Pickhardt Marcus, Biernat Jacek, Hübschmann Sabrina, Dennissen Frank J A, Timm Thomas, Aho Annukka, Mandelkow Eva-Maria, Mandelkow Eckhard
DZNE, German Center for Neurodegenerative Diseases, Bonn, Germany; CAESAR Research Institute, Bonn, Germany.
DZNE, German Center for Neurodegenerative Diseases, Bonn, Germany; CAESAR Research Institute, Bonn, Germany.
Neurobiol Aging. 2017 Sep;57:47-63. doi: 10.1016/j.neurobiolaging.2017.04.022. Epub 2017 May 3.
The aggregation of Tau protein is a hallmark of neurodegenerative diseases including Alzheimer's disease. Previously, we generated a cell model of tauopathy based on the 4-repeat domain with the FTDP-17 mutation ΔK280 (Tau) which is expressed in a regulatable fashion (tet-on). The deletion variant ΔK280 is highly amyloidogenic and forms fibrous aggregates in neuroblastoma N2a cells staining with the reporter dye Thioflavin S. The aggregation of Tau is toxic, contrary to wildtype or anti-aggregant variants of the protein. Using a novel approach for monitoring in situ Tau aggregation and toxicity by combination of microscopic analysis with FACS and biochemical analysis of cells enabled the dissection of the aggregating species which cause a time-dependent increase of toxicity. The dominant initiating step is the dimerization of Tau which leads to further aggregation and induces a strong increase in reactive oxygen species (ROS) and cytoplasmic Ca which damage the membranes and cause cell death. Tau-based treatments using Tau aggregation inhibitors reduce both soluble oligomeric and fully aggregated Tau species and decrease their toxicity.
tau蛋白的聚集是包括阿尔茨海默病在内的神经退行性疾病的一个标志。此前，我们基于具有FTDP - 17突变ΔK280（tau）的4重复结构域构建了一个tau蛋白病细胞模型，该模型以可调控的方式（四环素诱导）表达。缺失变体ΔK280具有高度淀粉样变性，可在神经母细胞瘤N2a细胞中形成纤维状聚集体，并用报告染料硫黄素S进行染色。与该蛋白的野生型或抗聚集变体相反，tau蛋白的聚集具有毒性。通过将显微镜分析与流式细胞术相结合以及对细胞进行生化分析的新方法来监测原位tau蛋白聚集和毒性，能够剖析导致毒性随时间增加的聚集物种。主要的起始步骤是tau蛋白的二聚化，这会导致进一步聚集，并诱导活性氧（ROS）和细胞质钙大量增加，从而损害细胞膜并导致细胞死亡。使用tau蛋白聚集抑制剂的基于tau蛋白的治疗可减少可溶性寡聚体和完全聚集的tau蛋白物种，并降低其毒性。
