乙肝病毒X蛋白对肝癌细胞组蛋白H3K4甲基化修饰的影响及机制

【目的】 探讨乙肝病毒X蛋白(HBx)与肝癌细胞组蛋白H3K4位点甲基化修饰的关系及其可能的机制?【方法】构建稳定表达HBx基因的HepG2-hbx及载体对照细胞系HepG2-vc,同时培养用于阳性对照的肝癌细胞系HepG2.2.15及阴性对照细胞系HepG2;用酶标法检测各组细胞组蛋白H3K4位点甲基转移酶活性,比较HBx对H3K4位点甲基转移酶活性的影响;qRT-PCR检测各组细胞中H3K4位点特异性甲基转移酶SMYD3 ( SET and MYND domain containing3 )基因转录水平变化;Western-blot 检测SMYD3蛋白表达水平变化;免疫组化方法检测84例肝癌组织(包含:HbsAg(+)58例,HbsAg(-)26例)中SMYD3与组蛋白H3K4me3表达情况并对结果进行统计分析?【结果】稳定表达HBx基因的肝癌细胞系其组蛋白H3K4位点甲基化活性显著高于对照组细胞(P < 0.05),且SMYD3表达上调(P < 0.05);肝癌组织组蛋白SMYD3表达与患者是否存在HBV感染有关,并且促进H3K4甲基化修饰?【结论】 HBx上调肝癌细胞组蛋白H3K4位点甲基转移酶活性,这可能与其介导的SMYD3表达上调有关,并且在肝癌组织中由HBx介导的SMYD3上调也增强了组蛋白H3K4甲基化修饰?

Effect of Hepatitis B Virus X Protein on Histone H3K4 Methylation in Hepatocellular Carcinoma and Its Potential Mechanism

【Objective】 To investigate the relation between HBx protein expression and H3K4 methylation in hepatocellular carcinoma and its potential mechnism. 【Methods】 Cell line HepG2HBx that is able to stably express HBx andcontrol cell line HepG2vc was constructed and cultured, as well as positive control HepG2.2.15 and negative control HepG2. Histone H3K4 methyltransferase activity was detected by using ultraviolet spectrophotometer, and we explored the effection of HBx on Histone H3K4 methyltransferase activity. Using qRTPCR and Westernblot to evaluate the change of expression level of SMYD3 ( SET and MYND domain containing3 ) in each cell groups. Detecting the expression level of SMYD3 and H3K4me3 with immunohistochemical staining (HbsAg+ 58 cases, HbsAg 26 cases) and finally statistically analyzing the results. 【Results】 Histone H3K4 methyltransferase activity and the expression level of SMYD3 in cell line that express HBx are stably significantly higher than that of negative control groups (P < 0.05). Moreover, the expression of SMYD3 is directly relevant to the HBV infection in HCC tissue sample, and promoting the methylation of histone H3K4. 【Conclusion】 HBx enhance the activity of histone H3K4 methyltransferase in hepatoma cells, and the potential mechanism may be related to HBxregulated expression of SMYD3, which also promote the methylation of histone H3K4 in HCC tissue.