Differential regulation of RANTES and IL-8 expression in lung adenocarcinoma cells

Lung Cancer. 2007 May;56(2):167-74. doi: 10.1016/j.lungcan.2006.12.003. Epub 2007 Jan 17.


In lung adenocarcinoma, expression of Regulated upon Activation, Normal T cell Expressed and presumably Secreted (RANTES) is a predictor of survival while that of interleukin (IL)-8 is associated with a poor prognosis. In several models, tumorigenesis is abolished by RANTES, while it is facilitated by IL-8. We studied the regulation of RANTES and IL-8 expression in A549 lung adenocarcinoma cells. The effects of tumor necrosis factor (TNF)-alpha and regulators of protein kinases C (PKC)alpha/beta were tested because these have been shown to modulate cancer development and progression. TNF-alpha stimulated expression of both chemokines, while the PKCalpha/beta activator 12-O-tetradecanoyl-phorbol-13-acetate (TPA) induced only expression of IL-8 and inhibited TNF-alpha-induced RANTES expression. The PKCalpha/beta inhibitor Gö 6976 increased TNF-alpha-induced RANTES production and prevented its down-regulation by TPA. In contrast, it decreased TNF-alpha or TPA-induced IL-8 release. The differential regulation of RANTES and IL-8 expression was further analyzed. Site-directed mutagenesis indicated that regulation of RANTES promoter activity required two nuclear factor (NF)-kappaB response elements but not its activator protein (AP)-1 binding sites. An AP-1 and a NF-kappaB recognition sites were necessary for full induction of IL-8 promoter activity by TNF-alpha and TPA. Moreover, electrophoretic mobility shift assays demonstrated that NF-kappaB response elements from the RANTES promoter were of lower affinity than that from the IL-8 promoter. Immunoblotting experiments showed that TPA was more potent than TNF-alpha to induce in a PKCalpha/beta dependent manner the p44/p42 mitogen-activated protein kinases (MAPK) signaling cascade which controls AP-1 activity. Conversely, TPA inhibited TNF-alpha-induced NF-kappaB signaling and was a weak activator of this pathway. Thus, TPA did not sufficiently activate NF-kappaB to increase transcription through the low affinity NF-kappaB binding sites on RANTES promoter and its inhibitory effect on TNF-alpha-induced NF-kappaB signaling resulted in a reduced transcription rate. On IL-8 promoter, increased transcription through the high affinity NF-kappaB binding site occurred even with poorly activated NF-kappaB and the functional AP-1 response element compensated any loss of transcription rate. These data provide a mechanistic insight into the differential regulation of IL-8 and RANTES expression by PKCalpha/beta in lung adenocarcinoma cells.

MeSH terms

  • Adenocarcinoma / metabolism*
  • Blotting, Northern
  • Blotting, Western
  • Carcinogens / pharmacology
  • Cell Line, Tumor
  • Chemokine CCL5 / biosynthesis*
  • Electrophoretic Mobility Shift Assay
  • Humans
  • Interleukin-8 / biosynthesis*
  • Lung Neoplasms / metabolism*
  • Mitogen-Activated Protein Kinases / drug effects
  • Mitogen-Activated Protein Kinases / metabolism
  • NF-kappa B / drug effects
  • NF-kappa B / metabolism
  • Promoter Regions, Genetic
  • Protein Kinase C / drug effects
  • Protein Kinase C / metabolism
  • Signal Transduction / drug effects
  • Signal Transduction / physiology*
  • Tetradecanoylphorbol Acetate / pharmacology
  • Transcription Factor AP-1 / drug effects
  • Transcription Factor AP-1 / metabolism
  • Transfection
  • Tumor Necrosis Factor-alpha / drug effects
  • Tumor Necrosis Factor-alpha / metabolism


  • Carcinogens
  • Chemokine CCL5
  • Interleukin-8
  • NF-kappa B
  • Transcription Factor AP-1
  • Tumor Necrosis Factor-alpha
  • Protein Kinase C
  • Mitogen-Activated Protein Kinases
  • Tetradecanoylphorbol Acetate