Figure 6

(a) Representative immunoblots of NHU whole-cell lysate showing GATA3, KRT13, FOXA1 and P63 protein expression after differentiation induction for 48 h following transfection with GATA3 siRNA. ACTB=β actin loading control. KRT13, FOXA1 and GATA3 were on the same membrane and normalised to the ACTB shown, and the P63 was on a separate membrane and normalised to a separate ACTB as shown in the supplementary data. (b) Densitometry measurements from immunoblots of three donors showing log₍₂₎ fold change in the expression of GATA3, KRT13 and FOXA1 in 48 h differentiation-induced NHU cells following transfection with GATA3 siRNA relative to control siRNA. Signals for P63 and KRT13 were normalised for loading to β-actin (ACTB) and fold change determined relative to the equivalent control siRNA transfection results. Statistical test performed was a repeated measures one-way ANOVA with Greenhous-Geisser correction and Sidak’s multiple comparison post test, with P-values indicated by *P≤0.05, **P≤0.01, ***P≤0.001 and ****P<0.0001. (c) RT-qPCR results combined from NHU cells from three independent donors showing change in abundance of RNA transcript for P63 and differentiation-associated genes after transfection with GATA3 siRNA followed by differentiation for 48 h. Log₍₂₎ fold change shown relative to control siRNA transfection with followed by 48 h differentiation. Statistics was performed using a two-way ANOVA with Dunnett’s multiple comparison post test, with P-values indicated by *P≤0.05, **P≤0.01, ***P≤0.001 and ****P<0.0001