(A – C) C6wt, C6deltaPro and C6D23A were co-transfected into COS-7 cells with mHTT1–586 or the empty vector as control, and treated with inhibitor #12 (10 μM) in the media as indicated. Lysates were labelled with LE22 before analysis by Western blot using C6 antibody HD91. HTT was detected with antibody MAB2166. mHTT1–586 increases LE22 labelling of the C6 zymogen for C6wt, C6deltaPro and C6D23A, and this increase is prevented when cells are incubated in the presence of inhibitor #12. The compound furthermore reduces basal activity of the C6 zymogen in samples without mHTT1–586.
Graphed data are pooled results of three independent experiments graphed with S.E.M., Western blots are representative images. Statistical significance was determined by 2-way ANOVA, with Tukey’s post-hoc correction ), wt C6: HTT p = 0.0016, compound p <0.0001; deltaPro C6: HTT p = 0.0049, compound p <0.0001; D23A C6: HTT p = 0.0023, compound p <0.0001. *: p <0.05, **: p <0.01, ***: p <0.001, n.s.: not significant. See also Suppl. Fig. S6.