Functions for Myosin VI in the kidney proximal tubule

肌球蛋白 VI 在肾近曲小管中的功能

• 批准号: 7034337
• 负责人: MARK S MOOSEKER
• 金额: $ 33.43万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7034337
• 关键词: actins; adenosine diphosphate; biomechanics; calcium ion; cell line; cytoskeleton; endocytosis; epithelium; genetic regulation; genetically modified animals; glomerular filtration rate; hormone regulation /control mechanism; laboratory mouse; molecular dynamics; myosins; parathyroid hormones; phenotype; phosphorylation; podocyte; protein structure function; renal glomerulus; renal ischemia /hypoxia; small interfering RNA; sodium hydrogen exchanger; streptozotocin; wound healing

The general objective of the proposed studies is to continue the molecular and functional characterization of myosin-VI (Myo6). Myo6 is unique among known members of the myosin superfamily of actin based molecular motors in that it moves in the opposite direction along the actin filament, toward the pointed/minus end. Past studies have implicated several potential functions for Myo6 including clathrin mediated endocytosis, Golgi traffic and cellular motility. The health relevance of the proposed studies has been well established as this myosin is a target for mutations that lead to inherited deafness and cardiac myopathies. It also has been shown to be a key contributor to the invasiveness of ovarian tumor cells. The goals of this project are three fold. The first will be to conduct an extensive characterization of the motor properties of native tissue purified Myo6. Our studies indicate that native Myo6 has properties significantly different from in-vitro expressed Myo6. Proposed studies will examine the motility of native Myo6 with respect to duty ratio and regulation by calcium, phosphorylation and load. A major focus will be the molecular basis for the conversion of Myo6 from a minus- to a plus- end directed motor. The second goal will be the phenotypic characterization of the cellular and physiologic defects in the proximal tubule epithelial (PTE) cells of the kidney in the Myo6 mutant mouse?Snell's waltzer. Like humans with Myo6 mutations, these mice are deaf due to degeneration of the neurosensory epithleum of the inner ear. Although this is the only overt dysfunction in these mice, our preliminary findings indicate that there are also serious cellular deficiencies in a number of tissues that express Myo6 including the renal proximal tubule. Studies will include assessment of defects in PTE architecture, endocytosis of glomerular filtrate, and responses to renal injuries including ischemia and chemically induced diabetes. The third goal will focus on the in vivo dynamics of fluorescently tagged Myo6 expressed in a PTE cell line and how cellular functions including endocytosis, wound healing and organization of the actin cytoskeleton are affected by targeted disruption of Myo6 function.

拟议的研究的一般目标是继续肌球蛋白VI的分子和功能表征（MyO6）。在基于肌动蛋白的分子电机的肌球蛋白超家族的已知成员中，MyO6是独一无二的，因为它沿着肌动蛋白细丝向相反的方向移动到尖头/负端。过去的研究暗示了MyO6的几个潜在功能，包括网格蛋白介导的内吞作用，高尔基体交通和细胞运动。拟议研究的健康相关性已得到很好的确定，因为这种肌球蛋白是导致遗传性耳聋和心脏肌病的突变的靶标。它也已被证明是卵巢肿瘤细胞侵入性的关键因素。该项目的目标是三倍。第一个是对电动特性的广泛表征 天然组织纯化的肌6。我们的研究表明，本地肌6具有与体外表达的Myo6的特性明显不同。拟议的研究将检查天然Myo6在占用比和钙，磷酸化和负载的调节方面的运动。主要的重点将是将Myo6从负端转换为正端导向电动机的分子基础。第二个目标是在Myo6突变小鼠？Snell的Waltzer中肾脏近端上皮（PTE）细胞中细胞和生理缺陷的表型表征。像具有肌6突变的人一样，由于内耳神经感觉上皮的变性，这些小鼠聋。尽管这是这些小鼠中唯一的明显功能障碍，但我们的初步发现表明，在许多表达MyO6（包括肾脏近端小管）的组织中也存在严重的细胞缺陷。研究将包括评估 PTE结构缺陷，肾小球滤液的内吞作用以及对包括缺血和化学诱导糖尿病在内的肾脏损伤的反应。第三个目标将集中在PTE细胞系中表达的荧光标记的荧光标记的体内动力学以及细胞功能如何受到肌动蛋白细胞骨架的内吞作用，伤口愈合和组织受到肌动蛋白的靶向破坏的影响。