Toxicity mechanisms of Aβ42 and Tau in aged cells

摘要 阿尔茨海默病(AD)的特征是神经元结构和功能的进行性丧失， 认知缺陷和痴呆症是其根本原因。神经元损伤与β的聚集有关 多肽和过度磷酸化的Tau蛋白。然而，减少这些神经毒性蛋白的临床结果 一直令人失望。这些失败表明阿尔茨海默病是一个多因素的过程，许多细胞缺陷 在移除Aβ/Tau证物后未被纠正。找到这些耐药的细胞缺陷是至关重要的 朝着AD的成功干预迈进一步。对人类的研究表明，衰老是最大的风险 AD的因素。然而，Aβ/Tau毒性的大多数细胞机制是通过对年轻细胞的研究而产生的。 由于衰老细胞具有不同于年轻细胞的蛋白质组和生理特性，因此研究Aβ42/Tau的细胞毒性 在老年细胞中是其对老年AD患者毒性的更好模型，并需要揭示耐药细胞 在老化过程中由Aβ42/Tau引起的缺陷。该提案的目标是确定Aβ/TAU、 与衰老一起，改变了细胞的区隔，以及是否使其中一些缺陷恢复活力 能保护细胞免受Aβ/Tau的毒性。确定这些年龄特定的Aβ/TAU敏感目标将揭示 β/Tau毒性的新机制和修复未纠正的细胞缺陷的策略 在去除Aβ/Tau沉积之后。我们建议使用酿酒酵母和果蝇 这两种模式生物被广泛用于研究分子 衰老与阿尔茨海默病的机制。酵母和苍蝇与人类有许多共同的生物过程，并会发育成年龄- 与人类衰老类似的相关细胞功能障碍。AD的酵母模型已经被用来揭示保守的 淀粉样蛋白聚集机制和A-β和Tau的细胞毒性及其遗传和化学作用 毒性修饰剂。然而，这些研究着眼于表达人类Aβ/Tau蛋白的年轻酵母细胞。 因此，我们希望结合酵母老化的强度和AD的研究来筛选蛋白质的缺陷 Aβ42/Tau在衰老细胞中的表达和定位我们的策略是使用陈年酵母来筛选 在AD的苍蝇模型中点击并测试保守的基因。我们打算通过决定如何实现我们的目标 Aβ和Tau蛋白干扰复制老化酵母细胞中的蛋白表达和定位，以及2) 评估新发现的长寿因子在AD果蝇模型中的有益作用。酵母菌筛选将是 通过使用我们实验室开发的一种新的高通量老化细胞显微筛选方法来完成。 从酵母筛选中获得的见解也将在AD的苍蝇模型中进行测试。这项工作将推动我们的 了解衰老和AD相关蛋白之间的相互作用，这些蛋白共同导致年龄相关的 神经退行性疾病。

ABSTRACT Alzheimer's disease (AD) is characterized by a progressive loss of neuronal structures and functions that underlie cognitive defects and dementia. The neuronal damage has been linked to the aggregation of Aβ peptides and hyperphosphorylated Tau proteins. However, clinical results of reducing these neurotoxic proteins have been disappointing. These failures suggest that AD is a multifactorial process and that many cellular defects were left uncorrected after removing the Aβ/Tau depositions. Finding these resistant cellular defects is a critical step towards the successful intervention of AD. Studies in humans have shown that aging is the biggest risk factor for AD. However, most cellular mechanisms of Aβ/Tau toxicity are generated by research in young cells. As aged cells have different proteomes and physiology than young cells, studying the cellular toxicity of Aβ42/Tau in aged cells is a better model for their toxicity in older AD patients and is required to reveal the resistant cellular defects that are induced by Aβ42/Tau during aging. The goal of this proposal is to determine how Aβ/Tau, together with aging, changes the cellular compartmentalization and whether rejuvenating some of these defects can protect cells from the toxicity of Aβ/Tau. Identifying these age-specific Aβ/Tau sensitive targets will reveal novel mechanisms of Aβ/Tau toxicity and enable strategies to rescue cellular defects that are left uncorrected after removing the Aβ/Tau depositions. We propose to use Saccharomyces cerevisiae (yeast) and Drosophila melanogaster (fly) for this pilot project as these two model organisms are widely used to study molecular mechanisms of aging and AD. Yeast and fly share many biological processes with humans and develop age- related cellular dysfunctions similar to human aging. The yeast model of AD has been used to reveal conserved mechanisms of amyloid aggregation and the cellular toxicity of Aβ and Tau, as well as the genetic and chemical toxicity modifiers. However, these studies looked at the young yeast cells expressing the human Aβ/Tau proteins. Therefore, we want to combine the strength of aging and AD research in yeast to screen for the defects of protein expression and localization caused by Aβ42/Tau in aged cells. Our strategy is to use aged yeast to screen for hits and test the conserved ones in fly models of AD. We intend to accomplish our goal by 1) determining how Aβ and Tau proteins interfere with protein expression and localization in replicatively old yeast cells, and 2) evaluating the beneficial effect of newly identified longevity factors in fly models of AD. The yeast screen will be accomplished by using a new high-throughput microscopic screening method developed in our lab for aged cells. The insights gained from the yeast screen will also be tested in the fly models of AD. This work will advance our understanding of the interactions between aging and AD-related proteins that together cause the age-related neurodegenerative diseases.