Characterizing the role of ER-shaping proteins during RNA virus infection

项目摘要 所有的正链RNA病毒都操纵宿主细胞膜，以使其 复制细胞器这些结构被认为集中了所需的细胞和病毒因子 并创造一个环境，帮助保护病毒免受先天免疫传感器的侵害。 肠道病毒，如柯萨奇病毒B3（CVB）和黄病毒，如登革病毒（DENV），代表 诱导形成不同复制类型的两个正链RNA病毒家族 细胞器肠道病毒感染导致细胞质单膜和双膜囊泡的形成 含有内质网（ER）和自噬标记。肠道病毒复制蛋白锚定在 在这些囊泡的外膜小叶上，它们协调病毒基因组的复制。除了 感染人类的大多数黄病毒必须在昆虫载体如蚊子和蜱中有效复制。 黄病毒诱导ER的囊泡内陷的形成，其含有通向细胞的小开口。 细胞质基因组复制在这些结构内由病毒非结构蛋白介导， 都集中在囊泡中尽管有证据表明急诊室参与了建立 对于这些不同的复制细胞器，缺乏关于ER相关的作用的知识。 肠病毒和黄病毒复制过程中的膜成形调节因子。ER是一个大型的，动态的 一种细胞器，以核周片和细胞质小管网络的形式存在，其中包括一个 连续管腔。这个细胞器的动力学和独特的形状是由几个表达调控的， ER形成蛋白的类别，其中许多是保守的从酵母到人类。我的初步结果 表明这些蛋白质中几种充当CVB和DENV感染的调节剂。因此，我们假设 肠道病毒和黄病毒靶向特定的ER形成蛋白以促进其复制。 第一组拟议的研究旨在描述网状结构、ER形成和细胞增殖的机制。 促进膜弯曲的蛋白质，调节CVB和DENV的感染。此外，我们将 确定昆虫网状蛋白同源物在昆虫细胞的DENV感染期间的作用。第二组 提出的研究将表征FAM 134 B在DENV感染期间的作用。FAM 134 B是一款 确定ER-成形蛋白，调节自噬介导的ER降解（ER-吞噬）。结果 这些研究将提供新的宿主-病原体相互作用的细节，介导两种RNA的复制 病毒，这是一个重大的公共卫生负担。从我们的结果中获得的信息将提供 对这些病毒的生物学有重要的了解，可用于开发抗病毒药物， 对抗疾病。此外，我们的研究结果也将有助于确定ER形成蛋白的具体作用， 调节ER的形状和动力学。

PROJECT SUMMARY All positive-stranded RNA viruses manipulate host cell membranes for the biogenesis of their replication organelles. These structures are thought to concentrate cellular and viral factors that are required for replication and create an environment that helps to protect the virus from innate immune sensors. Enteroviruses, such as coxsackievirus B3 (CVB), and flaviviruses, such as dengue virus (DENV), represent two families of positive-stranded RNA viruses that induce the formation of distinct classes of replication organelles. Enterovirus infection results in the formation of cytoplasmic single- and double-membrane vesicles that contain endoplasmic reticulum (ER) and autophagic markers. Enteroviral replication proteins are anchored on the outer membrane leaflets of these vesicles, where they coordinate viral genome replication. In addition to infecting humans, most flaviviruses must efficiently replicate in insect vectors, such as mosquitoes and ticks. Flaviviruses induce the formation of vesicular invaginations of the ER that contain a small opening to the cytoplasm. Genome replication is mediated within these structures by the viral nonstructural proteins, which are concentrated in the vesicles. Despite the evidence that exists for the involvement of the ER in establishing these distinct replication organelles there is a lack of knowledge regarding the role of ER-associated membrane shaping regulators during enterovirus and flavivirus replication. The ER is a large, dynamic organelle that exists as perinuclear sheets and a cytoplasmic network of tubules, which encompass a contiguous lumen. The dynamics and unique shape of this organelle are regulated by the expression of several classes of ER-shaping proteins, many of which are conserved from yeast to humans. My preliminary results indicate that several of these proteins act as regulators of CVB and DENV infection. Thus, we hypothesize that enteroviruses and flaviviruses target specific ER-shaping proteins to facilitate their replication. The first set of proposed studies are designed to delineate the mechanisms by which reticulons, ER-shaping proteins that promote membrane curvature, regulate infection by CVB and DENV. Furthermore, we will determine the role of the insect reticulon homolog during DENV infection of insect cells. The second set of proposed studies will characterize the role of FAM134B during DENV infection. FAM134B is a recently identified ER-shaping protein that regulates autophagy-mediated degradation of the ER (ER-phagy). Results from these studies will provide details on novel host-pathogen interactions that mediate replication of two RNA viruses, which represent a significant public health burden. Information derived from our results will provide significant insight into the biology of these viruses that can be utilized in the development of antivirals to combat disease. Furthermore, our results will also help to define the specific role of ER-shaping proteins in regulating the shape and dynamics of the ER.