The concept that cellular terminal differentiation is stably maintained once development is complete has been questioned by numerous observations showing that differentiated epithelium may undergo an epithelial-to-mesenchymal transition (EMT) program. EMT and the reverse process, mesenchymal-to-epithelial transition (MET), are typical events of development, tissue repair, and tumor progression. In this study, we aimed to clarify the molecular mechanisms underlying these phenotypic conversions in hepatocytes. Hepatocyte nuclear factor 4a (HNF4a) was overexpressed in different hepatocyte cell lines and the resulting gene expression profile was determined by real-time quantitative polymerase chain reaction. HNF4a recruitment on promoters of both mesenchymal and EMT regulator genes was determined by way of electrophoretic mobility shift assay and chromatin immunoprecipitation. The effect of HNF4a depletion was assessed in silenced cells and in the context of the whole liver of HNF4 knockout animals. Our results identified key EMT regulators and mesenchymal genes as new targets of HNF4a. HNF4a, in cooperation with its target HNF1a, directly inhibits transcription of the EMT master regulatory genes Snail, Slug, and HMGA2 and of several mesenchymal markers. HNF4a-mediated repression of EMT genes induces MET in hepatomas, and its silencing triggers the mesenchymal program in differentiated hepatocytes both in cell culture and in the whole liver. Conclusion: The pivotal role of HNF4a in the induction and maintenance of hepatocyte differentiation should also be ascribed to its capacity to continuously repress the mesenchymal program; thus, both HNF4a activator and repressor functions are necessary for the identity of hepatocytes. (HEPATOLOGY 2011;53:2063-2074 E pithelial-to-mesenchymal transition (EMT) is the process by which polarized cells of the epithelium lose cell-cell connections and acquire the mesenchymal characteristics of motility and invasiveness. The reverse process, mesenchymal-to-epithelial transition (MET), often occurs at a site secondary to the original EMT population. The dynamic EMT/MET processes are essential for embryonic development and wound repair and initiate the pathological states of fibrosis and metastatic cancer.

The notorious unresponsiveness of metastatic cutaneous melanoma to current treatment strategies coupled with its increasing incidence constitutes a serious worldwide clinical problem. Moreover, despite recent advances in targeted therapies for patients with BRAF V600E mutant melanomas, acquired resistance remains a limiting factor and hence emphasises the acute need for comprehensive pre-clinical studies to increase the biological understanding of such tumours in order to develop novel effective and longlasting therapeutic strategies. Autophagy and ER stress both have a role in melanoma development/progression and chemoresistance although their real impact is still unclear. Here, we show that BRAF V600E induces a chronic ER stress status directly increasing basal cell autophagy. BRAF V600E -mediated p38 activation stimulates both the IRE1/ASK1/JNK and TRB3 pathways. Bcl-XL/Bcl-2 phosphorylation by active JNK releases Beclin1 whereas TRB3 inhibits the Akt/mTor axes, together resulting in an increase in basal autophagy. Furthermore, we demonstrate chemical chaperones relieve the BRAF V600E -mediated chronic ER stress status, consequently reducing basal autophagic activity and increasing the sensitivity of melanoma cells to apoptosis. Taken together, these results suggest enhanced basal autophagy, typically observed in BRAF V600E melanomas, is a consequence of a chronic ER stress status, which ultimately results in the chemoresistance of such tumours. Targeted therapies that attenuate ER stress may therefore represent a novel and more effective therapeutic strategy for BRAF mutant melanoma.

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