Anaplasma phagocytophilum hijacking of host cell monoubiquitination

描述（由申请方提供）：嗜吞噬细胞无形体是一种专性细胞内细菌，可侵入中性粒细胞，引起新发和潜在致死性感染，即人粒细胞无形体病。A.嗜吞噬细胞菌驻留劫持再循环内体，这是细菌生存所必需的过程。这种独特的病原体占据的细胞器如何征用内吞分选还知之甚少。Monoubiquitination最近出现作为一种翻译后修饰，调节内吞分选，特别是回收内体途径。单泛素本身是一种分选信号，用该部分修饰的蛋白质与内吞机制广泛接触。我们发现A.嗜吞噬细胞菌空泡膜积累单泛素，而定位于空泡膜的效应子P100携带三个F盒。F-box基序首先在真核生物中被鉴定，并与SCF泛素连接酶复合物相互作用，SCF泛素连接酶复合物催化蛋白质的泛素化。单核泛素在真核细胞中异位表达时与GFP-P100共定位，在A.嗜吞噬细胞空泡膜含有F-box基序的P100区域暴露在A的胞质表面。嗜吞噬细胞菌空泡膜，并且当其在感染的细胞中异位表达时竞争性地抑制细菌生长。P100是一种新的细菌效应子，因为虽然许多微生物F盒蛋白利用宿主细胞聚泛素化，但P100是第一个选择单泛素化的F盒效应子的例子。我们的研究将检验P100 F盒募集SCF泛素连接酶复合物以指导A.嗜吞噬细胞空泡膜，这样做是重要的细菌生长。在这样做的过程中，我们将破译一个新发现的细菌致病机制，并将推进微生物的策略知识，其中微生物增选泛素途径在不同的，甚至杀微生物，宿主细胞茁壮成长。

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.