Identification of alpha-fetoprotein silencer and analysis of its regulatory mechanisms

甲胎蛋白(AEP)的沉默活性与出生后AFP表达的抑制有关。此外，在肝癌发生过程中，沉默蛋白活性的下调将导致AFP基因的重新表达。在本研究中，AFP Silecer与其增强子相互作用，导致增强子活性被抑制。此外，抑制AFP增强子活性抑制了AFP和白蛋白基因的表达。将视网膜母细胞瘤(Rb)基因或ras基因导入人肝癌细胞系HH-7中，转导Rb基因可上调肝细胞核因子-1(HNF-1)基因的表达，HNF-1基因是一种肝脏特异的转录因子，通过其启动子的激活刺激AFP和白蛋白基因的转录。转导ras基因可抑制AFP增强子活性，抑制AFP基因转录。此外，Rb基因在HH-7细胞中的转导不仅促进了HNF-1的表达，而且还促进了细胞生长的接触性抑制。因此，Rb基因被认为是分化诱导剂，尽管它不直接诱导AFP沉默活性。HMGI-C是参与DNA环形成的关键因子，在胎儿器官或某些恶性肿瘤细胞中高表达。用RT-PCR法分析HMGI-C在人肝癌细胞系中的表达，AFP产生细胞的HMGI-C表达明显高于非AFP产生细胞。这提示HMGI-C在人肝癌细胞中的表达与AFP沉默蛋白的活性有关。

The a-fetoprotein (AEP) silencer activity is relevant to repression of AFP expression after birth. Moreover, down-regulation of the silencer activity would lead to re-expression of the AFP gene in the hepatocarcinogenetic process. In the present study, the AFP silecer interacted with its enhancer, resulting in the inhibition of the enhancer activity. Furthermore, repression of the AFP enhancer activity depressed both the AFP and albumin gene expression. When retinoblastoma (Rb) gene or ras gene were transduced in HuH-7 human hepatoma cells, transduction of Rb gene up-regulated expression of the hepatocyte nuclear factor-1 (HNF-1) gene, a liver-specific transcriptional factor, which stimulated both the AFP and albumin gene transcription through activation of their promoters. Transduction of ras gene induced the suppression of the AFP enhancer activity and repressed the AFP gene transcription. In addition, the Rb gene transduction in HuH-7 cells not only stimulated HNF-1 expression but also promoted contact inhibition of cell growth. Thus the Rb gene was considered to function as a differeatiation inducer, although it did not directly induce the AFP silencer activity.HMGI-C is a key factor involved in DNA loop formation and expressed highly in fetal organs or certain malignant cells. Expression of HMGI-C was analyzed in human hepatoma cell lines by means of RT-PCR.Its expression was apparenty higher in AFP-producing cells than nonproclucing cells. This would suggest the association of HMGI-C expression with the AFP silencer activity in human hepatoma cells.