Fig. 3.
EMT is a prominent feature in Kras;N2ko PDAC. (A) Quantitative RT-PCR analysis of EMT-associated genes expressed by cancer cells from Kras and Kras;N2ko PDAC (n = 4 for each genotype). (B) Assessment of cell migration in wound closure assays performed in Kras and Kras;N2ko cells treated with TGF-β. Wound closure is delayed in Kras cells compared with Kras;N2ko cells. Quantification of wound closure is plotted as the percentage of the cell-free area over time. (C) Comparison of TGF-β gene sets by GSEA reveals significantly up-regulated TGF-β signatures in Kras;N2ko pancreata isolated from 7-d-old mice (dark blue, n = 2 and 4) and cancer cells (light blue, n = 6 each). A positive normalized enrichment score indicates elevated TFG-β–associated gene expression. Roman numbers refer to the detailed analysis in Tables S10 and S11. (D) Kras;N2ko cells reveal morphological and molecular responses characteristic of EMT in response to TGF-β, including loss of E-cadherin expression and nuclear translocation of SMAD4. (Scale bars: 50 μm.) (E) Treatment with the TGF-β receptor inhibitor SB431542 is sufficient to reverse the EMT-associated cadherin switch, suggesting that EMT in Kras;N2ko cells is dependent on a TGF-β autocrine loop.