Structure and Function of ROMK Channel in Kidney

肾脏ROMK通道的结构和功能

• 批准号: 7093104
• 负责人: STEVEN C HEBERT
• 金额: $ 32.97万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7093104
• 关键词: Bartter' s syndrome; X ray crystallography; Xenopus oocyte; binding sites; biological models; gene targeting; genetically modified animals; glycine; intermolecular interaction; kidney; laboratory mouse; magnesium ion; nucleotides; pharmacology; phosphatidylinositols; phosphorylation; potassium channel; protein kinase; protein structure function; secretion; structural biology; sulfonylurea

DESCRIPTION (provided by applicant): ROMK (Kir1.1) forms apical K channels in the distal nephron and collecting duct that provide the K secretory pathways for 1) K recycling necessary for NaCI absorption by the thick ascending limb and 2) K secretion by connecting duct and principal cells. Therefore, defining the function and regulation of the ROMK channels is important for understanding renal K handling and body K homeostasis. Loss-of-function mutations in ROMK cause Bartter's syndrome, a hypotensive renal salt wasting disease, that is due to loss of salt reabsorbing capacity by the thick ascending limb. We have developed the only ROMK Bartter's mouse with sufficiently high survival (>30%) to permit physiological study - and is the only mouse model of Bartter's with such high survival. We propose to continue to study the pathophysiology of mouse ROMK Bartter's to enhance our understanding of the role of ROMK in renal K handling and to suggest potential modalities for therapy of human Bartter's syndrome. We will also develop two new ROMK transgenic mice that will provide models for assessing both the specific roles of ROMK isoforms and the regulated trafficking of ROMK isoforms in native kidney cells. One of these isoforms, ROMK1, is only expressed in late distal tubule and collecting duct and has been suggested to play an important role in the regulation of renal K secretion by dietary K intake. We propose to generate a mouse deficient only in ROMK1 by selective deletion of the ROMK1-specific exon using a Cre-LoxP strategy. We hypothesize that this mouse will not have a Bartter's phenotype, but instead, have disordered K handling with increases or decreases in dietary K intake. We will also generate a ROMK1 BAC-transgenic mouse tagged with FLAG and EGFP to define ROMK trafficking in kidney epithelial cells and to determine associated proteins in the ROMK regulatory complex. These mice will also provide a resource for others interested in ROMK function in non- renal cells (Gl tract, brain, etc).

描述（由申请人提供）：ROMK（Kir1.1）在远端肾单位和集合管中形成顶端 K 通道，提供 K 分泌途径，用于 1）粗升肢吸收 NaCl 所需的 K 循环和 2）通过连接管和主细胞分泌 K 。因此，定义 ROMK 通道的功能和调节对于理解肾脏 K 处理和体内 K 稳态非常重要。 ROMK 的功能丧失突变会导致 Bartter 综合征，这是一种低血压肾盐消耗性疾病，这是由于粗大的升肢失去盐重吸收能力造成的。我们开发了唯一具有足够高存活率 (>30%) 以进行生理研究的 ROMK Bartter 小鼠，并且是唯一具有如此高存活率的 Bartter 小鼠模型。我们建议继续研究小鼠 ROMK Bartter 综合征的病理生理学，以增强我们对 ROMK 在肾脏 K 处理中的作用的理解，并提出治疗人类 Bartter 综合征的潜在方法。我们还将开发两种新的 ROMK 转基因小鼠，它们将为评估 ROMK 亚型的特定作用以及 ROMK 亚型在天然肾细胞中的调节运输提供模型。其中一种异构体 ROMK1 仅在晚期远端肾小管和集合管中表达，并被认为在饮食钾摄入调节肾钾分泌中发挥重要作用。我们建议通过使用 Cre-LoxP 策略选择性删除 ROMK1 特异性外显子来生成仅 ROMK1 缺陷的小鼠。我们假设这只小鼠不会具有巴特表型，而是随着膳食钾摄入量的增加或减少而出现钾处理紊乱。我们还将生成带有 FLAG 和 EGFP 标记的 ROMK1 BAC 转基因小鼠，以定义肾上皮细胞中的 ROMK 运输并确定 ROMK 调节复合物中的相关蛋白。这些小鼠还将为其他对非肾细胞（胃肠道、大脑等）中ROMK功能感兴趣的其他人提供资源。