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

肝脏特异性有机阴离子转运蛋白的转录调控及其临床应用

• 批准号: 12671195
• 负责人: UNNO Michiaki
• 金额: $ 2.56万
• 依托单位: TOHOKU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671195/
• 关键词: Liver specific organic anion transporter-1 (LST-1); transcriptional factor; HNF-1a; Luciferase assay; トランスポーター; NF-E2; HT17; AP-1; 遺伝子上流域; ケノデオキシコール酸

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.

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.使用HT 17核提取物进行的电泳迁移率变动分析（EMSA）显示，使用寡核苷酸检测到特异性蛋白质-DNA复合物（从-78到-48）。4.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。

项目成果

Abe,T.,M.Unno, et al.: ""LST-2, a human liver-specific organic anion transporter determines methotrexate sensitivity in the gastrointestinal cancers.""Gastroenterology. (in press). (2001)

Abe,T.,M.Unno 等人：“LST-2，一种人类肝脏特异性有机阴离子转运蛋白，决定胃肠癌中甲氨蝶呤的敏感性。”胃肠病学。

阿部高明, 海野倫明, 他: "有機アニオントランスポーターoatp/LSTとその機能"蛋白質核酸酵素. 46巻5号. 612-620 (2001)

Takaaki Abe、Michiaki Unno 等：“有机阴离子转运蛋白 OATP/LST 及其功能”《蛋白质核酸酶》第 46 卷，第 5 期。612-620（2001 年）

Ito A, Yamaguchi T, Onogawa T, Unno M, et al.: "Distribution of organic anion-transporting polypeptide2(oatp2) and oatp3 in the rat retina"Invest Ophthalmol Vis Sci. 43,(3). 858-863 (2002)

Ito A、Yamaguchi T、Onokawa T、Unno M 等人：“有机阴离子转运多肽 2(oatp2) 和 oatp3 在大鼠视网膜中的分布”Invest Ophasemol Vis Sci。

Nishio,T.,H.Adachi, et al.: "Molecular identification of a rat novel organic anion transporter moatl, which transports prostaglandin D (2), leukotriene C (4), and taurocholate."Biochem Biophys Res Commun. 275(3). 831-8 (2000)

Nishio,T.,H.Adachi 等人：“大鼠新型有机阴离子转运蛋白 moatl 的分子鉴定，该转运蛋白转运前列腺素 D (2)、白三烯 C (4) 和牛磺胆酸盐。”Biochem Biophys Res Commun。

Akiyama T.,Unno, M. et al.: "Activation of Reg gene, a gene for insulin-producing beta-cell regeneration : Poly (ADP-ribose) polymerase binds Reg promoter and regulates the transcription by autopoly (ADP-ribosyl) ation."Proc Natl Acad Sci USA. 98(1). 48-5

Akiyama T.、Unno、M. 等人：“Reg 基因的激活，一种用于产生胰岛素的 β 细胞再生的基因：聚 (ADP-核糖) 聚合酶结合 Reg 启动子并通过自聚 (ADP-核糖基) 调节转录