烟草花叶病毒双位点弱毒突变体的研发

doi:10.1016/j.jia.2024.02.019

Journal of Integrative Agriculture ›› 2024, Vol. 23 ›› Issue (7): 2318-2331.DOI: 10.1016/j.jia.2024.02.019

烟草花叶病毒双位点弱毒突变体的研发

收稿日期:2023-11-16 接受日期:2024-01-17 出版日期:2024-07-20 发布日期:2024-07-01

Development of a stable attenuated double-mutant of tobacco mosaic virus for cross-protection

Xiaojie Xu^{1, 2*#}, Shaoyan Jiang^2*, Chunju Liu^3*, Xujie Sun², Qing Zhu², Xiuzhai Chen⁴, Pengchao Jiang³, Fenglong Wang⁵, Yanping Tian^2#, Xiangdong Li^2#

¹ The Engineering Research Institute of Agriculture and Forestry, Ludong University, Yantai 264025, China
² Laboratory of Plant Virology, Department of Plant Pathology, College of Plant Protection, Shandong Agricultural University, Tai’an 271018, China
³ Weifang Tobacco Corporation, Weifang 261031, China
⁴ Linyi Tobacco Corporation, Linyi 276000, China
⁵ Tobacco Research Institute, Chinese Academy of Agricultural Sciences, Qingdao 266101, China

Received:2023-11-16 Accepted:2024-01-17 Online:2024-07-20 Published:2024-07-01
About author:Shaoyan Jiang, E-mail: Syan_Jiang@163.com; Chunju Liu, E-mail: liuchunju19790326@163.com; #Correspondence Xiaojie Xu, E-mail: xiaojiexua105@163.com; Yanping Tian, E-mail: yanping.tian@sdau.edu.cn; Xiangdong Li, Tel: +86-538-8242523, Fax: +86-538-8226399, E-mail: xdongli@sdau.edu.cn * These authors contributed equally to this study.
Supported by:
This work was supported by funds from ‘Taishan Scholar’ Construction Project, China (TS2022-028 and 202101-KN275).

摘要/Abstract

摘要：

烟草（Nicotiana tabacum）和番茄（Solanum lycopersicum）是我国两种主要的经济作物，其安全生产受到烟草花叶病毒（tobacco mosaic virus，TMV）的严重威胁。交叉保护是预防植物病毒病安全且环保的措施。然而，稳定的TMV弱毒突变体却很少。TMV基因组为单链RNA，其编码的p126蛋白参与病毒复制、细胞间运动、抑制寄主RNA沉默和病毒症状形成等过程。在本研究中我们发现，将p126蛋白中第196位的精氨酸（arginine，R¹⁹⁶）替换为天冬氨酸（aspartic acid，D），第614位的谷氨酸（glutamic acid，E⁶¹⁴）替换为甘氨酸（glycine，G），第643位的丝氨酸（serine，S⁶⁴³）替换为苯丙氨酸（phenylalanine，F），或者把第730位的D（D⁷³⁰）替换为S，均可以显著降低TMV的致病力和复制水平。然而，只有S643F突变可以降低TMV p126的RNA沉默抑制活性。由于RNA病毒的RNA聚合酶缺乏校对和修复活性，导致RNA病毒基因组易发生突变。一些携带单一致弱突变位点的病毒突变体可能会通过自发突变恢复其致病力，而在多个致弱位点有突变的突变体恢复强致病力的风险会相对较低。为了获得稳定的TMV弱毒突变体，我们在p126中两个位点同时引入突变，发现仅双位点突变体TMV-E614G-S643F可以系统侵染烟草植株，而且不引起明显的症状，因此将其作为用于交叉保护的候选突变体。我们将其在烟草植株中连续继代接种六次，发现双位点突变体TMV-E614G-S643F可以稳定遗传。交叉保护效果实验结果表明，双位点突变体TMV-E614G-S643F可以有效保护烟草和番茄植株免受野生型TMV侵染。本研究报道了一种具有应用前景的TMV双位点弱毒突变体，可用于通过交叉保护来防治烟草和番茄植株中的TMV。

Abstract:

Tobacco (Nicotiana tabacum) and tomato (Solanum lycopersicum) are two major economic crops in China. Tobacco mosaic virus (TMV; genus Tobamovirus) is the most prevalent virus infecting both crops. Currently, some widely cultivated tobacco and tomato cultivars are susceptible to TMV and there is no effective strategy to control this virus. Cross-protection can be a safe and environmentally friendly strategy to prevent viral diseases. However, stable attenuated TMV mutants are scarce. In this study, we found that the substitutions in the replicase p126, arginine at position 196 (R¹⁹⁶) with aspartic acid (D), glutamic acid at position 614 (E⁶¹⁴) with glycine (G), serine at position 643 (S⁶⁴³) with phenylalanine (F), or D at position 730 (D⁷³⁰) with S, significantly reduced the virulence and replication of TMV. However, only the mutation of S⁶⁴³ to F reduced the RNA silencing suppression activity of TMV p126. A double-mutant TMV-E614G-S643F induced no visible symptom and was genetically stable through six successive passages in tobacco plants. Furthermore, our results showed that TMV-E614G-S643F double-mutant could provide effective protection against the wild-type TMV infection in tobacco and tomato plants. This study reports a promising mild mutant for cross-protection to control TMV in tobacco and tomato plants.

Key words: cross-protection , double-mutant , p126 , tobacco mosaic virus , virulence

Xiaojie Xu, Shaoyan Jiang, Chunju Liu, Xujie Sun, Qing Zhu, Xiuzhai Chen, Pengchao Jiang, Fenglong Wang, Yanping Tian, Xiangdong Li. 烟草花叶病毒双位点弱毒突变体的研发[J]. Journal of Integrative Agriculture, 2024, 23(7): 2318-2331.

Xiaojie Xu, Shaoyan Jiang, Chunju Liu, Xujie Sun, Qing Zhu, Xiuzhai Chen, Pengchao Jiang, Fenglong Wang, Yanping Tian, Xiangdong Li. Development of a stable attenuated double-mutant of tobacco mosaic virus for cross-protection[J]. Journal of Integrative Agriculture, 2024, 23(7): 2318-2331.

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烟草花叶病毒双位点弱毒突变体的研发

Development of a stable attenuated double-mutant of tobacco mosaic virus for cross-protection

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