Elucidation of Arvcf-dependent mechanisms required for lens function

阐明晶状体功能所需的 Arvcf 依赖性机制

• 批准号: 10615869
• 负责人: Timothy F Plageman
• 金额: $ 39.68万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10615869
• 关键词: ARVCF gene; Address; Adherens Junction; Adhesions; Age Months; Animal Model; Architecture; Attenuated; Bilateral; Binding; Biochemical; Biological Models; Biology; Biomechanics; Blindness; Cadherin Domain; Cadherins; Cataract; Cell Adhesion; Cell Culture Techniques; Cell membrane; Cell physiology; Cellular Morphology; Cellular biology; Chromosome Deletion; Complex; Congenital chromosomal disease; Cytoskeletal Modeling; Data; Defect; Development; DiGeorge Syndrome; Disease; Distal; Electron Microscopy; Ensure; Epithelial Cells; Etiology; Event; Excision; F-Actin; Face; Generations; Genes; Genetic; Genetic Diseases; Guanosine Triphosphate Phosphohydrolases; Histologic; Human; Intercellular Junctions; Investigation; Knowledge; Length; Lens Fiber; Link; Longevity; MIP gene; Maintenance; Malignant Neoplasms; Maps; Measurement; Mediating; Microscopy; Molecular; Morphology; Mus; Mutation; N-Cadherin; Nature; Neoplasm Metastasis; Pathway interactions; Penetrance; Periodicity; Phenotype; Property; Protein Dynamics; Proteins; Proteomics; Public Health; Refractive Errors; Regulation; Schizophrenia; Shapes; Shprintzen syndrome; Signal Transduction; Structure; Techniques; Testing; Vision Disorders; Visual impairment; Water; age related; anticancer research; biophysical properties; brain research; cell type; cell water; experimental study; falls; fiber cell; genetic regulatory protein; inhibitor; lens; lens transparency; member; microscopic imaging; mutant; nanocluster; nervous system disorder; novel; preservation; protein complex; recruit; rho GTP-Binding Proteins; superresolution microscopy; translational impact; ultra high resolution

Abstract: The function of the lens requires the maintenance of its transparency and refractive properties throughout its lifespan but those mechanisms responsible are not well understood. While important cellular functions and pathways such as cell adhesion or GTPase signaling have long been hypothesized to play a role in preserving these functions the removal of genes associated with these cellular functions often severely disrupt the lens making the contribution of them challenging to interpret. We have overcome this barrier with the discovery that mice lacking the Arvcf gene develop bilateral, age- dependent cortical cataracts. Arvcf is a member of the p120-catenin subfamily of catenins that bind to a specific intracellular domain of cadherins and regulate Rho-GTPases and junctional protein dynamics and stability. We have additionally found that Arvcf is highly enriched within lens fiber cell junctional structures and is required for the recruitment of several proteins to the cadherin complex, normal lens refraction, preservation of fiber cell morphology, organization of the F-actin architecture, and the normal biomechanical properties of the lens. We propose three aims to address the central question: What molecular functions lie downstream of Arvcf to maintain lens transparency, refraction, cell morphology, and biomechanical properties? In the first aim we will determine which of these properties, precede, follow, or are simultaneous with cadherin complex and adherens junction instability in Arvcf deficient lenses. We will test the hypothesis that the initiating event is the reduction of cadherin complex proteins in fiber cell membranes through a combination of biophysical property measurements of whole lenses and fluorescent/electron microscopy of lens fiber cells. Our early investigations also found that Aquaporin 0 (AQP0) depends on Arvcf to associate with the cadherin complex and novel quantitative analysis of super-resolution microscopy images of interlocking protrusions demonstrated a significant reduction of AQP0 from the distal tips of these structures. Therefore, in the second Aim of this proposal we plan to test the hypothesis that Arvcf recruits AQP0 to the tips of interlocking protrusions to facilitate cell adhesion and water transport by determining whether AQP0 and the Arvcf/N-cadherin protein complex function together in adhesion and water transport. In the third aim we will test the hypothesis that altered GTPase signaling downstream of Arvcf contributes to lens function. The GTPase regulation domain of Arvcf and candidate GTPase regulatory proteins will be genetically altered to determine their contribution to Arvcf function, lens fiber cell biology, and lens transparency. Together, these project aims will identify functional mechanisms of a previously untested and important protein required for lens transparency and lens fiber cell function. Furthermore, they will elucidate mechanisms underlying the etiology of age-dependent cortical cataracts. Because of the association of murine Arvcf with cortical cataracts and the human ARVCF gene with genetic disorders, neurological diseases, and cancer, the advances made on this proposal will also have a broad impact.

抽象的： 镜片的功能需要在其整个使用寿命期间保持其透明度和折射特性，但是 那些负责的机制还没有被很好地理解。虽然重要的细胞功能和途径，例如细胞 长期以来，人们一直假设粘附或 GTP 酶信号传导在保留这些功能（去除基因）方面发挥着作用 与这些细胞功能相关的通常会严重破坏晶状体，使得它们的贡献具有挑战性 解释。我们克服了这一障碍，发现缺乏 Arvcf 基因的小鼠发育出双侧、年龄- 依赖性皮质白内障。 Arvcf 是连环蛋白 p120-连环蛋白亚家族的成员，可与特定的细胞内结合 钙粘蛋白结构域并调节 Rho-GTP 酶和连接蛋白动力学和稳定性。我们还发现 Arvcf 在晶状体纤维细胞连接结构中高度富集，并且是招募多种蛋白质所必需的 钙粘蛋白复合物、正常晶状体屈光、纤维细胞形态的保存、F-肌动蛋白的组织 结构以及晶状体的正常生物力学特性。我们提出三个目标来解决核心问题： Arvcf 下游有哪些分子功能来维持晶状体透明度、折射、细胞形态和 生物力学特性？在第一个目标中，我们将确定这些属性中的哪些先于、后于或同时存在 Arvcf 缺陷晶状体中钙粘蛋白复合物和粘附连接不稳定。我们将检验以下假设： 起始事件是通过生物物理的组合减少纤维细胞膜中的钙粘蛋白复合物蛋白 整个晶状体的特性测量和晶状体纤维细胞的荧光/电子显微镜。我们的早期调查 还发现水通道蛋白 0 (AQP0) 依赖于 Arvcf 与钙粘蛋白复合物结合并进行新的定量 对互锁突起的超分辨率显微镜图像的分析表明 AQP0 显着降低 从这些结构的远端尖端。因此，在该提案的第二个目标中，我们计划检验以下假设： Arvcf 将 AQP0 招募到互锁突起的尖端，通过确定来促进细胞粘附和水运输 AQP0 和 Arvcf/N-钙粘蛋白复合物是否在粘附和水运输中共同发挥作用。在第三个 我们的目标是检验 Arvcf 下游 GTPase 信号传导的改变有助于晶状体功能的假设。 GTP酶 Arvcf 和候选 GTPase 调节蛋白的调节域将被基因改变以确定它们的 对 Arvcf 功能、晶状体纤维细胞生物学和晶状体透明度的贡献。这些项目目标共同确定 晶状体透明度和晶状体纤维细胞所需的先前未经测试的重要蛋白质的功能机制 功能。此外，他们还将阐明年龄依赖性皮质白内障病因的潜在机制。因为 小鼠 Arvcf 与皮质性白内障以及人类 ARVCF 基因与遗传性疾病、神经系统疾病之间的关系 疾病和癌症方面，这项提案取得的进展也将产生广泛的影响。