Shao Junjie, Liu Xiaoying, Liang Yuying, Ly Hinh
Department of Veterinary and Biomedical Sciences, University of Minnesota, Twin Cities, 1988 Fitch Ave., 295 AS/VM Bldg., Saint Paul, MN, 55108, USA.
Methods Mol Biol. 2018;1604:169-178. doi: 10.1007/978-1-4939-6981-4_11.
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 precursor complex (GPC). GPC is processed by signal peptidase in the endoplasmic reticulum into the stable signal peptide (SSP) and GP1/GP2, which is further cleaved by the Golgi-resident subtilisin kexin isozyme-1 (SKI-1)/site-1 protease (S1P) into the cellular receptor-recognition subunit GP1 and the transmembrane subunit GP2, which helps promote the membrane fusion reaction to allow virus entry into the cell. This article describes assays to assess PICV GPC expression, proteolytic processing, fusion function, and GPC-mediated virus-like particle (VLP) entry into cells under tissue-culture conditions.
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）和糖蛋白前体复合物（GPC）。GPC在内质网中被信号肽酶加工成稳定信号肽（SSP）和GP1/GP2，后者再被高尔基体驻留枯草杆菌蛋白酶/kexin同工酶-1（SKI-1）/1位点蛋白酶（S1P）进一步切割成细胞受体识别亚基GP1和跨膜亚基GP2，这有助于促进膜融合反应，使病毒进入细胞。本文描述了在组织培养条件下评估PICV GPC表达、蛋白水解加工、融合功能以及GPC介导的病毒样颗粒（VLP）进入细胞的实验方法。
