用于证明沙粒病毒核蛋白（NPs）在病毒RNA合成及抑制I型干扰素过程中作用的检测方法
Assays to Demonstrate the Roles of Arenaviral Nucleoproteins (NPs) in Viral RNA Synthesis and in Suppressing Type I Interferon.
作者信息
Huang Qinfeng, Shao Junjie, Liang Yuying, Ly Hinh
机构信息
Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai, China.
Department of Veterinary and Biomedical Sciences, University of Minnesota, Twin Cities, MN, USA.
出版信息
Methods Mol Biol. 2018;1604:189-200. doi: 10.1007/978-1-4939-6981-4_13.
Arenaviruses, such as Lassa virus (LASV) and Pichindé virus (PICV), are enveloped viruses with a bi-segmented ambisense RNA genome. The large (L) genomic segment encodes the Z matrix protein and the L RNA-dependent RNA polymerase, whereas the small (S) genomic segment encodes the nucleoprotein (NP) and the glycoprotein (GPC). The NP encapsidates viral genome, is required for viral transcription and replication, and acts as a type I interferon (IFN) antagonist. This article describes assays to demonstrate that NP contains 3'-5' exoribonuclease (RNase) activity to degrade modeled RNA of the pathogen-associated molecular pattern type and suppresses the IFNβ promoter-driven luciferase reporter gene. The minigenomic (MG) assay is used to assess the role of NP in replicating and transcribing a viral promoter-driven luciferase reporter gene. These powerful assays demonstrate the versatility of NP in mediating viral replication as well as in modulating host innate immune responses.
沙粒病毒，如拉沙病毒（LASV）和皮钦德病毒（PICV），是具有双节段负链RNA基因组的包膜病毒。大（L）基因组节段编码Z基质蛋白和L RNA依赖性RNA聚合酶，而小（S）基因组节段编码核蛋白（NP）和糖蛋白（GPC）。NP包裹病毒基因组，是病毒转录和复制所必需的，并作为I型干扰素（IFN）拮抗剂发挥作用。本文描述了一些实验，以证明NP具有3'-5'外切核糖核酸酶（RNase）活性，可降解病原体相关分子模式类型的模拟RNA，并抑制IFNβ启动子驱动的荧光素酶报告基因。微型基因组（MG）检测用于评估NP在复制和转录病毒启动子驱动的荧光素酶报告基因中的作用。这些强大实验证明了NP在介导病毒复制以及调节宿主先天免疫反应方面的多功能性。
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