Fig. 4

Induction of PTC readthrough by G418 and enhancers in hiPSC-derived R493X^−/− KI astrocytes. a R493X^−/− KI hiPSC-derived astrocytes were treated with vehicle solution, G418 alone, G418 in combination with CDX5–288, and rec. Human PGRN at the indicated concentrations for 72 h. Expression of PGRN and GRN-2,3 peptides in treated WT and R493X^−/− KI astrocyte samples were analyzed by western blotting, using actin as the loading control. b Densitometric quantification of ~ 70 kDa PGRN (i) and GRN-2,3 peptide (ii) in astrocyte lysates (a) normalized to vehicle-treated (VT) WT levels. VT WT was excluded from ii due to oversaturation of GRN-2,3 signal in long exposure blot. For clarity, rec. Human PGRN treated R493X^−/− KI astrocytes expressed 20.9% ± 0.027 of VT WT GRN-2,3 levels based on quantification of the short exposure blot (data not shown). c R493X^−/− KI hiPSC-derived astrocytes were treated with vehicle solution, G418 in combination with CDX5–288, and G418 CDX5–288 combination with either 10 or 30 μM of Z-Phe-Phe-FMK for 72 h. Again, expression of PGRN in WT and R493X^−/− KI astrocyte lysates was also analyzed by western blotting, using actin as the loading control. d Densitometric quantification of full-length PGRN in astrocyte lysates (c) normalized to VT WT levels. n = 3 independent cultures (except in dn = 2); values are shown as mean ± SEM; p < 0.05, ** p < 0.01, *** p < 0.0001 was determined by one-way ANOVA with Tukey’s multiple comparison test