乙肝病毒X蛋白结合蛋白(HBXIP)增强肝癌细胞NF-κB转录活性的实验研究
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国家自然科学基因资助项目(30670959).


Promotion of Transcriptional Activity of NF-κB Mediated by HBXIP in Hepatoma Cells
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This work was supported by a grant from The National Natural Science Foundation of China (30670959).

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    摘要:

    前期研究结果表明,乙型肝炎病毒X蛋白结合蛋白 (hepatitis B virus X-interacting protein,HBXIP)具有促进细胞增殖的作用.为了进一步阐明其分子机制,观察了HBXIP对核因子κB (NF-κB) 转录活性的影响.实验中通过基因共转染将NF-κB报告基因质粒pNF-κB-Luc和HBXIP真核表达载体pcDNA3-hbxip导入人肝癌H7402细胞系中,进行荧光素酶活性分析.结果显示:H7402细胞过表达HBXIP后NF-κB的转录活性明显增强;此外,基因转染后经免疫印迹检测显示,与NF-κB二聚体结合的抑制亚基IκBα的磷酸化水平明显增加;同时,提取H7402细胞的核蛋白,然后应用免疫印迹检测细胞核中p65/NF-κB的水平.结果显示,H7402细胞中HBXIP过表达后细胞核中p65/NF-κB的水平明显增加.当应用RNA干扰技术抑制了细胞内源性的HBXIP基因表达后,则出现与上述结果相反的效果.上述结果提示,HBXIP可增加核内p65/NF-κB蛋白水平,进而发挥NF-κB促转录调控的作用.因此,HBXIP可通过调控NF-κB信号途径而促进细胞增殖.

    Abstract:

    Hepatitis B virus X-interacting protein (HBXIP), a cofactor of survivin, previously had been found to promote cell proliferation. In order to demonstrate the molecular mechanismsthe effect of HBXIP on transcriptional activity of NF-κB was investigated. The eukaryotic expression vector of HBXIP (pcDNA3-hbxip), or the RNA interference vector of HBXIP (pSilencer-hbxip) and NF-κB luciferase reporter (pNF-κB-Luc) were co-transfected into H7402 hepatoma cells. The pRL-TK vector was used as an internal control for normalization of luciferase activity, and the luciferase activity was determined by using the dual luciferase reporter assay system. The luciferase assay indicated that the over-expression of HBXIP could stimulated NF-κB transcriptional activity in H7402 hepatoma cells. RNA interference (RNAi) targeting HBXIP mRNA resulted in the opposite effects. Statistically no significant difference was observed between the control cells and pcDNA3 empty vector-transfected cells or pSilencer-control transfected cells. The phosphorylation level of IκBα, an inhibitor of NF-κB, was increased by Western blot analysis when over-expression of HBXIP in H7402 cells. The p65/NF-κB level in nucleus extraction from H7402 cells was examined by Western blot analysis after transfection. The data indicated that over-expression of HBXIP in H7402 cells was able to increase the levels of p65/NF-κB in the nucleus. However, down-regulation of HBXIP mRNA in the cells by RNAi resulted in the opposite results. In conclusion, HBXIP is involved in cell proliferation by regulating NF-κB signal pathway.

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王凤泽,沙丽,乔玲,吴莲英,张晓东,叶丽虹.乙肝病毒X蛋白结合蛋白(HBXIP)增强肝癌细胞NF-κB转录活性的实验研究[J].生物化学与生物物理进展,2007,34(11):1197-1201

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  • 收稿日期:2007-03-23
  • 最后修改日期:2007-06-10
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  • 在线发布日期: 2007-06-15
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