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  • 邓力宾,郭巾旭,马鹏娟,王刚,龙钢.多配体聚糖结合蛋白1过表达增加低密度丙型肝炎病毒颗粒的产生[J].第二军医大学学报,2018,39(3):263-268    [点击复制]
  • DENG Li-bin,GUO Jin-xu,MA Peng-juan,WANG Gang,LONG Gang.Overexpression of syntenin-1 enhances production of low-density hepatitis C virus particles[J].Acad J Sec Mil Med Univ,2018,39(3):263-268   [点击复制]
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多配体聚糖结合蛋白1过表达增加低密度丙型肝炎病毒颗粒的产生
邓力宾1,2,郭巾旭2,马鹏娟2,王刚1,龙钢2*
0
(1. 上海大学生命科学学院, 上海 201900;
2. 中国科学院上海巴斯德研究所, 上海 200031
*通信作者)
摘要:
目的 探讨多配体聚糖结合蛋白1(syntenin-1)对丙型肝炎病毒(HCV)颗粒组装的影响。方法 采用免疫印迹法分析人原代肝细胞中syntenin-1的含量。使用慢病毒质粒转染人肝癌细胞系Huh7.5.1构建syntenin-1过表达细胞系和绿色荧光蛋白(GFP)过表达细胞系。采用电转染法将HCV RNA转入Huh7.5.1细胞、syntenin-1过表达细胞和GFP过表达细胞后,用荧光素酶分析、免疫印迹分析和病毒滴度测定方法分别从RNA复制、病毒蛋白表达和感染性病毒颗粒的分泌方面研究syntenin-1过表达对HCV的影响。用密度梯度分析法检测HCV RNA和感染滴度在不同密度组分的分布情况。结果 3例不同人原代肝细胞中syntenin-1的含量均较Huh7.5.1细胞高(P<0.01)。Syntenin-1过表达不影响HCV RNA在宿主细胞中的复制;syntenin-1过表达细胞来源HCV病毒的感染性与Huh7.5.1细胞或GFP过表达细胞相比差异均无统计学意义。在syntenin-1过表达细胞培养上清液中,HCV感染性颗粒从主要集中在1.08~1.16 g/mL区域逐渐转变为有部分转移到密度为1.01~1.02 g/mL的低密度区域。结论 Syntenin-1过表达改变了HCV感染性颗粒的密度组分分布。
关键词:  丙型肝炎病毒  多配体蛋白聚糖结合蛋白  密度组分分布  病毒颗粒
DOI:10.16781/j.0258-879x.2018.03.0263
投稿时间:2017-10-24修订日期:2017-12-25
基金项目:国家重点基础研究发展计划(2015CB554301).
Overexpression of syntenin-1 enhances production of low-density hepatitis C virus particles
DENG Li-bin1,2,GUO Jin-xu2,MA Peng-juan2,WANG Gang1,LONG Gang2*
(1. College of Life Science, Shanghai University, Shanghai 201900, China;
2. Institute Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200031, China
*Corresponding author)
Abstract:
Objective To investigate the effect of syntenin-1 on the assembly of hepatitis C virus (HCV) particles. Methods The content of syntenin-1 in human primary hepatocytes was detected by Western blotting analysis. Human hepatocellular carcinoma cell line Huh7.5.1 was transfected with lentiviral plasmids to construct syntenin-1 overexpressed cell line and green fluorescent protein (GFP) overexpressed cell line. After transferring HCV RNA into Huh7.5.1 cells, syntenin-1 overexpressed cells and GFP overexpressed cells by electroporation, the effects of syntenin-1 overexpression on HCV in the cells were investigated in terms of RNA replication, viral protein contents and secretion of infectious virus particles by luciferase analysis, Western blotting and virus titer, respectively. The density of viral particles was assessed by isopycnic ultracentrifugation to analyze the distributions of HCV RNA and infective titer. Results The contents of syntenin-1 in three human primary hepatocytes were higher than that in the Huh7.5.1 cells (P<0.01). Syntenin-1 overexpression had no effect on HCV RNA replication in host cells; the infectivity of HCV derived from syntenin-1 overexpressed cells was not significantly different compared with the Huh7.5.1 cells or GFP overexpressed cells. In syntenin-1 overexpressed cell culture supernatants, some infectious HCV particles mainly concentrated in the region of concentration of 1.08-1.16 g/mL gradually transferred to the low-density region of 1.01-1.02 g/mL. Conclusion Syntenin-1 overexpression alters the distribution of density components of infectious HCV particles.
Key words:  hepatitis C virus  syntenin  density component distribution  virion