Journal
JOURNAL OF HEPATOLOGY
Volume 58, Issue 1, Pages 65-72Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jhep.2012.08.023
Keywords
TGF beta; HNF4 alpha; GSK3 beta; EMT; HCC; Gene therapy
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Funding
- Associazione Italiana per la Ricerca sul Cancro
- Ministero della Salute [35/07, 40H27]
- Ministero Universita e Ricerca Scientifica
- Tavola Valdese OPM
- Giorgio Ferraresi FIRC Fellowships for Cancer Research
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Background & Aims: The tumor fate derives from cell autonomous properties and niche microenvironmental cues. The transforming growth factor beta (TGF beta) is a major microenvironmental factor for hepatocellular carcinoma (HCC) influencing tumor dedifferentiation, induction of epithelial-to-mesenchymal transition (EMT) and acquisition of metastatic properties. The loss of the transcriptional factor HNF4 alpha is a predominant mechanism through which HCCs progress to a more aggressive phenotype; its re-expression, reducing tumor formation and repressing EMT program, has been suggested as a therapeutic tool for HCC gene therapy. We investigated the influence of TGF beta on the anti-EMT and tumor suppressor HNF4 alpha activity. Methods: Cell motility and invasion were analyzed by wound healing and invasion assays. EMT was evaluated by RT-qPCR and immunofluorescence. ChIP and EMSA assays were utilized for investigation of the HNR4 alpha DNA binding activity. HNF4 alpha post-translational modifications (PTMs) were assessed by 2-DE analysis. GSK3 beta activity was modulated by chemical inhibition and constitutive active mutant expression. Results: We demonstrated that the presence of TGF beta impairs the efficiency of HNF4 alpha as tumor suppressor. We found that TGF beta induces HNF4 alpha PTMs that correlate with the early loss of HNF4 alpha DNA binding activity on target gene promoters. Furthermore, we identified the GSK3 beta kinase as one of the TGF beta targets mediating HNF4 alpha functional inactivation: GSK3 beta chemical inhibition results in HNF4 alpha DNA binding impairment while a constitutively active GSK3 beta mutant impairs the TGF beta-induced inhibitory effect on HNF4 alpha tumor suppressor activity. Conclusions: Our data identify in the dominance of TGF beta a limit for the HNF4 alpha-mediated gene therapy of HCC. (C) 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.
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