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Rab1b-GBF1-ARFs mediated intracellular trafficking is required for classical swine fever virus replication in swine umbilical vein endothelial cells.
Veterinary Microbiology ( IF 3.3 ) Pub Date : 2020-06-01 , DOI: 10.1016/j.vetmic.2020.108743
Liang Zhang 1 , Tao Wang 1 , Mengzhao Song 1 , Mingxing Jin 1 , Shanchuan Liu 1 , Kangkang Guo 1 , Yanming Zhang 1
Affiliation  

Classical swine fever virus (CSFV), a plus-sense RNA virus, utilizes host intracellular membrane organelles for its replication. Our previous studies have shown that disruption of the intracellular membrane-trafficking events can inhibit CSFV replication. However, the underlying mechanism of this process in CSFV infection has not been elucidated. To determine the role of Golgi-associated anterograde and retrograde trafficking in CSFV replication, we revealed the effect of vesicular transport between Golgi and ER inhibitors Brefeldin A (BFA) and 2,2-methyl-N-(2,4,6,-trimethoxyphenyl) dodecanamide (CI-976), the GBF1 inhibitor golgicide A (GCA) on virus production. Our results showed that disruption of vesicular trafficking by BFA, CI-976, and GCA significantly inhibited CSFV infection. Subsequent experiments revealed that knockdown of Rab1b by lentiviruses and negative-mutant Rab1b-N121I transfection inhibited CSFV infection. Furthermore, we showed that the Rab1b downstream vesicular component effectors GBF1, and class I and class II ADP-ribosylation factors (ARFs) were also involved in virus replication. In addition, confocal microscopy assay showed that CSFV infection disrupted the Golgi apparatus resulting in extended Golgi distribution around the nucleus. We also showed that cell secretory pathway, measured using Gaussia luciferase flash assay, was blocked in CSFV infected cells. Taken together, these findings demonstrate that CSFV utilizes Rab1b-GBF1-ARFs mediated trafficking to promote its own replication. These findings also provide new insights into the intracellular trafficking pathways utilized for CSFV life cycle.



中文翻译:

Rab1b-GBF1-ARFs介导的细胞内运输是猪瘟病毒在猪脐静脉内皮细胞中复制的必需条件。

古典猪瘟病毒(CSFV)是一种正向RNA病毒,利用宿主细胞内膜细胞器进行复制。我们以前的研究表明,破坏细胞内膜运输事件可以抑制CSFV复制。但是,尚未阐明此过程在CSFV感染中的潜在机制。为了确定高尔基体相关顺行和逆行运输在CSFV复制中的作用,我们揭示了高尔基体和ER抑制剂布雷菲德菌素A(BFA)和2,2-甲基-N-(2,4,6,-三甲氧基苯基)十二烷酰胺(CI-976),GBF1抑制剂杀虫剂A(GCA)对病毒的产生。我们的结果表明,BFA,CI-976和GCA对水泡运输的破坏显着抑制了CSFV感染。随后的实验表明,慢病毒和负突变的Rab1b-N121I转染可抑制Rab1b抑制CSFV感染。此外,我们显示Rab1b下游水泡成分效应物GBF1,以及I类和II类ADP核糖基化因子(ARF)也参与病毒复制。此外,共聚焦显微镜分析表明,CSFV感染破坏了高尔基体,导致高尔基体在核周围的分布延长。我们还显示,使用高斯荧光素酶快速测定法测量的细胞分泌途径在CSFV感染的细胞中被阻断。综上所述,这些发现表明CSFV利用Rab1b-GBF1-ARF介导的贩运来促进其自身复制。这些发现也为用于CSFV生命周期的细胞内运输途径提供了新的见解。

更新日期:2020-06-01
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