Fig. 4
From: Targeting RNA structure in SMN2 reverses spinal muscular atrophy molecular phenotypes
PK4C9 increases SMN protein in SMA cells. a Western blot quantification showing SMN levels in GM03814B vs. GM03813C fibroblasts (top) and in DMSO-treated (0.04%) vs. PK4C9-treated (40 μM) GM03813C fibroblasts (bottom) (n = 3 biological replicates). b Double-antibody staining showing the increase in SMN (red) in the cytoplasm and nucleus of GM03814B fibroblasts after treatment with PK4C9 (40 μM), and its restored localisation in nuclear gems (coilin-p80, white). Scale bar, 6 μm. c Quantification of the number of gems per cell (left) and the percentage of cells with gems (right) from images in the experiment (b) (n = 30 cells from three biological replicates). d qRT-PCR showing the effect of PK4C9 (40 μM) on the splicing of the indicated SMN-sensitive transcripts from GM03813C, GM03814B and wild-type (WT, ND36091A) fibroblasts. RPS6KB1, LPIN1, PHF14 and INSR responded to PK4C9 in GM03813C and GM03814B cells but not in ND36091A, confirming that these changes are mediated by SMN recovery. PRRC2C, BPTF, LGALS8 and CPSF6 responded to PK4C9 in SMA and WT cells, suggesting that they are PK4C9 off-targets (n = 8; four biological replicates and two technical replicates). **p < 0.01, ***p < 0.001 and ****p < 0.0001. p-values were obtained by applying non-paired, two-tailed t tests with Welch corrections. Graphs represent mean values ± SEM
