Coactivation of AP-1 activity and TGF-β1 gene expression in the stress response of normal skin cells to ionizing radiation

Abstract

Activation of the AP-1 transcription factor and TGF-β1 growth factor by ionizing radiation was studied both in vivo in pig skin, and in vitro in human fibroblasts and keratinocytes. Three and 6 h after irradiation, the Fos and Jun proteins and their binding activity to an AP-1 consensus sequence were strongly induced by high doses of γ-rays. c-Fos, c-Jun and JunB proteins were found to be present in gel-shift complexes by probing with specific antibodies. Both keratinocytes and fibroblasts exhibited heightened AP-1 activity following irradiation. As we previously found that TGF-β1 is involved in the development of skin lesions induced by radiation, TGF-β1 gene expression was also examined. Two and 6 h after irradiation, the levels of TGF-β1 transcripts were increased in skin. By immunostaining, TGF-β1 protein levels were found to be increased in fibroblasts, keratinocytes and endothelial cells. As the TGF-β1 promoter contains AP-1 binding sites, the relation between AP-1 activity and TGF-β1 induction was addressed. The −365 TGF-β1 promoter fragment, which contains a high affinity AP-1 site, exhibited increased binding to Jun and Fos proteins following irradiation. These results suggest that stress-inducible TGF-β1 expression is mediated by the activation of AP-1 transcription factor.