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中华实验和临床感染病杂志(电子版) ›› 2017, Vol. 11 ›› Issue (05) : 441 -446. doi: 10.3877/cma.j.issn.1674-1358.2017.05.004

基础论著

HCV核心蛋白调控SREBP-1c表达水平的机制研究
李敏1, 王琦2, 刘顺爱2, 成军2,()   
  1. 1. 730000 兰州市,兰州大学第一医院感染科
    2. 100015 北京首都医科大学附属北京地坛医院传染病研究所
  • 收稿日期:2016-10-30 出版日期:2017-10-15
  • 通信作者: 成军
  • 基金资助:
    北京市医院管理局扬帆计划项目(肝炎专业)(No. ZYLX201402)

Molecular mechanism of SREBP-1c expression regulated by HCV core protein

Min Li1, Qi Wang2, Shun'ai Liu2, Jun Cheng2,()   

  1. 1. Department of Infection, The First Hospital of Lanzhou University, Lanzhou 730000, China
    2. Infectious Diseases Institute, Beijing Ditan Hospital, Capital Medical University, Beijing 100015, China
  • Received:2016-10-30 Published:2017-10-15
  • Corresponding author: Jun Cheng
引用本文:

李敏, 王琦, 刘顺爱, 成军. HCV核心蛋白调控SREBP-1c表达水平的机制研究[J]. 中华实验和临床感染病杂志(电子版), 2017, 11(05): 441-446.

Min Li, Qi Wang, Shun'ai Liu, Jun Cheng. Molecular mechanism of SREBP-1c expression regulated by HCV core protein[J]. Chinese Journal of Experimental and Clinical Infectious Diseases(Electronic Edition), 2017, 11(05): 441-446.

目的

探讨HCV核心蛋白调控SREBP-1c的分子生物学机制。

方法

构建HCV核心蛋白真核表达载体pcDNA3.1/myc-His(-)-Core(1b,3a),转染HepG2细胞系,48 h后提取细胞总RNA和总蛋白,采用real-time PCR、Western blot等方法检测脂代谢相关基因Sirt1和SREBP-1c的mRNA和蛋白表达水平;构建Sirt1启动子报告基因表达载体pGL4.10-Sirt1,与pcDNA3.1/myc-His(-)-Core(1b,3a)共转染HepG2细胞系24 h后检测核心蛋白对Sirt1启动子活性的影响;构建pmirGLO-SREBP-1c-3'-UTR报告基因表达载体,与pcDNA3.1/myc-His(-)-Core(3a)共转染HepG2细胞系48 h后检测SREBP-1c-3'-UTR相对luciferase活性。

结果

与对照组相比,1b型和3a型HCV核心蛋白表达组SREBP-1c的mRNA表达上调(分别上调1.358倍和1.337倍,P = 0.043、0.008)SREBP-1c蛋白表达水平上调(分别上调1.608倍和1.926倍,P = 0.042、0.008);与对照组相比,1b型和3a型HCV核心蛋白表达组Sirt1 mRNA表达上调(分别上调1.566倍和1.71倍,P = 0.037、0.006),Sirt1蛋白表达水平上调(分别上调1.436倍和1.588倍,P = 0.026、0.009);与对照组相比,HCV核心蛋白表达组Sirt1的启动子活性无显著变化;与对照组相比,HCV核心蛋白表达组SREBP-1c-3'-UTR相对luciferase活性下调(相对值为0.667,P = 0.008)。

结论

HCV核心蛋白上调SREBP-1c与Sirt1的表达,可能是HCV相关性肝脂肪变的发病机制之一。microRNA参与HCV核心蛋白对SREBP-1c的表达调控,而是否有microRNA对HCV调控Sirt1表达发挥作用尚待于进一步的研究证实。

Objective

To investigate the molecular mechanism of SREBP-1c expression regulated by HCV core protein.

Methods

HepG2 cells were transfected with pcDNA3.1/myc-His(-)-Core(1b, 3a), and the mRNA and protein expression levels of SREBP-1c and Sirt1 were determined by real time PCR and Western blot 48 h post-transfection. pGL4.10-Sirt1 and pcDNA3.1/myc-His(-)-Core(1b, 3a) were co-transfected into HepG2 cells, the Sirt1 promoter activity was measured by Luciferase assay 24h post-transfection. pmirGLO-SREBP-1c-3'-UTR and pcDNA3.1/myc-His(-)-Core (3a) were co-transfected into HepG2 cells, and the relative luciferase activity of SREBP-1c-3'-UTR was measured by Luciferase assay 48h post-transfection.

Results

HCV core proteins (1b, 3a) up-regulate the mRNA levels (1.358 times and 1.337 times respectively; P = 0.043, 0.008) and protein expression (1.608 times and 1.926 times respectively, P = 0.042, 0.008) of SREBP-1c. Both HCV core proteins of genotype 1b and 3a up-regulate the mRNA levels (1.566 times and 1.71 times respectively; P = 0.037, 0.006) and protein expression (1.436 times and 1.588 times respectively, P = 0.026, 0.009) of Sirt1. The Sirt1 promoter did not show significant transcriptional activity in the presence of HCV core proteins (1b, 3a). The relative luciferase activity of SREBP-1c-3'-UTR shown significant decrease in the presence of HCV core protein of genotype 3a (RQ = 0.667, P = 0.008).

Conclusions

HCV core proteins up-regulate the expression levels of SREBP-1c and Sirt1, which may be one of the pathogenesis of HCV related steatosis. MicroRNA also involved in the regulation of SEBP-1c by HCV core protein. Whether there are microRNAs contribute to the regulation of Sirt1 by HCV core protein still need to be investigated.

表1 real-time PCR定量检测脂代谢相关基因mRNA表达水平所用引物
图1 1b型HCV核心蛋白对脂代谢相关基因mRNA表达水平的影响
图2 HCV核心蛋白(1b、3a)上调SREBP-1c和Sirt1 mRNA表达水平
图3 HCV核心蛋白(1b、3a)上调SREBP-1c和Sirt1蛋白表达水平
图4 HCV核心蛋白对Sirt1启动子活性的影响
图5 microRNA对SREBP-1c的调控作用
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