Induction of an epithelial to mesenchymal transition in human immortal and malignant keratinocytes by TGF-beta1 involves MAPK, Smad and AP-1 signalling pathways

J Cell Biochem. 2005 Aug 1;95(5):918-31. doi: 10.1002/jcb.20458.

Abstract

Recent data indicate that transforming growth factor-beta1 (TGF-beta1) can act to promote tumour progression in the late stages of carcinogenesis. The mechanism by which this occurs is unknown although a ligand-induced epithelial-mesenchymal transition (EMT) is thought to be important. In this study, we demonstrate that active Ras is required for TGF-beta1-induced EMT in human keratinocytes and that epidermal growth factor (EGF) can substitute for mutant Ras. EMT was reversed by the removal of TGF-beta1. Under conditions of TGF-beta1-induced EMT, cells were growth inhibited by the ligand resulting in G1 arrest. In cells containing normal Ras, TGF-beta1-activated ERK and p38 mitogen-activated protein kinases (MAPKs), and levels of activation were further increased by co-treatment with EGF. Inhibition of MAPK pathways and Smad2/3 signalling blocked the induction of EMT by TGF-beta1. Further, inhibition of the AP-1 transcriptional complex by [6]-Gingerol, or by the ectopic expression of JDP2, blocked TGF-beta1-induced EMT and conversely, stimulation of AP-1 by 12-O-tetradecanoylphorbol 13-acetate (TPA) substituted for EGF in the induction of EMT by TGF-beta1 in cells containing normal Ras. The presence of oncogenic Ras, the treatment of cells with EGF, or the treatment of cells with TPA to activate AP-1, potentiated TGF-beta1-induced Smad-dependent transcription, an effect that was attenuated by the inhibition of MAPKs and AP-1. The results demonstrate that active Ras and TGF-beta1 co-operate to reversibly induce EMT in human keratinocytes by mechanisms that involve MAPKs, Smad2/3 and AP-1. Further we demonstrate that MAPK/AP-1 signalling enhances Smad transcriptional activity under conditions associated with TGF-beta1-induced EMT.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Catechols
  • Cell Cycle
  • Cell Proliferation
  • Cell Transformation, Neoplastic
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Epithelial Cells / cytology*
  • Epithelial Cells / metabolism
  • Fatty Alcohols / pharmacology
  • Genes, ras / physiology
  • Ginger
  • Humans
  • Keratinocytes / cytology*
  • Luciferases / metabolism
  • Mesoderm / cytology*
  • Mesoderm / metabolism
  • Mutagens / pharmacology
  • Phosphorylation
  • Plasmids
  • Signal Transduction
  • Skin Neoplasms / metabolism
  • Skin Neoplasms / pathology
  • Smad3 Protein
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Transcription Factor AP-1 / genetics
  • Transcription Factor AP-1 / metabolism*
  • Transforming Growth Factor beta / pharmacology*
  • Transforming Growth Factor beta1
  • p38 Mitogen-Activated Protein Kinases / metabolism*

Substances

  • Catechols
  • DNA-Binding Proteins
  • Fatty Alcohols
  • Mutagens
  • SMAD3 protein, human
  • Smad3 Protein
  • TGFB1 protein, human
  • Trans-Activators
  • Transcription Factor AP-1
  • Transforming Growth Factor beta
  • Transforming Growth Factor beta1
  • gingerol
  • Luciferases
  • p38 Mitogen-Activated Protein Kinases