Figure 2
From: Suppression of MAPK11 or HIPK3 reduces mutant Huntingtin levels in Huntington's disease models
HIPK3 and MAPK11 regulate HTT levels. (A) Left panels, representative western blots for the indicated proteins in STHdh cells upon transfection of the indicated siRNAs. Note that ab1 (N terminal antibody) detects both mHtt and wtHtt, and MW1 (polyQ antibody) detects mHtt only. Knocking down Hipk3 or Mapk11 reduces Htt levels in the HD (Q7/Q111) cells but not the WT (Q7/Q7) cells; Right panels, quantifications of the western blots. Hdh5 is an siRNA directly targeting the mouse Htt gene. (B) Left, A representative image showing the HD patient iPSC-derived neurons (blue, DAPI; green, Tuj1; scale bar, 50 μm); middle, mHTT levels of the HD patient iPSC-derived neurons measured by HTRF using the 2B7/MW1 antibody pair that detects mHTT specifically. Right, wtHTT levels of the control patient iPSC-derived neurons measured by HTRF using the 2B7/2166 antibody pair. (C) Representative western blots and quantifications showing that heterozygous deletion of Hipk3 reduces endogenous Htt levels in the striata of 10-month-old HD (HdhQ140/Q7) mice, but not WT (HdhQ7/Q7) mice. For quantifications, the Htt signal (measured by ab1) were first normalized to the loading control (β-Tubulin). The loading control corrected signals were then normalized to the average of WT controls at the same age. (D) Representative western blots and quantifications showing that heterozygous deletion of Mapk11 reduces endogenous Htt levels in the striata of 4- and 6.5-month-old HD (HdhQ140/Q7) mice, but not in WT (HdhQ7/Q7) mice. The quantification is the same as (C). For all bar plots in this figure, all signals were normalized to the average signal of the scrambled siRNA-transfected controls; the bars were plotted as mean and SEM; the number on top of each indicates the number of biological replicates, and the statistical analysis was performed by one-way ANOVA and post hoc Dunnett's tests for A and B, and two-way unpaired t tests for C and D, compared to the scramble siRNA-transfected control. *P < 0.05; **P < 0.01; ***P < 0.001; ns, P > 0.05.
