Fig. 1

Optimized inhibitor PRG-A-04 reduces SOD1 aggregation. a PRG-A-04 suppressed SOD1 oligomerization by MT-SOD1 in a dose-dependent manner, but did not affect dimer formation. b, c PRG-A-04 suppressed SOD1 inclusions in SK-N-SH cells. Cells with SOD1 inclusions (white arrows; strong intensity of SOD1) were counted and the percentages are shown with standard deviation. n = 3 independent experiments; two-tailed Student’s t-test. Scale bar, 10 μm. d, e PRG-A-04 reduced misfolding of MT-SOD1. d The affinity of misfolding-specific antibody to MT-SOD1 was inhibited by PRG-A-04. e Quantification of band intensity with Image J software. Mean ± SD. A.U., arbitrary units. *P < 0.05, **P < 0.01. f PRG-A-04 reduced the MT-SOD1 recombinant protein-induced increase of self-oligomerization in vitro. g WT-SOD1 and TDP-43 oligomerization induced by ectopic TDP-43 expression was abolished by PRG-A-04 treatment. h–k WT-SOD1 aggregation induced by ectopic TDP-43 expression was reduced by PRG-A-04 treatment. PRG-A-04 reduced SOD1 as well as TDP-43 aggregation. h SK-N-SH cells were transfected with tdtomato-tagged TDP-43 and/or WT-SOD1 for 24 h. Scale bars, 10 μm. i PRG-A-04 reduced WT-SOD1 inclusions triggered by TDP-43 overexpression. Inclusion-positive cells (white arrows in h, strong intensity of SOD1) were counted and the percentages are shown as mean ± SD. j PRG-A-04 reduced the ratio of cells with cytoplasmic TDP-43 inclusions to tdtomato-positive ones. Cells with cytoplasmic TDP-43 (white arrows) among tdtomato positive ones were counted in randomly selected fields by photomicrographs and the cytosol/nucleus ratio was shown with ± SD. k PRG-A-04 reduced cytoplasmic TDP-43 inclusion. Mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.005