Molecular Mechanism of αAN101D-Transgene-Induced Age-Related Cataract

αAN101D转基因诱发年龄相关性白内障的分子机制

• 批准号: 10597653
• 负责人: Om Prakash Srivastava
• 金额: $ 39.48万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10597653
• 关键词: Actins; Affect; Aging; Asparagine; Aspartic Acid; Binding; CDC42 gene; Cataract; Crystallins; Cytoskeleton; DNA Sequence Alteration; Defect; Deposition; Development; Disease; Epithelial Cells; Epithelium; Exhibits; Goals; Human; Hydrophobicity; In Vitro; K ATPase; Kinetics; Lens Opacities; Lens development; Link; Literature; Mass Spectrum Analysis; Membrane; Modeling; Modification; Molecular; Molecular Chaperones; Mus; Na(+)-K(+)-Exchanging ATPase; Pathogenicity; Phenotype; Play; Post-Translational Protein Processing; Precipitation; Predisposition; Property; Proteins; Resolution; Role; Structural Protein; Structure; Testing; Therapeutic; Tight Junctions; Transgenes; Transgenic Mice; Water; Wild Type Mouse; age related; aged; cell motility; crosslink; deamidation; fiber cell; in vivo; insight; lens; migration; mouse model; protein protein interaction; rho; rho GTP-Binding Proteins

The proposal has the overall goal to determine the molecular mechanisms of an age-related cortical cataract development. Cataracts are principally a lens structural proteins' (crystallins) aggregation and subsequent precipitation disease that are inducible by genetic mutations, and occurs due to aging. During cataract development, the increased sizes of aggregated and cross-linked crystallin multimers become so large that they finally become water insoluble and cause lens opacity. Several post-translational modifications (PTMs) of crystallins are known to cause age-related cataracts, and the deamidation of crystallins (the most abundant among post-translational modifications) is considered as a major cataract-causative factor. In spite of voluminous literature on in vitro studies of effects of crystallins' deamidation leading to cataract-development, no clear molecular mechanism has emerged that could implicate the deamidation-induced link of the in vitro effects to in vivo changes. Therefore, the PI's-generated unique αA-N101D mouse model, where asparagine 101 is deamidated to aspartic acid, provides an opportunity to directly link the in vitro studies to in vivo changes in phenotypic and crystallins' properties to the cortical cataract development. The model would provide information about the molecular mechanism of age-related cataract development. Based on our extensive results, we have hypothesized that the cataract in αAN101D mouse model is caused by the altered crystallin properties, cellular defects and synergistically increased deposit of αAN101D with membrane, resulting in membrane disruption and ionic imbalance. We plan to test the above hypothesis by seeking answers the following three questions using the above mouse model: (A) Aim 1: Does deamidated αAN101D causes temporal alterations in crystallin- crystallin interactions leading to aggregation among crystallins and their insolubilization in vivo? (B) Aim 2: What are the temporal sequence of lens phenotypes that cause cellular defects during the cortical cataract development? (C) Aim 3: Does temporal increase in binding of deamidated αAN101D to membrane leads to membrane disorganization and intracellular ionic imbalance? The results will be of significant therapeutic value to delay the development and progression of age-related cataracts.

该提案的总体目标是确定年龄相关性皮质性白内障的分子机制 发展白内障主要是透镜结构蛋白（晶体蛋白）聚集和随后的 沉淀性疾病是由基因突变诱导的，并且由于衰老而发生。在白内障 随着晶体蛋白多聚体的发育，聚集和交联的晶体蛋白多聚体的尺寸增加， 最终变成水不溶性并导致透镜不透明。几种翻译后修饰（PTM） 已知晶体蛋白引起年龄相关性白内障，并且晶体蛋白的脱酰胺化（最丰富的 在翻译后修饰中）被认为是白内障的主要致病因素。尽管有大量的 关于晶体蛋白脱酰胺作用导致白内障发展的体外研究的文献， 已经出现的分子机制可能涉及脱酰胺诱导的体外效应与胰岛素抵抗之间的联系。 体内变化。因此，PI生成的独特α A-N101 D小鼠模型，其中天冬酰胺101是 脱酰胺为天冬氨酸，提供了将体外研究与体内变化直接联系起来的机会， 表型和晶体蛋白的性质对皮质性白内障的发展。该模型将提供信息 关于年龄相关性白内障发展的分子机制。根据我们的广泛结果，我们有 假设α AN 101 D小鼠模型中的白内障是由晶状体蛋白性质的改变、细胞凋亡和白内障形成引起的。 缺陷并协同增加α AN 101 D与膜的存款，导致膜破裂， 离子失衡我们计划通过使用以下三个问题寻求答案来测试上述假设 上述小鼠模型：（A）目的1：脱酰胺α AN 101 D是否引起晶状体蛋白- 晶体蛋白相互作用导致晶体蛋白之间的聚集和它们在体内的不溶解？（B） 目的2：在皮质骨发育过程中，导致细胞缺陷的透镜表型的时间顺序是什么？ 白内障发展？(C)目的3：脱酰胺α AN 101 D与 膜导致膜解体和细胞内离子失衡？结果将是 对延缓年龄相关性白内障的发展和进展具有显著的治疗价值。