Delineating protein-protein interaction network of hyperphosphorylated tau in tauopathies

阿尔茨海默病（AD）目前在美国侵袭超过500万患者。如果没有一个有效 治疗或预防方案，到2050年，该国的AD患者将接近1400万， 医疗费用高达1.2万亿美元。AD和其他疾病的主要病理标志 统称为tau蛋白病的神经退行性疾病是由以下组成的神经纤维缠结（NFT）： 过度磷酸化的tau蛋白（p-tau）。病理性tau蛋白过度磷酸化干扰了 tau蛋白的正常轴突运输相关功能。多种证据也揭示了功能获得 p-tau的细胞毒性。神经退行性变的p-tau蛋白分子靶点的鉴定将 可能为早期诊断工具和治疗方法的发展提供新的靶点。在这个项目中， 我们建议使用互补的方法来进行蛋白质的第一次系统鉴定， 优先与p-tau相互作用。在生物化学方法中，我们将使用重组p-tau蛋白， PIMAX-Cat技术从神经母细胞瘤细胞提取物中分离p-tau结合蛋白。遗传 方法，我们将使用拴系催化/酵母双杂交（TC/Y2 H）系统，该系统旨在 鉴定由包括磷酸化的翻译后修饰诱导的蛋白质-蛋白质相互作用， 从人脑cDNA文库中筛选p-tau结合蛋白。从这些发现的所有候选命中 将在基于神经细胞的共纯化实验中验证和表征两种方法。数据库 挖掘和数据整合将揭示以p-tau蛋白为中心的蛋白质-蛋白质相互作用网络， p-tau蛋白引起的神经变性的模型。

Alzheimer's disease (AD) currently attacks more than five million patients in the US. Without an effective treatment or preventative regime, AD patients in this country will be close to 14 million by 2050, with annual medical expenses topping 1.2 trillion US dollars. A major pathological hallmark for AD and other neurodegenerative disorders collectively called tauopathies is the neurofibrillary tangles (NFTs) composed of hyperphosphorylated tau protein (p-tau). Pathological hyperphosphorylation of tau interferes with the normal, axonal transport-related function of tau. Multiple lines of evidence also reveal a gain-of-function cytotoxicity of p-tau. The identification of the molecular targets of p-tau that underlie neurodegeneration will likely provide novel targets for the development of early diagnostic tools and therapeutics. In this project, we propose to use complementary approaches to perform the first systematic identification of proteins that interact preferentially with p-tau. In the biochemical approach, we will use recombinant p-tau produced by the PIMAX-Cat technology to isolate p-tau binding proteins from neuroblastoma cell extracts. In the genetic approach, we will use the tethered catalysis/yeast two-hybrid (TC/Y2H) system, which is designed to identify protein-protein interactions induced by a post-translational modification including phosphorylation, to screen for p-tau binding proteins from a human brain cDNA library. All candidate hits discovered from these two methods will be verified and characterized in neural cell-based co-purification experiments. Database mining and data integration will reveal the protein-protein interaction network centering upon p-tau, and lead to models for p-tau inflicted neurodegeneration.