Generation and Application of a Luciferase Reporter Virus Based on Yellow Fever Virus 17D

Dan Li; Jing-Long Ye; Zhong-Yu Liu

doi:10.1007/s12250-021-00428-1

December 2021

Dan Li, Jing-Long Ye and Zhong-Yu Liu. Generation and Application of a Luciferase Reporter Virus Based on Yellow Fever Virus 17D[J]. Virologica Sinica, 2021, 36(6): 1456-1464. doi: 10.1007/s12250-021-00428-1

Citation: Dan Li, Jing-Long Ye, Zhong-Yu Liu. Generation and Application of a Luciferase Reporter Virus Based on Yellow Fever Virus 17D .VIROLOGICA SINICA, 2021, 36(6) : 1456-1464. http://dx.doi.org/10.1007/s12250-021-00428-1

基于黄热病毒疫苗株17D的萤光素酶报告病毒的构建及应用

李丹 ¹ ,
叶经龙 ¹ ,
刘忠钰 ^1,,

中山大学医学院，感染免疫中心

通讯作者： 刘忠钰, Liuzhy79@mail.sysu.edu.cn, ORCID: 0000-0001-7385-5681
收稿日期： 2021-05-13
录用日期： 2021-06-08
出版日期： 2021-08-03

摘要

黄热病毒（YFV）是一种再发病毒，人类感染后可导致高致死率的黄热病。尽管已有高效疫苗，但目前对YFV的复制机制知之甚少，且仍缺乏临床可用的特效药物。本研究通过反向遗传学手段，将海肾萤光素酶报告基因（Rluc）引入YFV疫苗株17D中，从而获得重组病毒17D-Rluc.2A。该报告病毒具有与亲本株相似的蚀斑形态和体外增殖特征，在感染17D-Rluc.2A的哺乳动物细胞和蚊虫细胞中均可检测到报告基因的高效表达，而且细胞内荧光素酶的表达与细胞外病毒粒子的载量间有良好的线性关系。在此基础上，我们将基于17D-Rluc.2A报告病毒的突变分析与选择性2'-羟基酰化引物延伸（SHAPE）技术相结合，确认了保守的5'-SLA元件在YFV复制过程中的关键作用。因此，17D-Rluc.2A在YFV复制研究中具有重要价值。最后，我们证实两种具有不同抗病毒机制的化合物可以有效抑制17D-Rluc.2A在不同细胞系中的增殖，表明该报告病毒可用于抗病毒药物的评价。综上，17D-Rluc.2A将为YFV复制研究及药物研发奠定有力的技术基础。
- 黄热病毒
- , 17D
- , 报告病毒
- , 顺式作用RNA元件
- , 抗病毒药物

Generation and Application of a Luciferase Reporter Virus Based on Yellow Fever Virus 17D

The Centre for Infection and Immunity Studies, School of Medicine, Sun Yat-Sen University, Guangzhou 510006, China

Corresponding author: Zhong-Yu Liu, Liuzhy79@mail.sysu.edu.cn
ORCID: 0000-0001-7385-5681
Received Date: 13 May 2021
Accepted Date: 08 June 2021
Published Date: 03 August 2021

Abstract

Yellow fever virus (YFV) is a re-emerging virus that can cause life-threatening yellow fever disease in humans. Despite the availability of an effective vaccine, little is known about the replication mechanism of YFV, and there are still no available specific anti-YFV medicines. Herein, by introducing the Renilla luciferase gene (Rluc) into an infectious clone of YFV vaccine strain 17D, we generated a recombinant virus 17D-Rluc.2A via reverse genetics approaches. The 17D-Rluc.2A had similar plaque morphology and comparable in vitro growth characteristics with its parental strain. Importantly, the reporter luciferase was efficiently expressed in 17D-Rluc.2A-infected mammalian and mosquito cells, and there was a good linear correlation between intracellular luciferase expression and extracellular infectious virion reproduction. Furthermore, by a combination of the 17D-Rluc.2A reporter virus and selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) technology, the conserved 5′-SLA element was shown to be essential for YFV replication, highlighting the capability of 17D-Rluc.2A in the investigation of YFV replication. At last, we demonstrated that two compounds with distinct anti-viral mechanisms can effectively inhibit the viral propagation in 17D-Rluc.2A-infected cells, demonstrating its potential application in the evaluation of anti-viral medicines. Taken together, the 17D-Rluc.2A serves as a useful tool for the study of YFV replication and anti-YFV medicine development.
- Yellow fever virus (YFV)
- , 17D
- , Reporter virus
- , Cis-acting RNA elements
- , Anti-viral medicines

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