Abstract
Treatment of normal human epidermal keratinocytes (NHEK) with interferon-gamma (IFN-gamma) causes a 9-fold increase in the level of cyclooxygenase-2 (COX-2) mRNA expression. Nuclear run-off assays indicate that this induction is at least partly due to increased transcription. Activation of the epidermal growth factor receptor (EGFR) signaling pathway due to the enhanced transforming growth factor alpha (TGFalpha) expression plays an important role in the induction of COX-2 by IFN-gamma. This is supported by the ability of TGFalpha to rapidly induce COX-2 and the inhibition of the IFN-gamma-mediated COX-2 mRNA induction by an EGFR antibody and EGFR-selective kinase inhibitors. Deletion and mutation analysis indicates the importance of the proximal cAMP-response element/ATF site in the transcriptional control of this gene by TGFalpha. The increase in COX-2 mRNA by TGFalpha requires activation of both the extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) pathways. Inhibition of p38 MAPK decreases the stability of COX-2 mRNA, while inhibition of MAPK/ERK kinase (MEK) does not. These results suggest that the p38 MAPK signaling pathway controls COX-2 at the level of mRNA stability, while the ERK signaling pathway regulates COX-2 at the level of transcription. In contrast to NHEK, IFN-gamma and TGFalpha are not very effective in inducing TGFalpha or COX-2 expression in several squamous carcinoma cell lines, indicating alterations in both IFN-gamma and TGFalpha response pathways.
Highlights
Treatment of normal human epidermal keratinocytes (NHEK) with interferon-␥ (IFN-␥) causes a 9-fold increase in the level of cyclooxygenase-2 (COX-2) mRNA expression
To examine the effects of IFN-␥ on the expression of COX in NHEK, cells were treated with IFN-␥, and the levels of COX-1 and COX-2 protein and mRNA were analyzed by Western blot and Northern blot analysis, respectively
The p38 mitogen-activated protein kinase (MAPK) inhibitor PD169316 caused a dramatic decrease in the stability of COX-2 mRNA, whereas the MAPK/ERK kinase (MEK) inhibitor PD98059 exhibited in two independent experiments little effect on COX-2 mRNA stability in comparison with untreated cells. These results suggest that the activation of p38 MAPK regulates the stability of COX-2 mRNA, while activation of the extracellular signal-regulated kinase (ERK) signaling pathway appears important in the transcriptional control of COX-2
Summary
Vol 274, No 41, Issue of October 8, pp. 29138 –29148, 1999 Printed in U.S.A. Regulation of Cyclooxygenase-2 by Interferon ␥ and Transforming Growth Factor ␣ in Normal Human Epidermal Keratinocytes and Squamous Carcinoma Cells. Activation of the epidermal growth factor receptor (EGFR) signaling pathway due to the enhanced transforming growth factor ␣ (TGF␣) expression plays an important role in the induction of COX-2 by IFN-␥. Recent studies showed that targeted expression of IFN-␥ to the suprabasal layers of the epidermis of transgenic mice induces increased proliferation, a thickened epidermis, perturbed differentiation, and eczema resembling contact dermatitis [27] These results demonstrate the importance of IFN-␥ in the regulation of inflammation and cellular proliferation and differentiation in the skin. We provide evidence indicating that this induction is mediated at least in part through activation of the epidermal growth factor receptor (EGFR; c-ErbB1) and is related to increased expression of growth factors such as TGF␣ This induction of COX-2 is regulated in part at the transcriptional level and involves the CRE/ATF site in the proximal COX-2 promoter region. The stimulation of TGF␣ synthesis and possibly other cytokines by IFN-␥ and the subsequent increase in PGE2 production are likely to be important signals involved in triggering the hyperproliferative transformation associated with many inflammatory diseases in the skin
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