Fig. 3: Maintenance of PrPC homeostasis is important for myoblast differentiation.
a Western blot for PrPC and the myogenic differentiation markers MyHC and MyoG during C2C12 mouse myoblasts (control), C2C12 myoblasts stably expressing full-length wild-type mouse PrPC (WT PrPC), C2C12 myoblasts stably expressing F198S PrPC, and C2C12 myoblasts KO for PrPC cultured until their confluence reached 90% and then incubated with differentiation medium for 0, 3, and 5 days, respectively. β-actin served as the protein loading control. The relative amount of PrPC (b), MyHC (c), or MyoG (d) in the above cell lines (control, orange; WT PrPC, blue; F198S PrPC, gray; and KO for PrPC, pink) (solid black circles shown in scatter plots) was expressed as the mean ± SD (with error bars) of values obtained in n = 3 independent experiments. One-way two-sided ANOVA and multiple comparisons with no adjustments were performed by SPSS 19.0, and different letters indicate significant differences at the level of p < 0.05. Immunofluorescence imaging of the above four cell lines incubated with differentiation medium for 3 and 5 days, respectively, using antibody against PrPC (red) (e) or MyHC (red) (f) and staining with DAPI (blue). Scale bars, 50 (e) and 75 μm (f), respectively. We have replaced the second row for Day 3 in (f) with a correct version of WT PrPC, in which WT PrPC-expressing cells did not have any MyHC signal at day 3 in most fields of view. Source data are provided as a Source Data file.
