Fiber Cell Membrane Organization-Role in Lens Architecture and Function

纤维细胞膜组织-在晶状体结构和功能中的作用

• 批准号: 9180703
• 负责人: P VASANTHA Rao
• 金额: $ 39.25万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-12-01 至 2019-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9180703
• 关键词: ANK2 gene; Actins; Address; Adhesions; Adhesives; Affect; Architecture; Area; Biochemical; Biological Assay; Blood Circulation; Calcium; Cataract; Cell Communication; Cell Shape; Cell membrane; Cell-Cell Adhesion; Cells; Characteristics; Complex; Connexins; Coupling; Crystalline Lens; Cytoskeletal Modeling; Cytoskeleton; Dystroglycan; Dystrophin; Etiology; Exhibits; Fiber; Gene Targeting; Genes; Geometry; Glycoproteins; Goals; Image; Immunoprecipitation; Impairment; In Vitro; Knock-in Mouse; Knock-out; Knockout Mice; L-Type Calcium Channels; Lens Fiber; MIP gene; Maintenance; Mechanics; Membrane; Membrane Proteins; Molecular; Mus; Myelin Sheath; Ocular Fixation; Optical Coherence Tomography; Optics; Phosphorylation; Physiological; Play; Property; Proteins; Proteomics; Radial; Regulation; Research; Research Project Grants; Resolution; Role; Scaffolding Protein; Scanning; Shapes; Spectrin; Structure; Tertiary Protein Structure; Testing; Tissues; Transmission Electron Microscopy; Visual Accommodation; Work; age related; base; fiber cell; filensin; in vivo; innovation; insight; lens; lens intrinsic protein MP 70; lens transparency; mechanical properties; mouse model; mutant; novel; null mutation; periaxin; ponsin; protein B; protein function; protein protein interaction; public health relevance; reconstruction; scaffold; skeletal

 DESCRIPTION (provided by applicant): Transparency and accommodation are crucial determinants of image focusing by the ocular lens and depend upon precise hexagonal geometry, radial packing, mechanical properties and membrane organization of fiber cells. The broad goals of this research project are to identify and characterize plasma membrane cytoskeletal scaffolding proteins regulating fiber cell shape, adhesion, tensile properties, membrane organization, and to determine how dysregulation of membrane cytoskeletal scaffolding activity impacts these vital characteristics of lens. This proposal will investigate th coordinated and interdependent scaffolding function(s) of Periaxin (PRX) and its interacting proteins- Ankyrin-B (AnkB) and the dystrophin- glycoprotein complex (DGC) in differentiating and mature fiber cells in the context of lens function. Our recent observations show that PRX and AnkB not only co-immunoprecipitate in a reciprocal manner but also exhibit a close spatial overlap of distribution profile in fiber cells. Importantly, mice with PRX null or heterozygous for AnkB null mutations (AnkB-/+) revealed prominent and similar disruptions in fiber cell shape, alignment and membrane clustering of connexin-50, filensin, spectrin, NrCAM and WAVE-2. Moreover, in AnkB-/+ lenses, PRX failed to anchor to the plasma membrane and in a vice versa manner, AnkB levels were decreased in PRX null lenses. Significantly in these lenses, membrane organization of the DGC was dramatically impaired indicating a close interaction among PRX, AnkB and the DGC. Although both PRX null and AnkB-/+ mouse lenses remained largely transparent, lens stiffness was found to be significantly decreased. These multiple and significant preliminary observations prompted us to hypothesize that the fiber specific PDZ- domain protein PRX serves as a key scaffolding protein via direct interaction with AnkB and the DGC, to maintain fiber cell hexagonal shape, radial and compact alignment, membrane domain organization and mechanical properties, which are critical determinants of lens transparency and accommodation. To determine the functional importance of the coordinated and essential scaffolding role(s) played by the PRX-AnkB-DGC axis in lens tissue, we will pursue the following three specific aims: 1. Detailed in vitro characterization of the interdependent scaffolding activities of PRX, AnkB and the DGC in mouse lens fibers, 2. Detailed in vivo analysis of how absence of PRX, AnkB and impaired DGC activity affects lens optical quality, fiber cell shape, adhesion and the membrane skeletal network, using gene targeted mice, and 3. Determine the role(s) of PRX-AnkB-DGC axis in organization and regulation of lens fiber cell channel protein membrane domains and activity, and in maintenance of lens mechanical properties using gene targeted mice. The completion of these unstudied aspects of fiber cells is expected to provide novel and important insights into how membrane cytoskeletal scaffolding interactions regulate the unique structure-function attributes of ocular lens in the context of its optical properties an accommodation.

 描述（由应用提供）：透明度和适应性是眼镜镜头焦点的至关重要决定者，并取决于精确的六边形几何形状，径向堆积，机械性能和纤维细胞的膜组织。该研究项目的广泛目标是识别和表征质膜细胞骨架脚手架蛋白质，以控制纤维细胞形状，粘合剂，拉伸性能，膜组织，并确定膜细胞骨架骨架活性的失调会影响这些镜头的这些重要特征。该提案将研究透镜功能的上下文中，在晶状体和成熟的纤维细胞中，该提案及其相互作用的蛋白质（PRX）及其相互作用的蛋白质及其相互作用的蛋白质 - 蛋白质蛋白 - Ankyrin-B（ANKB）和成熟的纤维细胞中的TH。我们最近的观察结果表明，PRX和ANKB不仅以相互的方式共免疫沉淀，而且还表现出纤维细胞中分布曲线的紧密空间重叠。重要的是，prx null或杂合子的小鼠适合 ANKB无效突变（ANKB - /+）揭示了纤维细胞形状，连接蛋白50，FILENSIN，SPECTIN，SPECTRIN，NRCAM和WAVE-2的纤维细胞形状，比对和膜聚类的显着干扰。此外，在ANKB - /+镜头中，PRX未能锚定在质膜上，反之亦然，PRX NULL透镜中的ANKB水平得到了提高。在这些镜头中，DGC的膜组织显着受损，表明PRX，ANKB和DGC之间存在密切相互作用。尽管PRX NULL和ANKB - /+小鼠透镜都在很大程度上保持透明，但发现透镜刚度得到了显着改善。这些多个和重要的初步观察结果促使我们假设纤维特异性的PDZ-romain PRX通过与ANKB和DGC直接相互作用，可以作为关键的脚手架蛋白质，以维持纤维细胞六边形的形状，辐射和紧凑的归档，膜结构域的组织和膜结构域的组织和机械性能。为了确定PRX-ANKB-DGC轴在镜头组织中扮演的协调和基本脚手架角色的功能重要性，我们将追求以下三个特定目标：1。详细介绍了PRX，ANKB和DGC在VIV和PRX中的prx和DGC的相互依存的钩子的体外表征。 DGC活性使用基因靶向小鼠影响镜头的光学质量，纤维细胞形状，粘合剂和膜骨骼网络，以及3。确定PRX-AKBB-DGC轴在组织中的作用，并使用基因靶标小鼠的镜头机械性能在组织中调节晶状体纤维细胞通道蛋白质膜和活性的镜头纤维细胞通道蛋白质膜和活性。预计这些未研究的纤维细胞的这些未研究的方面将提供新颖而重要的见解，以了解膜细胞骨架脚手架相互作用如何调节其光学特性中眼镜的独特结构 - 函数属性属性。