Transcriptional Regulation of Liver-specific Organic Anion Transporters and Those Clinical Application

1999年，我们克隆了一种新的人有机阴离子转运蛋白LST-1。LST-1在肝脏中强烈表达，其底物是胆汁酸、结合类固醇、类花生酸和一些药物。免疫组化显示LST-1定位于肝细胞窦膜。本研究旨在探讨人LST-1基因的转录调控机制及其临床应用。研究结果如下：1.采用HT-17细胞株进行荧光素酶分析，发现AP-1位点（转录起始位点上游-1100bp）和HNF-1位点（-60bp）是转录的重要区域.在HNF-1结合位点插入点突变的报告质粒荧光素酶活性降低.用HT 17细胞核提取物进行的电泳迁移率变动分析（EMSA）显示，用寡核苷酸（从-78到-48 μ g.加入HNF-1a抗体后，可观察到超移带，表明转录因子HNF-1a与LST-1基因上游约60 bp处结合，并被认为是LST-1基因在肝细胞中转录的最重要的转录因子。

In 1999, we have cloned a novel human organic anion transporter named LST-1. LST-1 is strongly expressed in liver and its substrates are bile acid, conjugated steroids, eicosanoids, and some kinds of drugs. Immunohistochemical analysis indicated that LST-1 was located at sinusoidal membrane of the hepatocytes. The aim of this study is to investigate the transcriptions]-regulation mechanism of human LST-1 gene and its clinical application. The results are follows;1. Luciferase assay using HT-17 cell lines derived from hepatocyte revealed that AP-1 site (-1100bp upstream from transcriptional starting site) and HNF-1 site (-60bp) were considered to be important region for transcription.2. Luciferase activity was decreased in the reporter plasmid which inserted point mutation in the HNF-1 binding site.3. Electrophoresis mobility shift assay (EMSA) using nuclear extract of HT17 revealed that specific protein-DNA complex was detected using oligonucleotide (from-78 to -48).4. Supershift band was observed by adding a HNF-1 a antibody.These results indicated that transcriptional factor, HNF-1 a was combined with about -60bp of LST-1 gene upstream and was considered to be most important by transcription of the LST-1 gene in hepatocytes.