Shen R F, Li Y, Sifers R N, Wang H, Hardick C, Tsai S Y, Woo S L
Howard Hughes Medical Institute, Department of Cell Biology, Baylor College of Medicine, Houston, TX 77030.
Nucleic Acids Res. 1987 Oct 26;15(20):8399-415. doi: 10.1093/nar/15.20.8399.
Human alpha 1-antitrypsin (AAT) is expressed in the liver, and a 318 bp fragment immediately flanking the CAP site of the gene was found to be sufficient to drive the expression of a reporter gene (CAT) specifically in hepatoma cells. The enhancing activity however, was orientation-dependent. The DNA fragment was separated into a distal region and a proximal region. A "core enhancer" sequence GTGGTTTC is present within the distal region and is capable of activity enhancement in both orientations when complemented by the proximal region in the sense orientation. The results strongly suggest that there are multiple cis-acting elements in the human AAT gene that confer cell specificity for its expression. Nuclear proteins prepared from the hepatoma cells bound specifically to the proximal region in a band-shifting assay that was resistant to competition by the globin promoter DNA. Foot-printing analysis showed a protected domain within the proximal region that contains a nearly perfect palindromic sequence TGGTTAATATTCACCA, which may be important in the regulation of AAT expression in the liver.
人α1 -抗胰蛋白酶（AAT）在肝脏中表达，发现该基因CAP位点紧邻的一个318 bp片段足以驱动报告基因（CAT）在肝癌细胞中特异性表达。然而，增强活性具有方向依赖性。该DNA片段被分为远端区域和近端区域。在远端区域存在一个“核心增强子”序列GTGGTTTC，当与有义方向的近端区域互补时，该序列在两个方向上均能增强活性。结果强烈表明，人AAT基因中存在多个顺式作用元件，这些元件赋予其表达的细胞特异性。在凝胶迁移实验中，从肝癌细胞制备的核蛋白与近端区域特异性结合，且该结合不受珠蛋白启动子DNA竞争的影响。足迹分析显示近端区域内有一个受保护的结构域，其中包含一个近乎完美的回文序列TGGTTAATATTCACCA，这可能对肝脏中AAT表达的调控很重要。
