title: TNF-alpha production in the skin.
abstract: Upregulation of TNF-alpha is a key early response to ultraviolet B (UVB) by keratinocytes (KCs), and represents an important component of the inflammatory cascade in skin. UVB irradiation induces TNF-alpha expression in both KCs and dermal fibroblasts, with TNF-alpha mRNA induction seen as early as 1.5 h after UVB. We previously reported that the effects are wavelength-specific: TNF-alpha expression and secretion are induced by UVB (290-320 nm), but not by UVA (320-400 nm). Moreover, we found that IL-1alpha, a cytokine also present in irradiated skin, substantially and synergistically enhances the induction of TNF-alpha by UVB, and the induction of TNF-alpha by this combination of UVB with IL-1alpha is mediated through increased TNF-alpha gene transcription. We investigated the molecular mechanism for UVB-induction of the TNF-alpha gene with a series of TNF-alpha promoter constructs, ranging from 1.2 kbp (from -1179 to +1 with respect to the TNF-alpha transcription initiation site) down to 0.1 kbp (-109 to +1), each driving expression of a CAT reporter. Our results showed a persistent nine to tenfold increase of CAT activity in all TNF-alpha promoter/reporter constructs in response to UVB (30 mJ/cm(2)) exposure. These results indicate the presence of UVB-responsive cis-element(s) located between -109 and +1 of the TNF-alpha promoter, a region that contains a putative AP-1 site and a putative NFkB site. UVB-induction was abolished when the TNF-alpha promoter was mutated by one base pair at the AP-1 binding site. Cells treated with SP600125, an AP-1 inhibitor that inhibits JNK (c-Jun N-terminal kinase), also showed suppression of the 0.1 kbp TNF-alpha promoter/reporter construct. The authentic endogenous gene in untransfected cells was also blocked by the inhibitor. Electrophoretic Mobility Shift Assay indicated new complexes from UVB-treated nuclear extracts and anti-phospho-c-Jun, a regulatory component of the AP-1 transcription factor, creating a supershift indicating increased phosphorylation of c-Jun and hence higher AP-1 activity. Keratinocyte-derived TNF-alpha is a component of the early induction phase of the inflammatory cascade.
authors.affiliation: Department of Dermatology, University of Pennsylvania, 2, Rhodes Pavilion, 3600 Spruce Street, Philadelphia, PA, 19104, USA.
authors.forename: M M
authors.initials: MM
authors.lastname: Bashir
authors.forename: M R
authors.initials: MR
authors.lastname: Sharma
authors.forename: V P
authors.initials: VP
authors.lastname: Werth
mesh_terms: D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D001665:Binding Sites / Q000235:genetics / Q000276:immunology; D005347:Fibroblasts / Q000276:immunology / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects; D005786:Gene Expression Regulation / Q000235:genetics / Q000276:immunology* / Q000528:radiation effects; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D015603:Keratinocytes / Q000276:immunology* / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects; D016297:Mutagenesis, Site-Directed; D015398:Signal Transduction / Q000276:immunology; D012867:Skin / Q000276:immunology* / Q000378:metabolism / Q000473:pathology / Q000528:radiation effects; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D014466:Ultraviolet Rays
chemical_list: D000873:Anthracenes; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; C432165:pyrazolanthrone; D048031:JNK Mitogen-Activated Protein Kinases