Induction of gene expression via activator protein-1 in the ascorbate protection against UV-induced damage

UV irradiation is a major insult to the skin. We have shown previously that exogenous vitamin C (ascorbate) accumulates in HaCaT keratinocytes, thus c onferring the ability to prevent radical formation and cell death elicited by UV-B. Here, we have investigated the potential mechanisms accounting for the cytoprotective effects exerted by this antioxidant. Using a cDNA micro array hybridization, we identified several genes whose expression was up-re gulated by ascorbate. We focused on the fra-1 gene, a member of the Fos fam ily of transcription factors that down-regulates activator protein-1 (AP-1) target genes. Both in HaCaT and in normal human epidermal keratinocytes, w e found Fra-1 mRNA induction as early as 2 h after ascorbate loading. Elect rophoretic mobility-shift assay and antibody supershift analysis revealed t hat ascorbate modulates AP-I DNA-binding activity and that Fra-1 is in AP-1 complexes in treated cells. Furthermore, transient-transfection studies, u sing an AP-1 reporter construct, showed that ascorbate was able to inhibit both basal and UV-B-induced AP-l-dependent transcription. Ascorbate also mo dulates UV-B-induced AP-1 activity by preventing the phosphorylation and ac tivation of the upstream c-Jun N-terminal kinase (JNK), thus inhibiting pho sphorylation of the endogenous c-Jun protein. These data suggest that ascor bate mediates cellular responses aimed at counteracting UV-mediated cell da mage and cell death by interfering at multiple levels with the activity of the JNK/AP-1 pathway and modulating the expression of AP-1-regulated genes.