Aggregation Characteristics of Tau Phosphorylated by Various Kinases as Observed by Quantum Dot Fluorescence Imaging

This study focused on the abnormal phosphorylation of tau and its aggregation process, characteristic of Alzheimer's disease, and aimed to compare the morphology and formation process of phosphorylated tau aggregates produced by four kinases: Cdk5/p25, GSK3<i>β</i>, MARK4, and p38<i>α</i>. Using quantum dots for 2D and 3D structural analysis, tau aggregates were confirmed in non-phosphorylated tau (non p-tau), as well as tau phosphorylated by GSK3<i>β</i> and MARK4. Aggregation initiation times were observed around 72 h for non-p-tau, and around 96 h for GSK3<i>β</i> and MARK4 phosphorylated tau. The thickness of non-p-tau aggregates was approximately 11 μm, while GSK3<i>β</i> aggregates were significantly thicker (13 μm) and exhibited increased density. TEM analysis suggested that tau forming wavy filaments was less prone to forming large aggregates. ThT assays and CD spectra showed an increased <i>β</i>-sheet structure for all kinases. Non-p-tau and GSK3<i>β</i> exhibited an increased right-twisted <i>β</i>-sheet structure, while Cdk5/p25, MARK4, and p38<i>α</i> showed an increased left-twisted <i>β</i>-sheet structure. The direct correlation between kinase activity and tau aggregate morphology revealed in this study provides a potential mechanistic basis for understanding disease heterogeneity and establishing novel therapeutic targets for AD specifically or for other neurodegenerative diseases as well.