【前沿进展】我国学者揭示乙脑病毒组装机制

    近日,中国农业科学院哈尔滨兽医研究所重要人兽共患病与烈性外来病团队在乙型脑炎病毒复制周期机制研究方面取得新进展。相关研究成果在线发表在病毒学国际期刊《Journal of Virology》上,论文题目为Host factor SPCS1 regulates the replication ofJapanese encephalitis virus 3 through interactions with transmembrane domainsof NS2B。研究首次阐明了宿主因子SPCS1参与乙型脑炎病毒组装的机制,进一步完善了乙型脑炎等黄病毒复制生命周期的机制细节,加深了宿主因子与病毒相互作用的理解,为研制新的抗病毒药物提供了新思路与方向。

 

 

  BiFC与Co-IP法分析SPCS1与NS2B蛋白相互作用

  背 景

  乙型脑炎病毒与登革病毒、西尼罗病毒、寨卡病毒等同属黄病毒科黄病毒属,由蚊媒传播,对动物和人类健康构成持续威胁,不断引发新的疫情和公共卫生危机。因次,深入研究该类病毒的致病机制,揭示病毒复制生命周期中各环节的调控机制,将为研制新型抗病毒药物与治疗策略提供理论依据。

  结果速览

  研究团队以乙型脑炎病毒为模型,利用RNA干扰技术与基因敲除细胞系技术验证了宿主因子SPCS1(Signal peptidase complex subunit 1)参与了病毒的复制调控(图1)。发现SPCS1分子除影响病毒多聚蛋白的剪切加工外,还影响病毒粒子的组装(图2)。进一步研究表明SPCS1与乙脑病毒NS2B蛋白发生相互作用,相互作用发生在NS2B蛋白的两个跨膜结构域与包含跨膜结构域的SPCS1分子的C端片段(图3)。

  

 

  Fig 1. Effects of JEV propagation in SPCS1 KO HEK-293 cells following 649 complementation with a plasmid expressing SPCS1

  

 

  Fig 2. Effects of loss of SPCS1 function on viral entry into cells, genome RNA 692 replication, and protein processing during JEV infection.

  

 

  Fig 3. Interaction of SPCS1 with the JEV NS2B protein in mammalian cells.

  该项研究进一步完善了乙型脑炎等黄病毒复制生命周期的机制细节,加深了宿主因子与病毒相互作用的理解,为研制新的抗病毒药物提供了新思路与方向。该研究得到国家重点研发计划项目(2016YFD0500403)的资助,该团队黄病毒研究组组长华荣虹副研究员和团队首席步志高研究员为论文通讯作者,硕士研究生马乐为第一作者。

  ABSTRACT

  Signal peptidase complex subunit 1 (SPCS1) is a newly identified hostfactor that regulates flavivirus replication, but the molecular mechanism isnot fully understood. Herein, using Japanese encephalitis virus (JEV) as amodel, we investigated the mechanism through which host factor SPCS1 regulatesthe replication of flaviviruses. We first validated the regulatory function ofSPCS1 in JEV propagation by knocking down and knocking out endogenous SPCS1.Loss of SPCS1 function markedly reduced intracellular virion assembly andproduction of infectious JEV particles, but did not affect virus cell entry,RNA replication, or translation. SPCS1 was found to interact with NS2B, whichis involved in post-translational protein processing and viral assembly. Serialdeletion mutation of the JEV NS2B protein revealed that two transmembranedomains, NS2B (1-49) and NS2B (84-131), interact with SPCS1. Furthermutagenesis analysis of conserved flavivirus residues in two SPCS1 interactiondomains of NS2B demonstrated that G12A, G37A, and G47A in NS2B (1-49), andP112A in NS2B (84-131), weakened the interaction with SPCS1. Deletion mutationof SPCS1 revealed that SPCS1 (91-169) which containing two transmembranedomains was involved in the interaction with both NS2B (1-49) and NS2B(84-131). Taken together, the results demonstrate that SPCS1 affects viralreplication by interacting with NS2B, thereby influencing post-translationalprocessing of JEV proteins and the assembly of virions.

  IMPORTANCE

  Understanding viral-host interactions is important for elucidating themolecular mechanisms of viral propagation, and identifying potential anti-viraltargets. Previous reports demonstrated that SPCS1 is involved in the flaviviruslife cycle, but the mechanism remains unknown. In this study, we confirmed thatSPCS1 participates in the post-translational protein processing and viralassembly stages of the JEV lifecycle, but not in the cell entry, genome RNAreplication, or translation stages. Furthermore, we found that SPCS1 interactswith two independent transmembrane domains of the Flavivirus NS2B protein. NS2Balso interacts with NS2A, which is proposed to mediate viral assembly.Therefore, we propose a protein-protein interaction model showing how SPCS1participates in the assembly of JEV particles. The findings expand ourunderstanding of how host factors participate in the flavivirus replicationlifecycle, and identify potential anti-viral targets for combatting flavivirusinfection.

  文章来源:

  1.Le Ma, Fang Li, Jing-Wei Zhang, Wei Li, Dong-Ming Zhao, Han Wang, Rong-HongHua and Zhi-Gao Bu. Host factor SPCS1 regulates the replication of Japaneseencephalitis virus through interactions with transmembrane domains of NS2B.JVI.2018.00197-18.

  Doi:10.1128/JVI.00197-18

  2.中国农科院哈尔滨兽医研究所:哈兽研重要人兽共患病与烈性外来病团队在乙型脑炎病毒复制机制研究方面取得新进展。

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