TAp73 Inhibits EMT and Cell Migration in Pancreatic Cancer Cells through Promoting SMAD4 Expression and SMAD4-Dependent Inhibition of ERK Activation

Pancreatic ductal adenocarcinoma (PDAC) is a fatal disease due to early metastatic spread, late diagnosis and the lack of efficient therapies. The aim of our study was to reveal if transcriptionally active p73 (TAp73), a homolog of the well-known tumor suppressor p53, inhibits tumor progression through promoting canonical TGF-beta/Smad signaling and by preventing non-canonical extracellular signal-regulated kinases (ERK)1/2-mediated TGF- beta signaling. Using PDAC-derived tumor cell lines, we showed that TAp73 suppresses epithelial-mesenchymal transition by inducing the expression of epithelial markers while suppressing that of mesenchymal markers. We further demonstrated that TAp73 upregulates the expression of the TGF-beta signaling intermediate SMAD4 and that SMAD4 acts a mediator of TAp73-induced inhibition of ERK activation and cell motility. Measuring the levels of TAp73 and/or SMAD4 could help to predict whether TGF-beta preferentially uses an oncogenic or a tumor suppressive pathway in a given patient and at a specific time. Pancreatic ductal adenocarcinoma (PDAC) is a fatal disease due to early metastatic spread, late diagnosis and the lack of efficient therapies. A major driver of cancer progression and hurdle to successful treatment is transforming growth factor (TGF)-beta. Recent data from pancreatic cancer mouse models showed that transcriptionally active p73 (TAp73), a p53 family member, inhibits tumor progression through promoting tumor suppressive canonical TGF-beta/Smad signaling, while preventing non-canonical TGF-beta signaling through extracellular signal-regulated kinases (ERK)1/2. Here, we studied whether this mechanism also operates in human PDAC. Using the PDAC-derived tumor cell lines PANC-1, HPAFII and L3.6pl, we showed that TAp73 induces the expression of the epithelial marker and invasion suppressor E-cadherin and the common-mediator Smad, SMAD4, while at the same time suppressing expression of the EMT master regulator SNAIL and basal and TGF-beta 1-induced activation of ERK1 and ERK2. Using dominant-negative and RNA interferencebased inhibition of SMAD4 function, we went on to show that inhibition of ERK activation by TAp73 is mediated through SMAD4. Intriguingly, both SMAD4 and the ff isoform of TAp73-but not the beta isoform-interfered with cell migration, as shown by xCELLigence technology. Our findings highlighted the role of TAp73-SMAD4 signaling in tumor suppression of human PDAC and identified direct inhibition of basal and TGF- beta-stimulated pro-invasive ERK activation as an underlying mechanism.