Regulation of phorbol ester-mediated TRAF1 induction in human colon cancer cells through a PKC/RAF/ERK/NF-kappaB-dependent pathway

Oncogene. 2004 Mar 11;23(10):1885-95. doi: 10.1038/sj.onc.1207312.


Tumor necrosis factor (TNF) receptor-associated factors (TRAFs) are cytoplasmic adapter proteins that link a wide variety of cell surface receptors to the apoptotic signaling cascade. The purpose of this study was to delineate the signaling pathways and TRAF1 promoter elements responsible for phorbol ester-mediated TRAF1 induction in human colon cancers. Here, we found that the PKC activators, phorbol 12-myristate 13-acetate (PMA) and bryostatin I, induced TRAF1 mRNA expression; pretreatment with actinomycin D blocked PMA-mediated TRAF1 expression suggesting induction at the transcriptional level. In contrast, expression of other TRAFs (TRAF2, 3 and 4) was minimally altered by PMA. Various PKC isoform-selective inhibitors blocked PMA-mediated TRAF1 mRNA and promoter stimulation; rottlerin, a selective PKCdelta inhibitor, had no effect suggesting that Ca(2+)-dependent PKC isoforms (e.g., PKCalpha and betaI) play a role in TRAF1 regulation. In addition, the MEK/ERK inhibitors, PD98059 and UO126, suppressed PMA-stimulated TRAF1 promoter activity indicating a role for ERK in TRAF1 induction. Moreover, cotransfection of a dominant-negative Raf-1 (Raf-C4) significantly reduced PMA-stimulated TRAF1 promoter activity whereas transfection of dominant-negative Ras or treatment with Ras inhibitors had minimal to no effect on TRAF1 induction suggesting dependence on Raf, but not Ras, activation. Finally, site-specific mutagenesis of functional NF-kappaB sites (particularly the most proximal site) in the TRAF1 promoter significantly decreased PMA-mediated promoter activity. In conclusion, our results demonstrate selective induction of TRAF1 in human colon cancer cells through a Ca(2+)-dependent PKC/Raf-1/ERK/NF-kappaB-dependent pathway.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Cell Line, Tumor
  • Colonic Neoplasms
  • Dactinomycin / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Flavonoids / pharmacology
  • Gene Expression Regulation, Neoplastic / drug effects*
  • Humans
  • Mitogen-Activated Protein Kinases / metabolism
  • NF-kappa B / metabolism
  • Promoter Regions, Genetic
  • Protein Kinase C / metabolism
  • Proteins / drug effects
  • Proteins / genetics*
  • Proto-Oncogene Proteins c-raf / metabolism
  • TNF Receptor-Associated Factor 1
  • Tetradecanoylphorbol Acetate / pharmacology*
  • Transcription, Genetic / drug effects


  • Enzyme Inhibitors
  • Flavonoids
  • NF-kappa B
  • Proteins
  • TNF Receptor-Associated Factor 1
  • Dactinomycin
  • Proto-Oncogene Proteins c-raf
  • Protein Kinase C
  • Mitogen-Activated Protein Kinases
  • Tetradecanoylphorbol Acetate
  • 2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one