Interaction of alpha-crystallin with cholesterol bilayer domains in cataract formation

α-晶状体蛋白与胆固醇双层结构域在白内障形成中的相互作用

• 批准号: 10653155
• 负责人: Laxman Mainali
• 金额: $ 29.04万
• 依托单位: BOISE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10653155
• 关键词: Affect; Affinity; Age; Animals; Binding; Biological; Cataract; Cell Nucleus; Cell membrane; Cholesterol; Crystalline Lens; Crystallins; Cytoplasm; Development; Electron Spin Resonance Spectroscopy; Eye; Goals; Health; Homeostasis; Human; Individual; Knowledge; Lead; Lecithin; Lipids; Measurement; Membrane; Membrane Lipids; Methods; Modeling; Mole the mammal; Molecular; Molecular Chaperones; Nuclear; Phosphatidylethanolamine; Phosphatidylserines; Phospholipids; Physical assessment; Physiological; Play; Property; Proteins; Race; Recording of previous events; Reporting; Research; Role; Sampling; Sphingomyelins; Structural Protein; Techniques; Testing; Work; age group; age related; alpha-Crystallins; aqueous; experience; fiber cell; fluidity; insight; lens; lens transparency; light scattering; membrane model; oxygen transport; physical property; prevent; sex; Affect; Affinity; Age; Animals; Binding; Biological; Cataract; Cell Nucleus; Cell membrane; Cholesterol; Crystalline Lens; Crystallins; Cytoplasm; Development; Electron Spin Resonance Spectroscopy; Eye; Goals; Health; Homeostasis; Human; Individual; Knowledge; Lead; Lecithin; Lipids; Measurement; Membrane; Membrane Lipids; Methods; Modeling; Mole the mammal; Molecular; Molecular Chaperones; Nuclear; Phosphatidylethanolamine; Phosphatidylserines; Phospholipids; Physical assessment; Physiological; Play; Property; Proteins; Race; Recording of previous events; Reporting; Research; Role; Sampling; Sphingomyelins; Structural Protein; Techniques; Testing; Work; age group; age related; alpha-Crystallins; aqueous; experience; fiber cell; fluidity; insight; lens; lens transparency; light scattering; membrane model; oxygen transport; physical property; prevent; sex

PROJECT SUMMARY/ABSTRACT The objective of this proposal is to understand the interaction of α-crystallin with membrane cholesterol (Chol) and cholesterol bilayer domains (CBDs) in the fiber-cell plasma membranes of the human eye lens. CBDs are formed in the fiber-cell plasma membrane of the eye lens and have positive physiological functions, helping to maintain lens transparency and possibly protect against cataract formation. The soluble lens protein, α-crystallin, is a major structural protein that, under healthy conditions, forms a transparent lattice in the lens and plays a major role in maintaining lens transparency. Several discoveries report that the level of α-crystallin in the lens cytoplasm declines with age and cataract progression, accompanied by a corresponding increase in the amount of membrane-bound α-crystallin. However, the mechanism by which α-crystallin associates with fiber-cell plasma membrane and how the age-related change in membrane lipid composition affects the α-crystallin binding is unclear. I hypothesize that the binding of α-crystallin to membrane is inhibited by CBDs, which decreases the light scattering and helps maintain lens transparency. In their proposed role, CBDs should increase the level of α-crystallin in the lens cytoplasm favoring its chaperone function and maintaining lens cytoplasm homeostasis. I discovered that the properties of CBDs change significantly with age and are related to the size of the CBD, which increases with age and is greater in nuclear than in cortical membranes. Based on my extensive experience working with CBDs in model and human lens membranes, I will (i) determine the lipid composition in fiber-cell plasma membranes that promotes or inhibits the binding of α-crystallin to membranes, (ii) test the hypothesis that CBDs inhibit the binding of α-crystallin to membranes, and finally (iii) determine the effects of CBD on the binding of α-crystallin in clear and cataractous human lens membranes of different age groups. The analysis will include donor's health history, sex, and race. I developed electron paramagnetic resonance (EPR) methods to study small-volume aqueous biological samples (3 µL at X-band or 150 nL at W-band), particularly for studies of lens membranes obtained from the eyes of a single donor. This technique provides a major advantage when studying the binding of α-crystallin in membranes of age-matched clear and cataractous lenses from human donors. In addition, the EPR approach has the unique ability to simultaneously provide information about the CBDs and the binding of α-crystallin. For the last eight years, my research has focused on understanding the molecular organization of lipids and proteins in plasma membrane of intact fiber cells of human eye lenses. Building upon the knowledge I acquired during these studies, here I propose moving my research in a new direction to focus on the interaction of CBDs with α-crystallin. There is a clear need for a more in-depth understanding of the roles of CBDs in the binding of α-crystallin in the fiber cell plasma membrane. The findings from this study will help fill this gap and produce valuable insights in maintaining lens transparency.

项目摘要/摘要 该提议的目的是了解α-晶状体与膜胆固醇（CHOL）的相互作用 人眼镜镜的纤维细胞质膜中的胆固醇双层域（CBD）。 CBD是 在眼镜的纤维细胞质膜中形成，具有正面的生理功能，有助于 保持透镜透明并可能防止白内障形成。固体透镜蛋白，α-晶状体， 是一种主要的结构蛋白，在健康条件下，在镜头中形成透明晶格并发挥 维持晶状体透明度的主要作用。几个发现报告说，镜头中的α-晶状体水平 细胞质随着年龄和白内障的进展而下降，伴随着相应的数量增加 膜结合的α-晶体蛋白。但是，α-晶状体蛋白与纤维细胞等离子体相关的机制 膜以及与年龄相关的膜脂质组成的变化如何影响α-晶体结合 不清楚。我假设CBD抑制了α-晶蛋白与膜的结合，这降低了 光散射并有助于维持晶状体透明度。在提议的角色中，CBD应提高 晶状体细胞质中的α-晶状体有利于其链酮功能并保持晶状体细胞质稳态。我 发现CBD的特性随着年龄的增长而发生显着变化，并且与CBD的大小有关 随着年龄的增长，核膜的增加，并且在核膜上更大。基于我的广泛 与CBD合作在模型和人透镜膜中工作的经验，我将（i）确定脂质组成 促进或抑制α-晶状体与膜结合的纤维细胞质膜，（ii）测试 CBD抑制α-晶状体与膜的结合的假设，最后（iii）确定了 CBD关于不同年龄组的清晰和白内障人透镜膜中α-晶状体的结合。这 分析将包括捐助者的健康历史，性别和种族。我开发了电子顺磁共振（EPR） 研究小体积水生物样品的方法（X波段3 µL或W频段时150 nL），尤其是 对于从单个供体的眼睛获得的镜头膜的研究。这项技术提供了专业 在研究年龄匹配的清晰和白内障镜头膜中α-晶状体蛋白的结合时的优势 来自人类捐助者。此外，EPR方法具有简单提供信息的独特能力 关于CBD和α-晶状体的结合。在过去的八年中，我的研究集中于 了解人类完整纤维细胞质膜中脂质和蛋白质的分子组织 眼镜。在这些研究中我获得的知识的基础上，我在这里提出了研究的研究 专注于CBD与α-晶蛋白相互作用的新方向。显然需要更深入 了解CBD在纤维细胞质膜中α-晶状蛋白结合中的作用。发现 从这项研究中，将有助于填补这一空白，并在维持晶状体透明度方面产生宝贵的见解。