Figure 2
Effect of PTC124 treatment on CHM3 fibroblasts. (A) qPCR analysis of CHM mRNA levels in PTC124-treated patient fibroblasts. CHM transcript levels of CHM3 fibroblasts treated with DMSO (grey bar) remained significantly lower than wild-type levels (black bar). Data are expressed as mean ± SEM, n = 3; Mann and Whitney test *p < 0.05. Regardless of the dose of PTC124 used, CHM transcript levels (dotted bars) are not altered, as compared to DMSO-treated cells; Kruskal-Wallis test p > 0.05. NT = non-treated. (B) A representative in vitro prenylation assay showing biotinylated Rab proteins in wild-type (WT), and in non-treated (NT), DMSO-treated or PTC124-treated CHM3 fibroblasts. β-actin serves as a loading control. Bands were cropped from the same gel. (C) Quantification of the biotinylated Rab pool (in relative units), after normalisation to β-actin loading, in multiple independent assays shows a significant 10-fold increase in biotinylated Rabs in the non-treated (NT) and DMSO-treated fibroblasts of patient CHM3 (grey bars), as compared to wild-type (WT; black bar). Data are expressed as mean ± SEM; n is indicated in the corresponding histogram bars; Mann and Whitney test *p < 0.05. Treatment with a range of PTC124 doses did not significantly alter biotinylated Rab levels in CHM3 fibroblasts (dotted bars) as compared to DMSO-treated controls (red dotted line); Kruskal-Wallis test p > 0.05.
