Yahya, K. | bioRxiv Science

bioRxiv2024

In situ generation of Aβ42 oligomers via secondary nucleation triggers neurite degeneration and synaptic dysfunction in human iPSC-derived glutamatergic neurons

The aggregation of A{beta}42 into misfolded oligomers is a central event in the pathogenesis of Alzheimers disease. In this study, we aimed to develop a robust experimental system that recapitulates A{beta}42 oligomerization in living cells to gain insight into their neurotoxicity and to provide a platform to characterize the effects of inhibitors of this process. Our strategy is based on the in situ generation of A{beta}42 oligomers via secondary nucleation by repeatedly treating the cells with A{beta}42 monomers in the presence of pre-formed A{beta}42 fibrils. This approach enables an accurate control over the levels of on-pathway soluble A{beta}42 oligomers and cell-associated aggregates, as well as the study of their neurotoxic effects. By implementing this approach in human glutamatergic neurons derived from induced pluripotent stem cells (iPSCs), we were able to replicate key aspects of Alzheimers disease, including neurite degeneration and synaptic dysfunction. Using BRICHOS, a molecular chaperone that specifically inhibits secondary nucleation, we confirmed that aggregation in this system occurs through secondary nucleation, and that quantitative parameters for comparing potential A{beta}42 aggregation inhibitors can be obtained. Overall, our results demonstrate that by in situ generation of on-pathway A{beta}42 oligomers, one can obtain translational cellular models of AD to bridge the gap between basic research and clinical applications.

cell biology↗

bioRxiv2023

Molecular mechanism of α-synuclein aggregation on lipid membranes revealed.

The central hallmark of Parkinsons disease pathology is the aggregation of the -synuclein protein, which, in its healthy form, is associated with lipid membranes. Purified monomeric -synuclein is relatively stable in vitro, but its aggregation can be triggered by the presence of lipid vesicles. Despite this central importance of lipids in the context of -synuclein aggregation, their mechanistic role in this process has not been established to date. Here, we use chemical kinetics to develop a detailed mechanistic model that is able to globally describe the aggregation behaviour of -synuclein in the presence of DMPS lipid vesicles, across a range of lipid and protein concentrations. Through the application of our kinetic model to experimental data, we find that the reaction is a co-aggregation process involving both protein and lipids and that lipids promote aggregation predominantly by enabling the elongation process. Moreover, we find that the initial formation of aggregates, via primary nucleation, takes place not on the surface of lipid vesicles but at the interfaces present in vitro. Our model will enable mechanistic insights, also in other lipid-protein co-aggregation systems, which will be crucial in the rational design of drugs that inhibit aggregate formation and act at the key points in the -synuclein aggregation cascade.

biophysics↗