Pseudotyped vesicular stomatitis virus for analysis of virus entry mediated by SARS coronavirus spike proteins.
Pseudotyped vesicular stomatitis virus for analysis of virus entry mediated by SARS coronavirus spike proteins.
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DOI：
10.1007/978-1-59745-181-9_23
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发表时间：
2008
期刊：
影响因子：
--
通讯作者：
Taguchi F
中科院分区：
文献类型：
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作者：
Fukushi S;Watanabe R;Taguchi F
严重急性呼吸综合征 (SARS) 冠状病毒 (CoV) 含有刺突 (S) 蛋白，该蛋白与受体分子（血管紧张素转换酶 2；ACE2）结合，诱导膜融合，并充当中和表位。为了研究 S 蛋白的功能，我们在此描述了使用 VSVΔG*/GFP 系统生成带有 SARS-CoV S 蛋白的水泡性口炎病毒 (VSV) 假型，其中 G 基因被绿色荧光蛋白 (GFP) 基因 (VSV-SARS-CoV-St19/GFP) 取代。 SARS-CoV S 蛋白（SARS-CoV-St19）胞质结构域的部分缺失可以有效地掺入 VSV 颗粒中，从而生成高滴度的假型病毒。抗 SARS-CoV 抗体中和试验表明，VSV-SARS-St19/GFP 假型感染是由 SARS-CoV S 蛋白介导的。 VSVΔG*/SEAP 系统分泌碱性磷酸酶而不是 GFP，也被生成为具有 SARS-CoV S 蛋白的 VSV 假型 (VSV-SARS-CoV-St19/SEAP)。该系统通过测量碱性磷酸酶活性，能够对 SARS-CoV S 蛋白介导的细胞进入进行高通量分析。因此，用 SARS-CoV S 蛋白假型化的 VSV 可用于开发快速检测系统，用于中和 SARS-CoV 感染特异性抗体，以及研究 S 介导的 SARS-CoV 细胞进入。
Severe acute respiratory syndrome (SARS) coronavirus (CoV) contains a spike (S) protein that binds to a receptor molecule (angiotensin-converting enzyme 2; ACE2), induces membrane fusion, and serves as a neutralizing epitope. To study the functions of the S protein, we describe here the generation of SARS-CoV S protein-bearing vesicular stomatitis virus (VSV) pseudotype using a VSV∆G∗/GFP system in which the G gene is replaced by the green fluorescent protein (GFP) gene (VSV-SARS-CoV-St19/GFP). Partial deletion of the cytoplasmic domain of SARS-CoV S protein (SARS-CoV-St19) allowed efficient incorporation into the VSV particle that enabled the generation of a high titer of pseudotype virus. Neutralization assay with anti-SARS-CoV antibody revealed that VSV-SARS-St19/GFP pseudotype infection is mediated by SARS-CoV S protein. The VSV∆G∗/SEAP system, which secretes alkaline phosphatase instead of GFP, was also generated as a VSV pseudotype having SARS-CoV S protein (VSV-SARS-CoV-St19/SEAP). This system enabled high-throughput analysis of SARS-CoV S protein-mediated cell entry by measuring alkaline phosphatase activity. Thus, VSV pseudotyped with SARS-CoV S protein is useful for developing a rapid detection system for neutralizing antibody specific for SARS-CoV infection as well as studying the S-mediated cell entry of SARS-CoV.