Function and Expression of the Bile Acid Transport Protein m-Epoxide Hydrolase

胆汁酸转运蛋白间环氧化物水解酶的功能和表达

• 批准号: 7339304
• 负责人: DANIEL S LEVY
• 金额: $ 28.62万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 1979
• 资助国家: 美国
• 起止时间: 1979-08-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7339304
• 关键词: 3' Untranslated Regions; 5' Flanking Region; Affect; Agonist; Amish; Antibodies; Bile Acids; Binding; Biological Assay; Carcinogens; Carrier Proteins; Cells; Code; DNA Sequence Analysis; Defect; Disease; Drug Metabolic Detoxication; EPHX1 gene; Electrons; Electrophoretic Mobility Shift Assay; Endogenous Factors; Endoplasmic Reticulum; Epoxide hydrolase; Etiology; Exogenous Factors; Exons; Fibrates; Funding; Genetic Polymorphism; Genetic Transcription; Genomics; Genotype; Glucuronides; Hepatic; Hepatocyte; Human; Hydrocarbons; Hydrolase Gene; Individual; Introns; Knockout Mice; Knowledge; Label; Laboratories; Liquid Chromatography; Liver diseases; Malignant Neoplasms; Mediating; Membrane; Metabolic; Metabolism; Microsomal Epoxide Hydrolase; Mutation; Nuclear Receptors; PPAR alpha; Physiological Processes; Plasma; Play; Poison; Population; Predisposition; Procedures; Proteins; RNA Splicing; Radiolabeled; Regulation; Regulatory Element; Research; Research Personnel; Role; Serum; Site; Site-Directed Mutagenesis; Sodium; Technology; Transfection; Translations; Transmembrane Transport; Uridine Diphosphate Glucuronic Acid; Urine; Western Blotting; Xenobiotic Metabolism; Xenobiotics; chromatin immunoprecipitation; deletion analysis; expression vector; genetic linkage analysis; glucuronide; kindred; programs; promoter; radiotracer; tandem mass spectrometry; transcription factor; uptake

Human microsomal epoxide hydrolase (mEH) is a bifunctional protein that plays an important role in the sodium-dependent hepatic uptake of bile acids and in the metabolism of numerous xenobiotic carcinogens. Aberrant expression of mEH caused by polymorphisms in regulatory elements of the mEH gene (EPHX1) has been shown to result in defects in bile acid uptake resulting in hypercholanemia, and along with poly- morphisms in the coding region, in the altered metabolism of numerous carcinogens resulting in alterations in the susceptibility to various forms of cancer. Despite the critical role that mEH plays in bile acid transport and xenobiotic metabolism, the mechanisms that serve to regulate EPHX1 expression by endogenous and exogenous factors are now just beginning to be elucidated by studies from our laboratory. The long-term objectives of this research program are therefore to characterize the regulation of EPHX1 expression, further elucidate the role of mEH in mediating bile acid transport and to identify and functionally characterize human EPHX1 polymorphisms that affect transport function in hypercholanemic subjects. The specific aims of this proposal are A) to elucidate the transcriptional mechanisms involved in the bile acid induction of EPHX1 expression mediated by HNF-3beta, PPARalpha and PXR/CAR; B) to characterize mEH mediated transport of bile acids into hepatocytes and into the lumen of the endoplasmic reticulum where bile acid detoxification may take place via glucuronidation and C) to characterize the effects of human EPHX1 polymorphisms at regulatory sites or in the coding region on the transcription of EPHX1 and/or functional expression of mEH as related to the etiology of hypercholanemia. EPHX1 expression studies will use promoter activity assays, Northern and Western blot analyses, transcription factor expression vectors, footprinting, site-directed mutagenesis, electrophoretic mobility shift assays and chromatin immunoprecipitation. Transports studies will be done on wild type and mEH knockout mice. Polymorphism studies will utilize sequencing, genotyping and functional expression procedures. These studies should substantially increase our knowledge of this important transport/metabolic protein that plays a critical role in numerous physiological processes and in the etiology of several diseases including cholestatic liver disease.

人微粒体环氧化物水解酶（Human microsomal epoxy hydrolase，mEH）是一种双功能蛋白，在人肝微粒体中起重要作用。 胆汁酸的钠依赖性肝摄取和许多异生物质致癌物的代谢。 mEH基因调控元件（EPHX 1）多态性引起的mEH异常表达 已显示导致胆汁酸摄取缺陷，从而导致高胆血症，并且沿着多聚 在编码区的形态，在许多致癌物质的代谢改变，导致改变 对各种癌症的易感性。尽管mEH在胆汁酸转运中起着关键作用， 和外源性物质代谢，通过内源性和外源性代谢调节EPHX 1表达的机制， 我们实验室的研究刚刚开始阐明外源因素。长期 因此，本研究计划的目的是表征EPHX 1表达的调节，进一步 阐明mEH在介导胆汁酸转运中的作用， 影响高胆固醇血症患者转运功能的EPHX 1多态性具体目标是 A）阐明胆汁酸诱导EPHX 1的转录机制 由HNF-3 β、PPAR α和PXR/CAR介导的表达; B）表征mEH介导的转运 胆汁酸进入肝细胞和内质网腔， 可能通过葡萄糖醛酸化发生，以及C）表征人EPHX 1多态性在 调节位点或编码区中的EPHX 1的转录和/或mEH的功能表达， 与高胆血症的病因有关。EPHX 1表达研究将使用启动子活性测定， 北方和西方印迹分析，转录因子表达载体，足迹法，定点 诱变、电泳迁移率变动测定和染色质免疫沉淀。运输研究 将在野生型和mEH敲除小鼠上进行。多态性研究将利用测序、基因分型 和函数表达式程序。这些研究应该大大增加我们对这一点的认识 一种重要的转运/代谢蛋白，在许多生理过程中起关键作用， 包括胆汁淤积性肝病在内的几种疾病的病因。