Glomerular Capillaries-- Normal and Pathologic

在过去的39年里，在这项资助下，我们的研究一直致力于了解 肾小球通透性和蛋白质吸收的细胞和分子机制以及 肾小球疾病中这些过程的紊乱。在此提出的实验 应用代表了我们正在进行的狭缝光阑表征工作的直接延续， GIV在足细胞损伤后足细胞功能和存活中的作用。我们将重点关注三个具体目标： 具体目标#1：表征和确定新型假定狭缝光阑的功能 通过MS和定量细胞器蛋白质组学在狭缝隔膜组分中鉴定组分。我们将 验证它们的连接定位并确定它们在足细胞中的结合伴侣和功能。 具体目标#2：研究GIV在被动免疫中介导Akt信号传导和细胞存活中的作用。 海曼肾炎、局灶性肾小球硬化和糖尿病肾病。在PAN肾病中，GIV是 上调，组装VEGFR 2/GIV/Gal 3复合物，并起到维持足细胞存活的作用。我们 将确定GIV是否在与足细胞损伤相关的其他疾病中发挥类似作用。 具体目标#3：进一步研究GIV影响的机制和时空方面， 足细胞我们的初步数据表明，GIV可能在局灶性粘连（FA）和/或裂隙处起作用。 因为它与黏着斑蛋白和VEGFR 2共定位在FA处，并且它结合CD 2AP。我们将调查 通过活细胞成像，使用邻近性，动态观察GIV与VEGFR 2和CD 2AP之间的相互作用 连接分析和超分辨率IF成像（STORM），以确定相互作用发生的位置，并测试 GIV缺失或GIV突变体的表达是否影响FA或连接形成。 这些研究有望为理解细胞和分子生物学提供新的见解。 肾小球滤过和肾小球损伤机制及其在肾小球疾病中的变化 与蛋白尿有关。 相关性（参见说明）： 足细胞损伤是许多肾小球疾病的起始原因。计划中的研究将提供关键的 深入了解调节足细胞组织，功能和生存的信号网络， 与蛋白尿相关的疾病中的肾小球损伤。

During the last 39 years under this grant our research has been directed toward understanding the cellular and molecular mechanisms of glornerular permeability and protein absorption as well as the derangements in these processes that occur in glomerular diseases. The experiments proposed in this application represent a direct continuation of our ongoing work on characterization of slit diaphragms and the role of GIV in podocyte function and survival after podocyte injury. We will focus on three specific aims: Specific Aim # 1 : To characterize and determine the functions of novel, putative slit diaphragm components identified by MS and quantitative organellar proteomics in slit diaphragm fractions. We will verify their junctional localization and determine their binding partners and functions in podocytes. Specific Aim #2: To investigate the role of GIV in mediating Akt signaling and cell survival in passive Heymann nephritis, focal glomerulosclerosis, and diabetic nephropathy. In PAN nephrosis GIV is upregulated, assembles a VEGFR2/GIV/Gal3 complex, and functions to maintain podocyte survival. We will determine if GIV plays a similar role in other diseases associated with podocyte injury. Specific Aim #3: To further investigate the mechanisms and spatial-temporal aspects of GIV's effects on podocytes. Our preliminary data indicate that GIV might function at focal adhesions (FA) and/or at slit diaphragms as it colocalizes with vinculin and VEGFR2 at FA and it binds CD2AP. We will investigate the dynamics ofthe interactions between GIV and VEGFR2 and CD2AP by live cell imaging, use proximity ligation assays and superresolution IF imaging (STORM) to determine where interaction occurs, and test whether GIV depletion or expression of GIV mutants affect FA or junction formation. These studies can be expected to provide novel insights into understanding ofthe cellular and molecular mechanisms of glomerular filtration and glomerular injury and their alterations in glomerular diseases associated with proteinuria. RELEVANCE (See instructions): Podocyte injury is the initiating cause of many glomerular diseases. The studies planned will provide key insights into the signaling neworks that regulate podocyte organization, functions and survival in response to glomerular injury in diseases associated with proteinuria.