Tau protein fibrils are aggregated polymers of pathological forms of tau proteins present in the brains of Alzheimer’s disease (AD) patients and patients suffering from other types of neurodegenerative tauopathies. Tau pathology spreads between neurons and propagates misfolding in a prion-like manner throughout the brain, leading to significant neuronal death. Despite the advancements in understanding tau pathology, the relationships between initial tau misfolding, the formation of fibrils, pathology propagation across connected neurons, and subsequent cytotoxicity on the level of individual neurons remain unclear.
Recently cryo-EM showed disease-specific structures of tau fibrils isolated from patients' brains depending on the type of tauopathy [1, 2]. Moreover, heparin-induced recombinant tau fibrils, frequently used as model filaments for AD research, formed different types of filaments [3]. These findings show that tau protein can adopt structurally distinct conformations depending not only on the tau protein variant (isoform, phosphorylation, truncation) but also on buffer conditions (ionic inducers, pH).
This project addresses the question of tau fibril structure on two levels; in-vitro and in-situ. We try to understand what tau modifications and external conditions cause tau protein fibrilization, what kind of structure these fibrils adopt, and which conditions lead to the formation of AD-specific fibril topology. We also investigate the ultrastructural aspects of the intake and seeding of various tau fibril variants directly inside neurons.
In this poster, we present an ongoing systematic screening of full-length 2N4N tau isoform fibrilization conditions (type of buffer, temperature, presence or absence of agitation, and additives) monitored by Thioflavin T assay, negative stain EM, AFM, and cryo-EM. We also show preliminary data from in-situ cryo-ET of AD-tau fibril intake by neurons.
This work has received funding from European Union Excellence Hub (101087124) and Czech Science Foundation (22-15175I). CIISB research infrastructure project LM208127 funded by MEYS CR is gratefully acknowledged for the financial support of the measurements at the CEITEC Cryo-Electron Microscopy and Tomography Core Facility, CEITEC Proteomics Core Facility, and CEITEC Nanobiotechnology Core Facility.