Introduction 1 2 3 4 5 6 7 8 11 12 13 14 17 10 18 19 20 21 22 18 23 24 25 The results of the experiments presented in this paper demonstrate that celastrol is effective in preventing both the aggregation and toxicity of polyglutamine expression in cells and that it mediates these effects via the HSF1-mediated gene expression pathway. These results support the potential of this drug as a possible therapeutic agent for treating polyglutamine expansion diseases. The results also suggest that other drugs that stimulate HSF1 activity leading to hsp gene expression may also have beneficial activity against these disease states as well as other human diseases that are caused by protein misfolding. Materials and methods Plasmids, cell culture, and celastrol n Trypan blue cell viability assay 6 Fluorescence microscopy 1 Extract preparation and Western blot assay 2 26 β Filtration assay 2 27 SDS solubility assay β Statistical analysis t P Results Celastrol protects against polyglutamine toxicity 28 1 18 1 1 Fig. 1 a β upper panel lower panel b P P P  Celastrol protective effect are correlated with decreased number of cells containing polyglutamine aggregates 2 2 9 29 1 2 Fig. 2 a P P P b  Protective effects of celastrol treatment in PC12 cells 3 1 3 3 Fig. 3 a b c a b P P a P P b  HSF1−/− cells exhibit increased polyglutamine aggregation and toxicity 8 11 30 4 4 4 Fig. 4 a b c a b P a P b  Protective effects of celastrol require HSF1 18 1 2 3 5 Fig. 5 a b c b P P c  1 2 3 5 1 3 5 5 Discussion 31 21 22 32 34 8 11 19 20 35 8 11 19 20 32 34 18 21 22 36 5 Electronic supplementary material Below is the link to the electronic supplementary material.
 Figure S1 (A) (B) (C)