Fibril-forming motifs are essential and sufficient for the fibrillization of human Tau.

BACKGROUND: The misfolding of amyloidogenic proteins including human Tau protein, human prion protein, and human α-synuclein is involved in neurodegenerative diseases such as Alzheimer disease, prion disease, and Parkinson disease. Although a lot of research on such amyloidogenic proteins has been done, we do not know the determinants that drive these proteins to form fibrils and thereby induce neurodegenerative diseases. In this study, we want to know the role of fibril-forming motifs from such amyloidogenic proteins in the fibrillization of human Tau protein. METHODOLOGY/PRINCIPAL FINDINGS: As evidenced by thioflavin T binding and turbidity assays, transmission electron microscopy, and circular dichroism, fibril-forming motifs are essential and sufficient for the fibrillization of microtubule-associated protein Tau: only when both of its fibril-forming motifs, PHF6 and PHF6*, are deleted can recombinant human Tau fragment Tau(244-372) lose its ability to form fibrils, and the insertion of unrelated fibril-forming motifs from other amyloidogenic proteins, such as human prion protein, yeast prion protein, human α-synuclein, and human amyloid ß, into the disabled Tau protein can retrieve its ability to form fibrils. Furthermore, this retrieval is independent of the insertion location on Tau(244-372). CONCLUSIONS/SIGNIFICANCE: We demonstrate for the first time that insertion of fibril-forming motifs can replace PHF6/PHF6* motifs, driving human Tau protein to form fibrils with different morphologies and different kinetic parameters. Our results suggest that fibril-forming motifs play a key role in the fibrillization of human Tau protein and could be the determinants of amyloidogenic proteins tending to misfold, thereby causing the initiation and development of neurodegenerative diseases. Our study also touches on the importance of amyloid "strains": changes to the amyloidgenic driver region results in altered structural morphologies at the macromolecular level.

Low micromolar zinc accelerates the fibrillization of human tau via bridging of Cys-291 and Cys-322.

A hallmark of a group of neurodegenerative diseases such as Alzheimer disease is the formation of neurofibrillary tangles, which are principally composed of bundles of filaments formed by microtubule-associated protein Tau. Clarifying how natively unstructured Tau protein forms abnormal aggregates is of central importance for elucidating the etiology of these diseases. There is considerable evidence showing that zinc, as an essential element that is highly concentrated in brain, is linked to the development or progression of these diseases. Herein, by using recombinant human Tau fragment Tau(244-372) and its mutants, we have investigated the effect of zinc on the aggregation of Tau. Low micromolar concentrations of Zn(2+) dramatically accelerate fibril formation of wild-type Tau(244-372) under reducing conditions, compared with no Zn(2+). Higher concentrations of Zn(2+), however, induce wild-type Tau(244-372) to form granular aggregates in reducing conditions. Moreover, these non-fibrillar aggregates assemble into mature Tau filaments when Zn(2+) has been chelated by EDTA. Unlike wild-type Tau(244-372), low micromolar concentrations of Zn(2+) have no obvious effects on fibrillization kinetics of single mutants C291A and C322A and double mutant C291A/C322A under reducing conditions. The results from isothermal titration calorimetry show that one Zn(2+) binds to one Tau molecule via tetrahedral coordination to Cys-291 and Cys-322 as well as two histidines, with moderate, micromolar affinity. Our data demonstrate that low micromolar zinc accelerates the fibrillization of human Tau protein via bridging Cys-291 and Cys-322 in physiological reducing conditions, providing clues to understanding the relationship between zinc dyshomeostasis and the etiology of neurodegenerative diseases.