Fig. 1

AGR2 expression is suppressed by TGF-β. a Four different cell lines A549, BT-474, MCF-7 and Panc1 were treated with TGF-β for 24 h. α-tubulin was used as a loading control and for normalization of protein levels calculated by densitometric analysis. Fold changes in protein level were calculated in relation to TGF-β untreated cells for each cell line independently. b mRNA levels of AGR2 were analyzed by RT-qPCR, GAPDH mRNA served as an endogenous control for data normalization. Data for qPCR are the mean +/− standard deviation obtained from three independent experiments. The decrease of AGR2 mRNA level in response to TGF-β in comparison with untreated cells was statistically significant (P < 0.05) for all analyzed cell lines. c Selected cell lines were treated with TGF-β and the cellular lysates were analyzed by Western blot with specific antibodies as indicated. β-actin was used as a loading control and for normalization to determine fold changes that were calculated in relation to TGF-β untreated cells for each cell line independently. d A549 and Panc1 cells were pre-treated with PD98059 for 2 h and then co-treated with TGF-β as indicated for 24 h. The changes in AGR2 level, phosphorylation of both p44/42 and Smad2 were analyzed by Western blot. α-tubulin was used as a loading control and for normalization of protein levels determined by densitometric analysis. Fold changes in protein level were calculated in relation to untreated cells for both cell lines independently. e A549 and Panc1 cells were pre-treated with SB431542 for 2 h and then co-treated with TGF-β as indicated for 24 h. The changes in AGR2 protein level were analyzed by Western blot. β-actin was used as a loading control and for normalization to calculate fold changes that were determined in relation to untreated cells for both cell lines independently