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: 7663051
• 负责人: CHING-HWA SUNG
• 金额: $ 32.63万
• 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/7663051
• 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; 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; 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（由申请人提供）：视网膜色素上皮（RPE）是脊椎动物视网膜的多功能和必不可少的成分。它的不对称性，专门的膜结构和膜运动性对于其许多功能至关重要，在很大程度上取决于高度有序的细胞骨架。目前，关于调节细胞孔介导的RPE功能的机械和分子机制的了解相对较少。我们的实验室发现，最近鉴定出的肌动蛋白相关蛋白的Clic4在顶端RPE微绒毛中大量表达。在培养的RPE细胞中，Clic4似乎是吞噬杯中的关键成分，是吞噬感光体外部段的结构。为了研究Clic4在体内RPE中的作用，我们通过将siRNA（小干扰RNA）转染到啮齿动物的眼睛RPE中进行了CLIC4沉默。 clic4抑制的RPE细胞发生了几种形态学变化，包括缩短微绒毛和细胞接触的分解。此外，这些动物发展出深刻的视网膜脱离和感光受体萎缩，类似于先前描述的用于增殖性玻璃体病的表型。提出了三个具体目标。 AIM 1将识别Clic4在RPE和RPE-Photoreceptor互插的微伏和连接结构的起源中的直接参与。 AIM 2将研究Clic4和Ezrin之间的分子相互作用，肌动蛋白 - 质膜接头蛋白，以及这种相互作用在体内Clic4介导的RPE形态发生中的重要性。 AIM 3。将获得CLIC4通过RPE参与外部段吞噬作用的功能证据，并剖析涉及CLIC4的特定步骤。我们认为，这些研究将提供对正常和患病RPE中细胞骨架组织和调节的关键作用的新见解。这些不仅是重要的细胞生物学问题，而且与我们对增生性玻璃体病的病因以及其他退行性视网膜疾病的理解高度相关。