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首页> 外文期刊>Molecular and Cellular Biology >Identification of protein-binding sites in the hepatitis B virus enhancer and core promoter domains.
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Identification of protein-binding sites in the hepatitis B virus enhancer and core promoter domains.

机译：鉴定乙型肝炎病毒增强子和核心启动子结构域中的蛋白质结合位点。

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摘要

We have investigated the role of liver-specific trans-acting factor(s) in the regulation of hepatitis B virus (HBV) gene expression. A recorder plasmid (pEcoAluCAT; HBV nucleotides 1 through 1878) was constructed containing the HBV enhancer and the promoter region of the pregenomic RNA, which was ligated to the bacterial chloramphenicol acetyltransferase (CAT) gene. Upon transfecting this plasmid into various cell lines, the CAT gene was expressed only in cells of liver origin. Moreover, competition cotransfections with pEcoAluCAT and plasmids containing HBV enhancer sequences in human hepatoblastoma-derived HepG2 cells indicated the presence of titratable trans-acting factor(s) in these cells. Gel mobility shift assays using HBV enhancer and core promoter domains confirmed the existence of sequence-specific DNA-binding proteins in liver cell nuclear extract which bound to these regions. These binding sites encompass 17- and 12-nucleotide palindromes in the HBV enhancer and core promoter domains, respectively, when mapped by the methylation interference assay.
机译：我们已经研究了肝特异性反式作用因子在乙型肝炎病毒（HBV）基因表达调控中的作用。构建了一个记录质粒（pEcoAluCAT; HBV核苷酸1至1878），其中包含HBV增强子和前基因组RNA的启动子区域，并与细菌氯霉素乙酰转移酶（CAT）基因连接。将这种质粒转染到各种细胞系中后，CAT基因仅在肝起源的细胞中表达。此外，在人肝母细胞瘤衍生的HepG2细胞中与pEcoAluCAT和含HBV增强子序列的质粒竞争共转染表明这些细胞中存在可滴定的反式作用因子。使用HBV增强子和核心启动子结构域的凝胶迁移率变动分析证实了与这些区域结合的肝细胞核提取物中存在序列特异性DNA结合蛋白。当通过甲基化干扰测定法作图时，这些结合位点分别在HBV增强子和核心启动子结构域中包含17和12个核苷酸的回文。

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