FIGURE 4.

NFAT expression is required for c-Myc induction by TGF-β. A, cancer cells were transfected with control siRNA or siRNA against NFAT proteins. Twenty-four hours post-transfection, cells were serum-starved and then treated with either medium alone or medium containing 10 ng/μl TGF-β. cDNA was prepared and subjected to qRT-PCR to analyze the effect of NFAT knockdown on c-Myc mRNA expression in Panc-1 (left) and PaTu8988t (right) cells. Values were calculated relative to basal mRNA expression levels in control siRNA-transfected cells, which were arbitrarily set to 1 for each experiment. Displayed are mean values from three independent experiments ± standard deviations. B, protein extracts from TGF-β-treated cancer cells transfected with either nonspecific control siRNA or NFAT-siRNA were subjected to immunoblotting with antibodies specific for NFATc1, NFATc2 or c-Myc. Protein loading was controlled using anti-β-actin antibodies.