Fig. 4

The CHIP mutations cause tau aggregation. A After 48-h transfection, SH-SY5Y and BE2-M17 cells were paraformaldehyde-fixed and DAPI (blue) was used to stain the nuclear DNA. The images (× 630) were acquired using a Zeiss LSM780 laser scanning fluorescence confocal microscope. Scale bar, 10 μm. B, C Quantified results in A are shown as the percentage of tau aggregated foci in SH-SY5Y and BE2-M17 cells. D Ectopic expression of CHIP WT, p.Glu278fs, or Δ278–303 in A was examined using Western blot analysis. E CHIP mutants increase SDS-insoluble aggregation of tau in SH-SY5Y and BE2-M17 cells. Tau aggregation was detected by the filter-trap assay in cells transfected with the CHIP WT, p.Glu278fs, or Δ278–303 plasmid. The lysate was diluted in SDS and filtered through nitrocellulose membranes. Tau immunostaining was detected using the tau antibody. A representative image and the densitometry data are shown (a.u., arbitrary unit). The values of tau aggregation were normalized to the amount of aggregation in the empty vector control (one-way ANOVA, *p < 0.05, ***p < 0.001)