Identification of host factors required by the tick-borne Powassan virus

项目摘要 黄病毒在世界范围内引起显著的发病率和死亡率。病毒的突然出现和传播 例如寨卡病毒，说明了我们的脆弱性，并强调需要做好准备， 病毒除蚊媒黄病毒外，蜱媒黄病毒的成员在全球范围内引起越来越多的感染。 我们缺乏有效的抗病毒药物。 Powassan病毒（POWV）是一种通过感染硬蜱叮咬传播的新兴病毒。的 人类中的POWV血清阳性率大多是未知的，因为感染通常可能没有症状。然而，在这方面， 有症状的POWV感染可发展为严重的、有时是致命的脑病， 约10%的发病率，存活的患者通常患有使人衰弱的长期神经症状。严重 POWV和其他蜱传黄病毒感染的临床后果以及缺乏特异性 治疗强调迫切需要更好地了解这些病毒在其蜱和哺乳动物宿主中的作用 这样就可以开发出预防性和治疗性的治疗方法。 该项目的目标是确定POWV所需的主机因素，这些因素也可能影响广泛的 现有的和新出现的黄病毒。这些宿主蛋白可能是抗病毒治疗的合适靶点。 干预，我们假设，受损的病毒复制，由于他们的抑制可能会允许先天性和 适应性免疫来控制感染此外，我们建议比较一组黄病毒，包括 POWV，以及它们对脊椎动物和无脊椎动物细胞中鉴定的宿主因子的需求。我们认为 这些方法将使我们能够为未来的药物开发研究优先考虑候选基因。 拟议的研究将利用CRISPR/Cas9筛选技术敲除基因组中的每一个基因。 人类基因组，目的是确定促进POWV感染的宿主因素。我们将使用类似的 以靶向阵列形式评价各种人和蜱细胞系中已鉴定宿主因子的方法 感染不同的黄病毒。 我们希望这项工作将扩大我们对蜱媒和蚊媒黄病毒的理解， 特别是在临床表型（脑炎与出血性疾病）和对宿主的共同需求方面， 因素此外，这将有助于确定哪些基因控制宿主物种和组织嗜性。在一起，这项工作 将为黄病毒生物学的许多方面提供丰富的新见解。

PROJECT SUMMARY Flaviviruses cause significant morbidity and mortality worldwide. The sudden emergence and spread of viruses such as Zika illustrate our vulnerability and emphasize the need for preparedness against related emerging viruses. In addition to mosquito-borne flaviviruses, members of tick-borne flaviviruses pose an increasing global threat for which we lack effective antivirals. Powassan virus (POWV) is an emerging virus transmitted by the bite of infected Ixodes ticks. The seroprevalence of POWV in humans is mostly unknown as infections may often be asymptomatic. However, symptomatic POWV infections may progress in to severe and sometimes fatal encephalic disease with mortality rates of ~10% and surviving patients often suffer from debilitating long-term neurologic symptoms. The severe clinical consequences of infections with POWV and other tick-borne flaviviruses and the lack of specific treatments highlight an urgent need for a better understanding of these viruses in their tick and mammalian hosts so that preventative and therapeutic treatments can be developed. The goal of this project is to identify host factors required by POWV that may also impact a wide range of existing and emerging flaviviruses. These host proteins may represent suitable targets for antiviral interventions, and we hypothesize that impaired virus replication due to their inhibition may allow innate and adaptive immunity to control the infection. Further, we propose to compare a panel of flaviviruses, including POWV, and their requirements for the identified host factors in both vertebrate and invertebrate cells. We believe these approaches will allow us to prioritize candidate genes for future drug development studies. The proposed studies will utilize CRISPR/Cas9 screening technology to knock out every gene in the human genome with the goal of identifying host factors that facilitate POWV infection. We will use a similar approach in a targeted, arrayed format to evaluate the identified host factors in various human and tick cell lines during infection with diverse flaviviruses. We expect that this work will broaden our understanding of tick- and mosquito-borne flaviviruses, specifically in terms of clinical phenotype (encephalitic vs hemorrhagic disease) and the shared need for host factors. In addition, it will help to define which genes govern host species and tissue tropism. Together, this work will provide a wealth of new insights into many aspects of flavivirus biology.