Abstract
Assembly of tau protein into paired helical filaments and straight filaments is a key feature of Alzheimer's disease. Aggregation of tau has been implicated in neurodegeneration, cellular toxicity and the propagation, which accompanies disease progression. We have reported previously that a region of tau (297–391), referred to as dGAE, assembles spontaneously in physiological conditions to form paired helical filament-like fibres in vitro in the absence of additives such as heparin. This provides a valuable tool with which to explore the effects of tau in cell culture. Here we have studied the cellular uptake of soluble oligomeric and fibrillar forms of dGAE and examined the downstream consequences of tau internalisation into differentiated SH-SY5Y neuroblastoma cells using fluorescence and electron microscopy alongside structural and biochemical analyses. The assembled dGAE shows more acute cytotoxicity than the soluble, non-aggregated form. Conversely, the soluble form is much more readily internalised and, once within the cell, is able to associate with endogenous tau resulting in increased phosphorylation and aggregation of endogenous tau, which accumulates in lysosomal/endosomal compartments. It appears that soluble oligomeric forms are able to propagate tau pathology without being acutely toxic. The model system we have developed now permits the molecular mechanisms of propagation of tau pathology to be studied in vitro in a more physiological manner with a view to development of novel therapeutic approaches.
Highlights
A common pathological process amongst neurodegenerative diseases is the accumulation of amyloid aggregates formed by disease-specific proteins in the cytoplasm, nucleus or extracellular space [1]
The Alexafluor-tag® labels amino groups at the N terminus and on lysine residues, and we examined the tagged and untagged dGAE using transmission electron microscopy (TEM), SDS-PAGE and Circular dichroism (CD) spectroscopy to determine whether the label affects aggregation and/or filament formation
After fibrillisation induced by agitation for 72 h, there was less of the 12-kDa monomeric form of dGAE compared with dGAE-488, and more of both preparations were retained in the gel well [41]
Summary
A common pathological process amongst neurodegenerative diseases is the accumulation of amyloid aggregates formed by disease-specific proteins in the cytoplasm, nucleus or extracellular space [1]. The misfolding, self-assembly and accumulation of tau protein in neurofibrillary tangles is a major pathological feature shared by tauopathies, the most common of which is Alzheimer's disease (AD). The identity of the species of tau that is most toxic has been debated [2,3,4,5]. It was thought that neurofibrillary tangles are an integral feature of tau toxicity, since their number and distribution in the brain correlate with cognitive decline in AD [6,7,8]. As with other amyloidogenic proteins implicated in creativecommons.org/licenses/by/4.0/)
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