The formation of tangles of PHF-like and SF from in
vitro hyperphosphorylated τ. τ, 0.5 mg/ml, was incubated
with rat brain extract in the presence of ATP to induce
hyperphosphorylation of τ (a) or incubated with
nonhydrolyzable ATP, AMP–PNP, as a control (b). τ
hyperphosphorylation (≈12–15 mol of phosphate per mol of protein)
induced its self-polymerization into straight and PHF-like
(Inset) filaments (a), irrespective of
the τ isoform used for the phosphorylation assay. The magnification
bars in a and b represent 500 nm and, in
the Inset, 50 nm. Intertwining 4-nm filaments generate
small PHF-like structures (arrows) as follows: τ4 (c);
PHF, τ3L (d); τ protofilaments forming PHF, τ4S
(e); τ protofilaments forming bigger PHF, τ4S
(f); ≈15-nm straight filament, τ3
(g); mixture of all six τ isoforms hyperphosphorylated
(h), and Inset shows a PHF from the
tangle formed; and control sample incubated without ATP, τ4S
(i). Bars represent 40 nm in c–g and
i, 200 nm in h, and 25 nm in
Inset.