The roles of ceramide and its derivatives in A. phagocytophilum pathogenesis

就发病率而言，专性细胞内细菌是全世界感染性疾病的主要原因， 严重性。它们寄生于宿主细胞代谢物中生存的机制将得到进一步的研究 了解微生物的发病机制，并可能导致开发新的治疗方法， 它们引起的感染。嗜吞噬细胞无形体（Anaplasma phagocytophilum，Ap）是一种专性细胞内细菌， 导致潜在的致命的人畜共患病，人类粒细胞无形体病。我们发现， 劫持富含鞘磷脂的囊泡从trans-Golgi网络（TGN）到其液泡在Rab 10- 以依赖的方式驱动其从非传染性转化为传染性形式。AP感染性子代 富含由酸性鞘磷脂酶（ASMase）产生的宿主鞘脂神经酰胺。 介导的鞘磷脂水解。值得注意的是，宿主ASMase也被路由到Ap液泡（ApV）。 敲低或抑制Rab 10或ASMase可以阻止Ap感染周期， 感染ASMase-/-小鼠。由于Ap寄生于TGN囊泡，因此TGN顺行交通的增加将导致TGN的增加。 受益感染。事实上，我们发现Ap通过上调宿主细胞来诱导这种现象 生产生物活性鞘脂，CERK衍生的神经酰胺-1-磷酸（C1 P）。显然， C1 P通过与UVRAG相互作用诱导高尔基体不稳定和顺行交通诱导。 我们假设Ap诱导高尔基体C1 P形成，其募集UVRAG以诱导 Rab 10阳性、富含鞘磷脂的囊泡的顺行运输被ApV拦截。我们进一步 在ApV中，被劫持的ASMase将鞘磷脂转化为神经酰胺， 后代生产目的1将确定为什么Rab 10对Ap毒力至关重要。目标2将定义 ASMase和神经酰胺在AP病理生物学中的作用。目标3将确定C1 P和UVRAG在 使用新型转基因小鼠模型在体内劫持TGN运输和Ap感染。C1 P作为 最近出现了多种细胞过程（包括癌症和炎症）的有效调节剂。 在这里，我们将首次阐明C1 P在传染病中也起着关键作用。的 我们的研究最终将确定神经酰胺的新的和以前未知的机制 细胞内细菌的寄生。如果我们的假设得到验证，ASMase和CERK将成为靶点。 用于开发新一代的治疗方法来对抗这些类型的病原体。总的来说，这项工作将 具有广泛而强大的影响力。

Obligate intracellular bacteria are major causes of infectious disease worldwide in terms of incidence and severity. Deciphering mechanisms by which they parasitize host cell metabolites to survive will advance understanding of microbial pathogenesis and may lead to development of novel therapeutics for the infections that they cause. Anaplasma phagocytophilum (Ap) is an obligate intracellular bacterium that causes the potentially deadly zoonosis, human granulocytic anaplasmosis. We discovered that Ap hijacks sphingomyelin-rich vesicles from the trans-Golgi network (TGN) to its vacuole in a Rab10- dependent manner to drive conversion from its non-infectious to infectious form. Ap infectious progeny are enriched in the host sphingolipid, ceramide, which is produced by acid sphingomyelinase (ASMase)- mediated hydrolysis of sphingomyelin. Notably, host ASMase is also routed to the Ap vacuole (ApV). Knocking down or inhibiting Rab10 or ASMase arrests the Ap infection cycle, and Ap cannot productively infect ASMase-/- mice. Since Ap parasitizes TGN vesicles, an increase in TGN anterograde traffic would benefit infection. Indeed, we discovered that Ap induces this very phenomenon by upregulating host cell production of the bioactive sphingolipid, CERK-derived ceramide-1-phosphate (C1P). Conspicuously, C1P induces Golgi destabilization and anterograde traffic induction through its interaction with UVRAG. We hypothesize that Ap induces C1P formation at the Golgi, which recruits UVRAG to induce anterograde trafficking of Rab10-positive, sphingomyelin-rich vesicles that the ApV intercepts. We further posit that, at the ApV, hijacked ASMase converts sphingomyelin to ceramide, which drives infectious progeny production. Aim 1 will determine why Rab10 is critical for Ap virulence. Aim 2 will define the roles of ASMase and ceramide in Ap pathobiology. Aim 3 will determine the roles of C1P and UVRAG in hijacking TGN traffic and Ap infection in vivo using novel transgenic mouse models. C1P’s role as a potent regulator of diverse cellular processes including cancer and inflammation has recently emerged. Here, we stand to illuminate for the first time that C1P also plays a critical role in infectious disease. The culmination of our studies will define novel and previously unsurmised mechanisms for ceramide parasitism by intracellular bacteria. If our hypotheses are validated, ASMase and CERK become targets for developing new generations of therapeutics against these types of pathogens. Overall, this work will have a broad and powerful impact.