Figure 2.

Hsp70 associates with tau aggregates. (A) Tau was aggregated in the presence of varying concentrations of Hsp70 for 5 h. After centrifugation, the resulting supernatant (S) and pellet (P) fractions were analyzed by immunoblotting using anti-Hsp70 and anti-tau antibodies. In the absence of AA (−AA), both Hsp70 and hTau40 remain in the supernatant fractions. When AA is added (+AA), Hsp70 cosediments with tau in pellet fractions. Electron micrographs demonstrate immunogold labeling of Hsp70 on amorphous tau oligomers. The ratio at the bottom left is the Hsp70:tau ratio. (B) Hsp70 was added to preformed oligomeric and filamentous tau aggregates. Electron micrographs demonstrate abundant Hsp70 immunogold labeling of oligomeric structures (arrowheads) but very limited labeling of tau filaments (arrow). (C) Quantification of Hsp70 immunogold labeling. Hsp70 was 3 times more likely to label ≤100 nm structures than >100 nm structures. *p < 0.01 (Student's t test). (D) Double immunogold labeling of Hsp70 (arrow) and the His tag on tau. Several amorphous aggregates were labeled for both. Also shown is a tau filament that is labeled for Hsp70. Hsp70 is labeled with 6 nm gold particles. His-tagged hTau40 is labeled with 10 nm gold particles. The scale bars are 50 nm.