Myo1c Participates in Podocyte Junction Formation Through Interaction with Neph1

描述（由申请人提供）：导致肾滤过损害性变化的肾小球疾病是全国范围内ESRD（终末期肾病）的一个日益增加的原因。不幸的是，管理ESRD发展的分子机制仍然知之甚少。足细胞蛋白的鉴定，包括Neph 1及其相关的蛋白复合物，是肾脏滤过的关键组成部分，是理解狭缝隔膜组织的一个重大进展。最近的一些研究，包括我们的研究，现在表明，Neph 1和它的相关蛋白，包括Grb 2，ZO-1，Nephrin和Par 3之间的动态相互作用调节肌动蛋白细胞骨架在狭缝隔膜。数据还表明，Neph 1及其复合物经常被发现在各种亚细胞隔室远离细胞膜肾小球损伤后。将这些蛋白质重新靶向回到它们的膜过滤狭缝位置可能需要基于分子马达的主动机制。我们现在有了令人兴奋的结果，确定了Neph 1与Myo 1c的一种新的相互作用，Myo 1c是非常规肌球蛋白家族的成员，其成员是与肌动蛋白结合并可以产生力的ATP酶马达蛋白。Myo 1c还具有膜结合和钙调蛋白结合结构域，并已显示在不同细胞类型的膜细胞骨架事件中发挥作用。我们最近发表了一篇文章，证明肾小球损伤导致Neph 1复合物的错误定位，导致细胞连接的丧失和蛋白尿，而从损伤中恢复与Neph 1复合物返回到细胞外周相关（瓦格纳等人，JBC 2008）。我们认为，相互作用的Neph 1与Myo 1c在调节组织的Neph 1复合物中起着至关重要的作用。在这项研究中，我们建议调查的Neph 1和Myo 1c在体外和体内条件下的相互作用。本研究的具体目标是：1）确定足细胞中Myo 1c的生理和病理生理组织。2)确定Myo 1c-Neph 1相互作用在响应肾小球损伤的Neph 1复合物组装和易位至滤过狭缝中的意义。3)确定Myo 1c及其与Neph 1的相互作用在体内条件下维持肾小球功能的重要性。这项研究的结果将增加我们对裂膜蛋白在生理和非生理条件下如何组织的理解，从而为旨在改善肾小球损伤后临床结果的研究开辟新的途径。 公共卫生相关性：在美国，导致终末期肾病的肾小球疾病是肾功能衰竭的主要原因。我们以前的研究表明，对维持肾脏过滤系统至关重要的蛋白质的功能和生化变化导致肾小球疾病的发展。在这项研究中，我们将确定一种新的蛋白质Myo 1c的作用，并确定用于开发新疗法的新靶点，旨在恢复各种肾小球疾病期间肾脏的选择性滤过功能。

DESCRIPTION (provided by applicant): Glomerular diseases that lead to damaging changes in kidney filtration are an increasing cause of ESRD (end stage renal disease) nationwide. Unfortunately, the molecular mechanisms governing the development of ESRD remain poorly understood. Identification of podocyte proteins, including Neph1 and its associated protein complex, that are critical components of kidney filtration was a major advancement in understanding the organization of slit diaphragm. A number of recent studies, including ours, now demonstrate that dynamic interactions between Neph1 and its associated proteins including Grb2, ZO-1, Nephrin and Par3 regulates the actin cytoskeleton at the slit diaphragm. Data also suggest that Neph1 and its complex are frequently found in various sub cellular compartments away from the cell membrane following glomerular injury. Retargeting these proteins back to their membrane filtration slit location is likely to require an active molecular motor-based mechanism. We now have exciting results that identify a novel interaction of Neph1 with Myo1c, a member of the un-conventional Myosin family whose members are ATPase motor proteins that bind to actin and can generate force. Myo1c also has membrane binding and Calmodulin binding domains and has been shown to have a role in membrane cytoskeletal events in diverse cell types. We recently published an article demonstrating that glomerular injury results in mis-localization of the Neph1 complex leading to loss of cell junctions and proteinuria, whereas recovery from injury correlates with the Neph1 complex's return to the cell periphery (Wagner et al., JBC 2008). We believe that the interaction of Neph1 with Myo1c plays a critical role in regulating the organization of the Neph1 complex. In this study we propose to investigate the Neph1 and Myo1c interaction under in-vitro and in- vivo conditions. The specific aims proposed for this study are: 1) Define the physiologic and pathophysiologic organization of Myo1c in podocytes. 2) Determine the significance of Myo1c-Neph1 interaction in the assembly and translocation of Neph1 complex to the filtration slit in response to glomerular injury. 3) Determine the significance of Myo1c and its interaction with Neph1 in maintaining the glomerular function under in-vivo conditions. The results from this study will increase our understanding of how slit diaphragm proteins organize under both physiological and nonphysiological conditions resulting in new avenues of research aimed at improving clinical outcome following glomerular injury. PUBLIC HEALTH RELEVANCE: Glomerular diseases leading to end stage renal disease are major cause of kidney malfunction in United States. Our previous studies have shown that functional and biochemical changes in the proteins that are central to the maintenance of the kidney filtration system leads to the development of glomerular disorders. In this study we will determine the role of a novel protein Myo1c and identify novel targets for use in the development of novel therapies aimed at restoring the selective filtration function of kidneys during various glomerular disorders.