Identification of Host Directed Drug Targets for SARS-CoV-2 Using Transposon Mutagenesis

摘要 正在进行的SARS-CoV-2大流行使人们迫切需要有效的新疗法 对抗广谱冠状病毒。识别参与这两种病毒复制的重要宿主基因 抗病毒防御可能揭示关键的宿主定向病毒疗法，这些疗法有可能对 SARS-CoV-2和未来的冠状病毒爆发。我们实验室的首要目标是确定 针对病毒感染的宿主防御机制。此应用程序旨在识别主机编码的 抵抗SARS-CoV-2感染的机制以及对病毒复制至关重要的细胞因素。 根据新出现的数据和我们的初步研究，我们的中心假设是宿主的上调 阻止病毒进入、抑制宿主细胞中的病毒复制或提高细胞能力的“限制因素” 抵御病毒诱导的细胞病变是宿主防御的重要策略。我们的理论基础是 拟议的工作是，确定这些因素将为治疗干预提供新的目标，并且 与病毒编码的靶标相比，这些靶标可能不太容易产生耐药性。我们将使用一种新的正向遗传 在BSL3中筛选抵抗SARS-CoV-2感染的宿主基因的方法 病毒。这将使我们能够在病毒感染的所有阶段确定关键目标。然后我们将验证新主机 并利用这些洞察力合理地制定抗病毒治疗策略。我们的筛选方法是 创新是因为它既可以识别与抗性有关的寄主基因，也可以识别寄主基因 感染所需，通常在传统的基于RNAi的筛查中检测到。这部作品具有 有可能发现新的靶点，包括非编码的RNA元件，而使用现有的 接近了。

SUMMARY The ongoing SARS-CoV-2 pandemic has brought into stark relief the need for novel therapeutics that can work against a broad spectrum of coronaviruses. Identifying essential host genes involved in both virus replication and antiviral defense could reveal key host-directed viral therapies that have the potential to work against SARS-CoV-2 and future coronavirus outbreaks. The overarching goal of our laboratory is to identify mechanisms of host defense against viral infections. This application aims to identify both host-encoded mechanisms of resistance to infection by SARS-CoV-2 as well as cellular factors critical for virus replication. Based on emerging data and our preliminary studies, our central hypothesis is that the up-regulation of host ‘restriction factors’ that prevent viral entry, constrain virus replication in host cells, or increase the ability of cells to withstand viral-induced cytopathy represent an important strategy in host defense. Our rationale for the proposed work is that identification of such factors will provide new targets for therapeutic intervention, and that these may be less susceptible to resistance than viral-encoded targets. We will use a novel forward-genetic approach to screen for host genes that confer resistance to SARS-CoV-2 infection in BSL3 using the native virus. This will allow us to identify key targets at all stages of virus infection. We will then validate new host targets and use these insights to rationally develop antiviral therapeutic strategies. Our screening approach is innovative because it allows identification of both host genes that confer resistance as well as host genes required for infection, which are normally detected in conventional RNAi-based screens. This work has the potential to identify new targets, including non-coding RNA elements, which would be missed using existing approaches.