Cytoskeleton's role in RPE's structure and function

细胞骨架在 RPE 结构和功能中的作用

• 批准号: 7475045
• 负责人: CHING-HWA SUNG
• 金额: $ 31.97万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7475045
• 关键词: Actins; Adhesions; Animals; Apical; Arts; Atrophic; Attenuated; Belief; Binding; Biochemistry; Biological; Biological Assay; Blood-Retinal Barrier; Cell membrane; Cells; Cytoskeletal Modeling; Cytoskeleton; Distal; Dominant-Negative Mutation; Electrons; Etiology; Event; Eye; F-Actin; Fostering; Homeostasis; In Vitro; Kinetics; Laboratory Finding; Life; Link; Localized; Mediating; Membrane; Microscopic; Modeling; Molecular; Morphogenesis; Names; Neural Retina; Phagocytosis; Phagosomes; Phenocopy; Phenotype; Photoreceptors; Physiological; Plant Roots; Process; Proteins; RNA Interference; Recruitment Activity; Regulation; Research Personnel; Resolution; Retina; Retinal; Retinal Detachment; Retinal Diseases; Rodent; Role; Series; Small Interfering RNA; Structure; Structure of retinal pigment epithelium; System; Techniques; Tight Junctions; Tissues; Transport Process; apical membrane; cell motility; cellular imaging; cellular microvillus; ezrin; gastrointestinal microvillus; human CLIC4 protein; in vitro Assay; in vivo; insight; mutant; novel; photoreceptor degeneration; polymerization; programs

DESCRIPTION (provided by applicant): The retinal pigment epithelium (RPE) is a multifunctional and indispensable component of the vertebrate retina. Its asymmetry, specialized membrane structures, and membrane motility, which are essential for many of its functions, rely heavily on a highly ordered cytoskeleton. At present, relatively little is known about the machinery and the molecular mechanism regulating cytoskelton-mediated RPE functions. Our lab found that CLIC4, a recently identified actin-associated protein, was abundantly expressed in apical RPE microvilli. In cultured RPE cells, CLIC4 appears to be a key component in the phagocytic cup, the structure that engulfs the photoreceptor outer segment. To study CLIC4's role in RPE in vivo, we performed CLIC4 silencing by transfecting siRNA (small interfering RNA) into RPE of rodent eyes. The CLIC4-suppressed RPE cells developed several morphological changes including shortening of microvilli and breakdown of cell- cell contacts. Moreover, these animals developed profound retinal detachment and photoreceptor atrophy, resembling those phenotypes previously described for proliferative vitroretinopathy. Three specific aims are proposed. Aim 1 will identify the direct involvement of CLIC4 in the genesis of the microvillar and junctional structure of RPE and RPE-photoreceptor interdigitation. Aim 2 will investigate the molecular interactions between CLIC4 and Ezrin, an actin-plasma membrane linker protein, and the importance of such interactions in CLIC4-mediated RPE morphogenesis in vivo. Aim 3. will obtain functional evidence for CLIC4's involvement in outer segment phagocytosis by RPE and dissect the specific step(s) in which CLIC4 is involved. It is our belief that these studies will provide novel insights into the pivotal role of cytoskeletal organization and regulation in both normal and diseased RPE. These are not only important cell biological questions but also highly relevant for our understanding of the etiology of proliferative vitroretinopathy and perhaps also other degenerative retinal diseases.

描述（申请人提供）：视网膜色素上皮（RPE）是脊椎动物视网膜的多功能且不可或缺的组成部分。它的不对称性、特殊的膜结构和膜运动性对其许多功能至关重要，在很大程度上依赖于高度有序的细胞骨架。目前，对于调节细胞骨架介导的 RPE 功能的机制和分子机制知之甚少。我们的实验室发现 CLIC4（一种最近发现的肌动蛋白相关蛋白）在顶端 RPE 微绒毛中大量表达。在培养的 RPE 细胞中，CLIC4 似乎是吞噬杯的关键组成部分，吞噬杯是吞噬光感受器外节的结构。为了研究 CLIC4 在体内 RPE 中的作用，我们通过将 siRNA（小干扰 RNA）转染到啮齿动物眼睛的 RPE 中来进行 CLIC4 沉默。 CLIC4 抑制的 RPE 细胞出现了多种形态变化，包括微绒毛缩短和细胞-细胞接触破坏。此外，这些动物出现严重的视网膜脱离和光感受器萎缩，类似于先前描述的增殖性玻璃体视网膜病变的表型。提出了三个具体目标。目标 1 将确定 CLIC4 在 RPE 的微绒毛和连接结构以及 RPE-光感受器交叉的发生中的直接参与。目标 2 将研究 CLIC4 和 Ezrin（一种肌动蛋白-质膜连接蛋白）之间的分子相互作用，以及这种相互作用在 CLIC4 介导的体内 RPE 形态发生中的重要性。目标 3. 将获得 CLIC4 参与 RPE 外节吞噬作用的功能证据，并剖析 CLIC4 参与的具体步骤。我们相信，这些研究将为细胞骨架组织和调节在正常和患病 RPE 中的关键作用提供新的见解。这些不仅是重要的细胞生物学问题，而且与我们对增殖性玻璃体视网膜病变以及其他退行性视网膜疾病的病因学的理解高度相关。