乙型肝炎病毒HBeAg上调S100钙结合蛋白A11基因启动子表达活性的研究

目的 探讨乙型肝炎病毒e抗原(HBeAg)对S100钙结合蛋白A11(calgizzarin S100A11)启动子转录的激活作用。方法 以我室构建的HBeAg反式调节基因的cDNA文库抑制性消减杂交(SSH)筛选结果为基础，利用生物信息学技术确定S100A11的启动子区域(S100A11-P)，聚合酶链反应(PCR)扩增S100A11-P，克隆至真核报告载体PCAT3中，构建pCAT3-S100-p报告载体，以该质粒转染肝癌细胞系HepG2细胞系，用酶联免疫吸附法(ELISA)检测氯霉素乙酰转移酶(CAT)的表达活性，并与pcDNA3．1(-)-HBeAg共转染HepG2细胞系，用ELISA法检测CAT的表达活性。结果pCAT3-S100-P在HepG2细胞中能够指导CAT的表达；共转染实验中pCAT3-S100-P-pcDNA3.1(-)-HBeAg组CAT的表达活性是pCAT3-S100-P组的6．1倍。结论 我室克隆的S100A11启动子有顺式激活下游基因的活性，HBeAg具有对S100A11的反式激活作用。本实验进一步验证了我室利用SSH技术研究HBeAg反式激活作用的结果。

The study of up-regulating effect of calgizzarin S100A11 gene promoter by hepatitis B virus e antigen

Objective To investigate activity of hepatitis B virus e antigen (HBeAg) on calgizzarin S100A11 gene promoter. Methods The sequence of calgizzarin S100A11 gene promoter was identified in GenBank by bioinformatics and amplified from HepG2 genome by polymerase chain reaction (PCR). The amplified product was cloned into pCAT3 reporter vector to construct PCAT3-S100-p. The HepG2 cell line was transfected by pCAT3-S 100-p, and was co-tranfected by pCAT3-S100-p and pcDNA3.1(-)-HBeAg. The choloraphenical acetyltransferase (CAT) activity was detected by enzyme-linked immunosorbent assay (ELISA) kit did.Results pCAT3-SI00-p had higher activity of CAT than pCAT3-basic by ELISA kit did. The expression of CAT in co-transfection of pCAT3-S 100-p and pcDNA3.1(-)-HBeAg was 6.1 times higher than that in pCAT3-S100-p plasmid.Conclusion HBeAg can transactivate the expression of S100A11 protein, and it testified our previous results by suppression subtractive hybridization (SSH).