A method for the generation of pseudotyped virus particles bearing SARS coronavirus spike protein in high yields
A method for the generation of pseudotyped virus particles bearing SARS coronavirus spike protein in high yields
复制标题
一种高产率产生带有SARS冠状病毒刺突蛋白的假型病毒颗粒的方法
DOI：
10.1101/2021.07.30.454063
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Ohba Yusuke
中科院分区：
文献类型：
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作者：
Fujioka Yoichiro;Kashiwagi Sayaka;Yoshida Aiko;Satoh Aya O.;Fujioka Mari;Amano Maho;Yamauchi Yohei;Ohba Yusuke
持续的严重急性呼吸系统综合征冠状病毒2（SARS-CoV-2）大流行威胁着人类健康和全球经济。迫切需要开发更多的疫苗和疗法，但这种活病毒的开发必须在生物安全3级限制下进行。假型病毒已被广泛用于病毒进入和药物开发的研究，以克服这一限制。在这里，我们描述了一种改进的方案，以产生水泡性口炎病毒（VSV）假型与SARS-CoV或SARS-CoV-2刺突蛋白在高产量。我们发现用常规瞬时表达系统产生的假病毒体在其表面缺乏冠状病毒刺突蛋白，这是由于通过过表达该蛋白来抑制亲本VSV感染的结果。具有冠状病毒刺突蛋白的最佳表达水平的稳定细胞系的建立允许用刺突蛋白修饰的子代假病毒的有效生产。这种改进的VSV假病毒生产方法将有助于冠状病毒进入的研究和抗病毒剂的开发。
The ongoing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has threatened human health and the global economy. Development of additional vaccines and therapeutics is urgently required, but such development with live virus must be conducted with biosafety level 3 confinement. Pseudotyped viruses have been widely adopted for studies of virus entry and pharmaceutical development to overcome this restriction. Here we describe a modified protocol to generate vesicular stomatitis virus (VSV) pseudotyped with SARS-CoV or SARS-CoV-2 Spike protein in high yield. We found that pseudovirions produced with the conventional transient expression system lacked coronavirus Spike protein at their surface as a result of inhibition of parental VSV infection by overexpression of this protein. Establishment of stable cell lines with an optimal expression level of coronavirus Spike protein allowed the efficient production of progeny pseudoviruses decorated with Spike protein. This improved VSV pseudovirus production method should facilitate studies of coronavirus entry and development of antiviral agents.