UNCONVENTIONAL MYOSINS IN RENAL FUNCTIONS

肾功能中的非常规肌球蛋白

• 批准号: 6105919
• 负责人: MARK MOOSEKER
• 金额: $ 18.46万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-06-01 至 1999-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6105919
• 关键词: actins; binding proteins; cell cell interaction; cell differentiation; cellular polarity; cytoskeleton; epithelium; genetically modified animals; kidney function; laboratory mouse; membrane proteins; myosins; nucleic acid sequence; protein isoforms; protein purification; renal ischemia /hypoxia; renal tubule; tight junctions; transfection

Description: (Abstract of the project) The proposed studies will continue the investigation of the role of the actin-based cytoskeleton in renal function and disease. We will continue to delineate the functional properties of the multiple actin-based motors expressed in the kidney with an emphasis on understanding the role of myosins in ischemic injury. Kidney ischemia leads to rapid changes in the actin cytoskeleton and a loss of membrane polarity in the proximal tubule cells. The hypothesis to be tested is that the ATP-dependent mechanochemical activity of one or more of the proximal tubule myosins is essential for maintenance of renal proximal tubule epithelial cell polarity, as well as for recovery of polarity following non-lethal ischemic injury. To understand these roles, we propose to purify and enzymatically characterize two proximal tubule expressed myosins, myosin-VI and myosin-Vlla. These studies will also include identification of myosin tail binding proteins that may link these myosins to the membrane cytoskeleton. We also propose to investigate the locations and cytoskeletal associations of an array of renal myosins during renal injury and recovery. These studies will include both immunolocation and biochemical extraction techniques to assess whether association of myosins with the membrane cytoskeleton is regulated during injury. As the characterization of renal myosins is still incomplete, we propose to identify, using a PCR-based screen, new renal myosins. Specifically, we will look for novel glomerular, distal tubule and collecting duct myosins as well as continue our analysis of proximal tubule myosins. Finally, we propose to assess whether mouse mutants with mutations in myosin-VI (Snell's waltzer) and myosin-VIIa (shaker-1) exhibit kidney dysfunction. These mutant mice will be assayed for sensitivity to renal injury as well as for susceptibility to dietary stress. By isolating and characterizing renal myosins we hope to identify which myosins are involved in the cytoskeletal and membrane rearrangements observed during recovery from ischemic injury.

产品描述：（项目摘要）拟议的研究将继续调查肌动蛋白为基础的细胞骨架在肾功能和疾病中的作用。我们将继续描述肾脏中表达的多种肌动蛋白为基础的马达的功能特性，重点是了解肌球蛋白在缺血性损伤中的作用。肾缺血导致肌动蛋白细胞骨架的快速变化和近端小管细胞膜极性的丧失。待检验的假设是，一种或多种近端小管肌球蛋白的ATP依赖性机械化学活性对于维持肾近端小管上皮细胞极性以及非致死性缺血损伤后极性的恢复至关重要。为了理解这些作用，我们建议纯化和酶法表征两种近端小管表达的肌球蛋白，肌球蛋白-VI和肌球蛋白-VIIa。这些研究还将包括鉴定可能将这些肌球蛋白连接到膜细胞骨架的肌球蛋白尾结合蛋白。我们还建议调查的位置和细胞骨架协会的一系列肾肌球蛋白在肾损伤和恢复。这些研究将包括免疫定位和生化提取技术，以评估肌球蛋白与膜细胞骨架的关联是否在损伤过程中受到调节。由于肾肌球蛋白的表征仍然是不完整的，我们建议确定，使用基于PCR的屏幕，新的肾肌球蛋白。具体而言，我们将寻找新的肾小球，远曲小管和集合管肌球蛋白，以及继续我们的近曲小管肌球蛋白的分析。最后，我们建议评估是否与肌球蛋白-VI（斯内尔的华尔兹）和肌球蛋白-VIIa（shaker-1）突变的小鼠突变体表现出肾功能障碍。将测定这些突变小鼠对肾损伤的敏感性以及对饮食应激的易感性。通过分离和鉴定肾肌球蛋白，我们希望确定哪些肌球蛋白参与缺血性损伤恢复过程中观察到的细胞骨架和膜重排。