Fig. 3.

BMP4-induced EMT is SMAD1 dependent. qRT-PCR of mRNA transcript levels for a SMAD1 and b SNAI1 and SNAI2 in OvCa429 and Panc1 cells transiently infected as described in methods with shRNA to SMAD1 or Scr controls and treated with 10 nM BMP4 for 24 h. Values are normalized to their respective untreated control (black horizontal line in b). c Images of shSMAD1- or control shScr–OvCa429 and Panc1 cells untreated or treated with BMP4 (10 nM), on Matrigel coated transwells after invasion for 18 h. Images are representative of two independent biological trials done in triplicate. Right graph represents the number of invading cells from four independent fields and two independent trials. d Immunofluorescence images of OvCa429, Panc1, and Py2T cells plated on soft (0.5 kPa) and rigid (8 kPa) fibronectin-coated hydrogels with or without 10 nM BMP4 treatment for 1 h followed by immunostaining with antiSMAD1. Overlay images are shown with the nuclear stain 4’6-diamidino-2-phenylindole. Scale bar = 20 μm. Quantitation of nuclear to cytoplasmic fluorescence intensity ratio is presented for each cell line (right graph). Error bars represent the standard error of the mean. *p < 0.05, **p < 0.01, ***p < 0.001