Characterization of TNF-α– and IL-17A–Mediated Synergistic Induction of DEFB4 Gene Expression in Human Keratinocytes through IκBζ

Abstract

Human β-defensin 2 (hBD2), encoded by the DEFB4 gene, is an antimicrobial peptide playing an essential role in inflammatory processes in the skin. hBD2 expression is regulated synergistically by tumor necrosis factor-α (TNF-α) and IL-17A; however, the underlying regulatory mechanisms are unknown. The purpose of this study was to characterize the molecular mechanism by which TNF-α and IL-17A synergistically induce hBD2 expression. In cultured human keratinocytes we show that a constitutive noninducible binding of the transcription factor organic cation transporter 1 (OCT1) to the DEFB4 promoter is crucial for IL-17A/TNF-α-mediated synergistic induction of hBD2 but not the synergistic induction of CCL20, IL8, IL17C and LCN2. Interestingly, stimulation with IL-17A results in a p38 mitogen-activated protein kinase-dependent accumulation of inhibitor of nuclear factor κB ζ (IκBζ), which is a necessity for synergistic induction of hBD2. Finally, co-stimulation with TNF-α induces DNA binding of NF-κB and activator protein 1 (AP-1) to two specific sites in the DEFB4 promoter region. Hence, our study shows how two inflammatory stimuli are integrated by three different signaling pathways into the regulation of one specific target gene involving the three specific transcription factors OCT1, NF-κB, and AP-1 as well as the transcriptional cofactor IκBζ. These findings may be important in psoriasis, where TNF-α and IL-17A have been identified as key pathogenic cytokines.