Tau protein can form various fibrillar structures with different morphologies and molecular arrangements. These distinct polymorphs are linked to specific tauopathies or neurodegenerative diseases. Evaluation of the reaction mechanisms of these diverse aggregation pathways simultaneously requires monitoring of the single aggregates/fibrils. Here, we use total internal reflection fluorescence microscopy (TIRFM) to monitor, in real time, the growth of the aggregates of tau in the presence of polyU RNA. TIRFM imaging identifies the formation of a large population of novel nanoaggregates, in addition to the conventional amyloid fibrils. Kinetic analysis suggests that these aggregates are formed in a nucleation dependent manner with a critical nuclei size of ≈2, rate constants of nucleation of ≈2.8 × 10-2, and a very slow rate of growth (-1). Under the same conditions, the fibrils exhibit a slower rate of nucleation (≈1 × 10-4 M-1 s-1) but much faster elongation (≈3-4 μm h-1), giving rise to fewer but longer fibrils. Electron microscopy and circular dichroism spectroscopy reveal that the nanoaggregates are spherical in shape with a diameter of about 20 nm consisting of β-sheet proteins. Formation of these aggregates can be inhibited using NaCl, KCl, and NH4Cl, but the preformed aggregates are resistant to dissolution upon addition of salt, 1,6-hexanediol, or even 4 M GdnHCl, indicating high stability. Therefore, we hypothesize that the nanoaggregates are assembled via weak electrostatic interactions involving both tau and RNA but subsequently stabilized via strong H-bonding and hydrophobic interactions between the β-sheet monomers. In contrast, fibril growth remains largely unaffected by the presence of salts. We propose that the tau-polyanion nanoaggregates may form in intracellular environments and play important roles in seeding and proliferation of the tau fibrils in vivo.
Sahu et al. (Tue,) studied this question.