Isolation and characterization of rat kidney active urea transporter

大鼠肾活性尿素转运蛋白的分离和表征

• 批准号: 7531555
• 负责人: Guangping Chen
• 金额: $ 23.24万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-10 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7531555
• 关键词: Carrier Proteins; Cells; Cloning; Complementary DNA; DNA amplification; Electrodes; Epithelial Cells; Family; Genes; Goals; Homeostasis; Kidney; Label; Lead; Mediating; Messenger RNA; Methods; Molecular; Movement; Names; Nature; Nephrons; Nitrogen; Osmolar Concentration; Perfusion; Physiological; Play; Population; Property; Public Health; Rattus; Recycling; Role; Screening procedure; Sodium; Source; System; Testing; Tubular formation; Urea; Water; Xenopus oocyte; Yeast Model System; Yeasts; cDNA Library; expression vector; gene cloning; novel; solute; subtraction hybridization; success; uptake; urea transporter; urinary; voltage clamp; water conservation

DESCRIPTION (provided by applicant): Urea plays an important role in the urinary concentrating mechanisms. In the past decade, significant progress has been made in understanding urea reabsorption and recycling in kidney including the cloning of the two facilitated urea transporter genes, UT-A and UT-B. However, strong evidence from physiological studies demonstrated that the existence of a sodium- dependent active transporter in mammalian kidney, which can uphill move urea in some nephron segments. Though the active urea transporter was proposed over 40 years ago, none has yet been characterized. The overall objective of this project is to clone this novel active urea transporter from kidney inner medulla (IM) by using two independent approaches: 1) modified two-tester suppression subtraction hybridization (ttSSH) and 2) expression screening using a urea uptake defective yeast strain. After having successfully identified the novel urea transporter, we will investigate the nature of the active urea transporter. We hope the two approaches we apply here will lead us to discover the new genes. We believe the success of this project will fill in the gap with the molecular evidence how urea is actively transported in kidney and will extend our understanding on the urinary concentrating mechanism and water homeostasis. PUBLIC HEALTH RELEVANCE Strong physiological evidence demonstrated the existence of active urea transport activity in mammalian kidney over 40 years. The overall objective of this project is to clone the novel active urea transporter from kidney IM by using two independent approaches. The success of this project will extend our understanding on the urinary concentrating mechanism and water homeostasis.

描述（由申请人提供）：尿素在尿液浓缩机制中起着重要作用。在过去的十年中，在理解肾脏中的尿素重吸收和循环方面取得了重大进展，包括克隆两个促进尿素转运蛋白基因 UT-A 和 UT-B。然而，生理学研究的有力证据表明，哺乳动物肾脏中存在钠依赖性主动转运蛋白，它可以在某些肾单位节段中向上移动尿素。尽管活性尿素转运蛋白在 40 多年前就被提出，但尚未得到表征。该项目的总体目标是通过使用两种独立的方法从肾内髓质 (IM) 克隆这种新型活性尿素转运蛋白：1) 改良的双测试仪抑制消减杂交 (ttSSH) 和 2) 使用尿素吸收缺陷酵母菌株进行表达筛选。在成功鉴定出新型尿素转运蛋白后，我们将研究活性尿素转运蛋白的性质。我们希望我们在这里应用的两种方法能够引导我们发现新基因。我们相信该项目的成功将填补尿素如何在肾脏中主动转运的分子证据的空白，并将扩展我们对尿液浓缩机制和水稳态的理解。 公共卫生相关性 强有力的生理学证据证明哺乳动物肾脏中存在活跃的尿素转运活性超过 40 年。该项目的总体目标是通过使用两种独立的方法从肾脏 IM 克隆新型活性尿素转运蛋白。该项目的成功将扩展我们对尿液浓缩机制和水稳态的理解。