Novel Cilia Trafficking Mechanisms

新颖的纤毛贩运机制

• 批准号: 7753753
• 负责人: BENJAMIN L MARGOLIS
• 金额: $ 35.09万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7753753
• 关键词: Affect; Apical; Binding; Binding Proteins; Cell Nucleus; Cell division; Cells; Centrosome; Cilia; Cyst; Cystic Kidney Diseases; Epithelial Cells; Etiology; Guanosine Triphosphate; Heart; Importins; Integral Membrane Protein; Kidney; Kidney Failure; Laboratories; Liquid substance; Mammalian Cell; Mediating; Molecular; Monomeric GTP-Binding Proteins; Movement; Mutation; Nuclear; Nuclear Export; Nuclear Import; Organelles; Play; Polycystic Kidney Diseases; Protein Import; Protein Isoforms; Proteins; RNA Splicing; Regulation; Retinitis Pigmentosa; Role; Running; Structure; Surface; System; Testing; Transmembrane Domain; Travel; Work; alpha Karyopherins; base; cilium biogenesis; insight; kinetosome; novel; protein transport; public health relevance; trafficking

DESCRIPTION (provided by applicant): Recent evidence implicates cilia abnormalities as important in the etiology of Polycystic Kidney Disease. Our laboratory has defined an important role for polarity proteins, such as the apical transmembrane protein Crumbs3, in ciliogenesis. In the course of our study on Crumbs3, we have identified a novel cilia and centrosomal trafficking mechanism that involves the Ran/Importin system. Ran is a small GTPase that is concentrated in the nucleus and regulates the import of proteins into the nucleus in combination with Importins. Importins travel into the nucleus carrying their cargo and when they bind to RanGTP, the cargo is released leading to overall nuclear import. However Ran is also concentrated at other points in the cell including the centrosome. We have found that Importin beta1 can bind to a splice form of Crumbs3, called Crumbs3-CLPI, and target this isoform of Crumbs to the centrosome. We also have obtained evidence that another cilia protein, called RP2, also uses the Importin system for trafficking to the cilia. Our hypothesis that guides this proposal is that the Ran/Importin system plays a major role in regulation of cilia and centrosomal trafficking. To test this hypothesis, we propose three specific aims. The first specific aim will be to identify the molecular basis for the Crumb3-CLPI interaction with Importins and determine how mutations that alter this interaction affect the trafficking of this isoform of Crumbs3. The second specific aim will test the role of other Importins, including Importin alphas, in this trafficking mechanism and in cilia function in general. In the last specific aim we will manipulate the levels of RanGTP at the centrosome and determine the effect of this manipulation on centrosome and cilia trafficking. We will also work to identify Ran and Importin alpha binding proteins at the centrosome. At the conclusion of these studies we will have a greater understanding of the role of the Ran/Importin system in cilia trafficking networks. This will have important implications for our understanding of Polycystic Kidney Disease and other ciliopathies. PUBLIC HEALTH RELEVANCE: The aim of this proposal is to study mechanisms that control the movements of proteins into a cell organelle called the cilia. Recent studies have suggested that defective function of the cilia leads to cystic diseases of the kidney. Cystic diseases of the kidney, where the normal kidney is replaced by non-functioning fluid filled cysts, are a major cause of kidney failure.

描述（由申请人提供）：最近的证据表明纤毛异常在多囊肾病的病因学中非常重要。我们的实验室已经确定了极性蛋白（例如顶端跨膜蛋白 Crumbs3）在纤毛发生中的重要作用。在我们对 Crumbs3 的研究过程中，我们发现了一种涉及 Ran/Importin 系统的新型纤毛和中心体运输机制。 Ran是一种小型GTP酶，集中在细胞核中，与Importins结合调节蛋白质进入细胞核。输入蛋白携带其货物进入细胞核，当它们与 RanGTP 结合时，货物被释放，导致整体核输入。然而，Ran 也集中在细胞的其他位置，包括中心体。我们发现 Importin beta1 可以与 Crumbs3 的剪接形式（称为 Crumbs3-CLPI）结合，并将这种 Crumbs 异构体靶向中心体。我们还获得了证据表明另一种纤毛蛋白 RP2 也使用 Importin 系统运输至纤毛。指导这一提议的假设是 Ran/Importin 系统在纤毛和中心体运输的调节中发挥着重要作用。为了检验这一假设，我们提出了三个具体目标。第一个具体目标是确定 Crumb3-CLPI 与 Importins 相互作用的分子基础，并确定改变这种相互作用的突变如何影响 Crumbs3 亚型的运输。第二个具体目标将测试其他导入蛋白（包括导入蛋白 alpha）在这种贩运机制和纤毛功能中的作用。在最后一个具体目标中，我们将操纵中心体的 RanGTP 水平，并确定这种操纵对中心体和纤毛运输的影响。我们还将致力于鉴定中心体的 Ran 和 Importin α 结合蛋白。在这些研究结束时，我们将对 Ran/Importin 系统在纤毛贩运网络中的作用有更深入的了解。这对于我们了解多囊肾病和其他纤毛病具有重要意义。公共健康相关性：该提案的目的是研究控制蛋白质运动进入称为纤毛的细胞器的机制。最近的研究表明，纤毛功能缺陷会导致肾脏囊性疾病。肾脏囊性疾病是导致肾衰竭的主要原因，其中正常肾脏被无功能的充满液体的囊肿取代。