FIGURE 10.

AGR2 induces AREG expression through YAP1. A, immunoblotting with isoform-specific antibodies for phosphorylated and total YAP1 protein in H460 cells. The H460 cells shown include controls (WT), AGR2 KD, and AGR2-KD_AREG cells where AREG is overexpressed after transfection. B, qPCR of YAP1 after expression of YAP1-specific shRNAmir in H460, OE33, and JH-EsoAd1 cells. Before qPCR was performed, the cells were FACS sorted for high GFP expression. OE33_AGR2 represent cells transfected with AGR2 cDNA. C, qPCR of AREG using the same cells as in panel B. D, ELISA for AREG protein in the tissue culture media of OE33 and JH-EsoAd1 cells. H460, OE33, and JH-EsoAd1 cells were FACS-sorted for GFP expression 72 h after RNAi exposure followed by another 48 h in culture before the media was collected for the AREG ELISA assay. E, protein immunoblotting of OE33 cell lysates for phosphorylated and total YAP1, and β-actin with and without AGR2 overexpression. F, plots derived from densitometry of the protein immunoblots (E) quantified with ImageJ.