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Porcine Viperin protein inhibits the replication of classical swine fever virus (CSFV) in vitro

机译:猪Viperin蛋白在体外抑制经典猪瘟病毒(CSFV)的复制

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Classical swine fever virus (CSFV) is the causative pathogen of Classical swine fever (CSF), a highly contagious disease of swine. Viperin is one of the hundreds of interferon-stimulated genes (ISGs), and possesses a wide range of antiviral activities. The aim of this study was to explore whether porcine Viperin has the anti-CSFV activity. The influences of CSFV infection on Viperin expression and Newcastle disease virus (NDV)/Pseudorabies virus (PRV)-induced Viperin expression were examined in 3D4/21 cells and porcine peripheral blood mononuclear cells (PBMCs). Porcine Viperin gene was amplified to generate cell line PK-Vi over-expressing Viperin. CSFV was inoculated in the cell lines and viral load was detected by qRT-PCR, virus titration and Western blot. The influence of Viperin expression on CSFV binding, entry and release in the cells was also examined. The co-localization of Viperin with CSFV and its proteins (E2, NS5B) was determined by confocal laser scanning microscopy test. Co-IP assay was performed to check the interaction of Viperin with CSFV proteins. CSFV infection could not induce Viperin expression in vitro while significantly inhibiting NDV/PRV-induced Viperin expression at 12, 24 and 48?h post infection (hpi; P?
机译:古典猪瘟病毒(CSFV)是古典猪瘟(CSF)的致病性病原体,是猪的一种高度传染性疾病。毒蛇蛋白是数百种干扰素刺激基因(ISG)之一,并具有广泛的抗病毒活性。这项研究的目的是探讨猪Viperin是否具有抗CSFV活性。在3D4 / 21细胞和猪外周血单核细胞(PBMC)中检查了CSFV感染对Viperin表达和新城疫病毒(NDV)/伪狂犬病病毒(PRV)诱导的Viperin表达的影响。猪Viperin基因被扩增以产生过表达Viperin的细胞系PK-Vi。将CSFV接种在细胞系中,并通过qRT-PCR,病毒滴定和Western印迹检测病毒载量。还检查了Viperin表达对CSFV结合,进入和释放在细胞中的影响。通过共聚焦激光扫描显微镜测试确定了Viperin与CSFV及其蛋白(E2,NS5B)的共定位。进行Co-IP分析以检查Viperin与CSFV蛋白的相互作用。 CSFV感染不能在体外诱导Viperin表达,而在感染后12、24和48h显着抑制NDV / PRV诱导的Viperin表达(hpi; P 0.05)。与对照组细胞(表达EGFP的PK-C1)相比,CSFV在PK-Vi的增殖在24、48和72hhpi受到显着抑制(P 0.05)。细胞培养上清液和细胞沉淀中的病毒均降低。 CSFV结合和进入细胞不受Viperin表达的干扰。这些结果表明其抗CSFV功能发生在基因组和/或蛋白质合成步骤中。共聚焦激光扫描显微镜测试显示,在感染了CSFV的PK-Vi细胞中,Viperin-EGFP蛋白与CSFV E2蛋白共定位。进一步的实验表明,Viperin蛋白与CSFV的E2和NS5B蛋白在转染的293ΔT细胞中共定位。此外,Co-IP分析证实了Viperin与E2蛋白(而非NS5B)的相互作用。猪Viperin可能通过Viperin与CSFV E2蛋白在细胞质中的相互作用而在体外有效抑制CSFV复制。该结果为进一步研究Viperin与CSFV和其他病毒的相互作用提供了基础。

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