuolar membrane accumulates monoubiquitin and that P100, an effector that localizes to the vacuolar membrane, carries three F-boxes. The F-box motif was first identified in eukaryotes and interacts with the SCF ubiquitin ligase complex, which catalyzes ubiquitination of proteins. Monoubiquitin colocalizes with GFP-P100 when it is ectopically expressed in eukaryotic cells and with endogenous P100 on the A. phagocytophilum vacuolar membrane. The P100 region that contains the F-box motif is exposed on the cytosolic face of the A. phagocytophilum vacuolar membrane and competitively inhibits bacterial growth when it is ectopically expressed in infected cells. P100 is a novel bacterial effector because, while many microbial F-box proteins exploit host cell polyubiquitination, P100 is the first example of an F-box effector that co-opts monoubiquitination. Our investigations will test the hypothesis that the P100 F-boxes recruit the SCF ubiquitin ligase complex to direct monoubiquitination of host proteins on the A. phagocytophilum vacuolar membrane and that doing so is important for bacterial growth. In doing so, we will decipher a newly discovered bacterial pathogenic mechanism and will advance knowledge of the strategies by which microbes co-opt ubiquitin pathways to thrive in diverse, even microbiocidal, host cells.

描述（由申请人提供）：嗜吞噬细胞无形体是一种专性细胞内细菌，它侵入中性粒细胞，引起新出现的、可能致命的感染——人粒细胞无形体病。嗜吞噬细胞曲霉所在的宿主细胞衍生的液泡会劫持回收内体，这是细菌生存所必需的过程。人们对这种独特的病原体占据的细胞器如何进行内吞分选知之甚少。单泛素化最近作为一种翻译后修饰出现，调节内吞分选，特别是再循环内体途径。单泛素本身就是一种分选信号，用该部分修饰的蛋白质与内吞机制进行广泛的接触。我们发现 A. phagocytophilum 液泡膜积累单泛素，而 P100（一种定位于液泡膜的效应子）携带三个 F-box。 F-box 基序首先在真核生物中被发现，并与 SCF 泛素连接酶复合物相互作用，催化蛋白质的泛素化。当单泛素在真核细胞中异位表达时，它与 GFP-P100 共定位，并与 A. phagocytophilum 液泡膜上的内源性 P100 共定位。含有 F-box 基序的 P100 区域暴露在嗜吞噬细胞液泡膜的胞质面上，当它在感染细胞中异位表达时，会竞争性抑制细菌生长。 P100 是一种新型细菌效应子，因为虽然许多微生物 F-box 蛋白利用宿主细胞多泛素化，但 P100 是共选择单泛素化的 F-box 效应子的第一个例子。我们的研究将检验以下假设：P100 F-box 招募 SCF 泛素连接酶复合物来指导嗜吞噬细胞囊膜上宿主蛋白的单泛素化，并且这样做对于细菌生长很重要。在此过程中，我们将破译新发现的细菌致病机制，并加深对微生物利用泛素途径在多种甚至杀微生物宿主细胞中繁衍生息的策略的了解。