Cellular Dysfunction in Exfoliation Glaucoma

项目摘要/摘要：剥脱性青光眼综合征的细胞功能障碍 (XFS)是一种与年龄相关的疾病，涉及聚集的纤维物质(XFM)在 细胞外基质。它的主要发病是在眼睛，在那里XFM形成在表面 前段组织。XFM导致剥脱性青光眼(XFG)，这是一种进展迅速的疾病 与全球约三分之一的开角型青光眼(POAG)病例有关。XFG 表现出强烈的年龄依赖性，与许多符合条件的年龄相关疾病相似 作为聚集性疾病。LOXL1，一种基质交联酶，催化 原弹性蛋白用于合成弹性纤维，是XFM的主要成分，已与XFG联系在一起 然而，通过基因组学广泛关联研究，仍然不清楚LOXL1蛋白是如何 有助于疾病病理。了解XFS/G的细胞基础的进展 由于缺乏实验模型而变得缓慢。与原代人TENON成纤维细胞(TF)合作 来源于XFG患者和对照组的小梁切除术(年龄匹配的POAG患者和非POAG患者)。 青光眼患者)，我们最近发现，XFG-TF显示了许多功能特征 在其他年龄相关聚集性疾病的细胞中观察到，如帕金森氏症、阿尔茨海默氏症和 AMD包括溶酶体定位、自噬、微管组织和功能缺陷， 以及线粒体的健康状况。将LOXL1与XFG的开发联系起来，我们发现 Xfg-tf a)显示通过旨在中和错误折叠的机制增加LOXL1出口 新生多肽；b)通过自噬来处理LOXL1，表明存在 这种蛋白质的错误折叠或变性单位；c)在细胞内积累蛋白质聚集体 更高的内源性表达；以及d)过表达聚集反应伴侣中的簇- 像蛋白质一样。最后，对LOXL1二级结构的电子检测表明，LOXL1 蛋白质在N-末端具有基本结构，预计这些结构会增加 聚合倾向。此外，对LOXL1全蛋白或缺失的实验表明 确实，LOXL1具有很高的聚集倾向，并且这种倾向的位置位于 在N端的特定伸展范围内。因此，我们试图从根本上回答 关于支持XFG病理的因素的问题。我们建议，SpA1)来研究分子 导致微管功能缺陷的机制；SpA2)以研究细胞应激 响应，以及LOXL1输出和错误折叠的蛋白质机制之间的联系，以及SpA3) 鉴定LOXL1序列的最小氨基酸序列和结构特征 调节其聚集倾向。

Project Summary/Abstract: Cellular Dysfunction in Exfoliation Glaucoma Exfoliation syndrome (XFS) is an age-related disease involving the deposition of aggregated fibrillar material (XFM) in extracellular matrices. Its main morbidity is in the eye, where XFM forms on the surface of anterior segment tissues. XFM causes exfoliation glaucoma (XFG), a rapidly progressing disease associated with approximately 1/3 of open-angle glaucoma (POAG) cases worldwide. XFG demonstrates a sharp age-dependence in similarity to the many age-related diseases qualified as aggregopathies. LOXL1, a matrix cross-linking enzyme that catalyzes the crosslinking of tropoelastin for the synthesis of elastic fibers and a major component of XFM, has been linked to XFG by Genomics Wide Association Studies, however, it is still unclear how LOXL1 protein contributes to disease pathology.Progress in understanding the cellular basis for XFS/G has been slowed by a lack of experimental models. Working with primary human tenon fibroblasts (TF) derived from trabeculectomies of XFG patients and controls (age-matched POAG patients and No- Glaucoma patients), we recently found that, XFG-TFs display many of the functional features observed in cells from other age-related aggregopathies such as Parkinson's, Alzheimer's, and AMD including defects in lysosomal positioning, autophagy, microtubule organization and function, and mitochondrial health status. Linking LOXL1 to the development of XFG, we have found that XFG-TF a) display increased LOXL1 export by a mechanism aimed at neutralizing misfolded nascent polypeptides; b) process LOXL1 through autophagy indicating the presence of misfolded or denatured units of this protein; c) accumulate intracellular protein aggregates with higher endogenous expression; and d) overexpress clusterin an aggregate-responsive chaperone- like protein. Finally, an in silico examination of LOXL1 secondary structure indicated that LOXL1 protein has elemental structures within the N-terminus that are predicted to confer an increased aggregation propensity. Furthermore, experiments of LOXL1 whole protein or with deletions shows that indeed LOXL1 has high aggregation propensity and that the loci of this propensity reside within specific stretches of the N-terminus. Accordingly, we seek to answer fundamental questions on the factors that underpin XFG pathology. We propose, SpA1) to study the molecular machinery leading to the defects in microtubule function; SpA2) to investigate cellular stress responses, and the connection between LOXL1 export and misfolded protein machinery, and SpA3) to identify the minimal amino acid sequences and structural features of the LOXL1 sequence that mediate its aggregation propensity.