P069 Inhibition of IL-17 receptor signal transduction

Abstract Introduction IL-17A (IL-17) is a proinflammatory cytokine that mediates host defense against extracellular pathogens but also contributes to the pathogenesis of various autoimmune diseases, such as rheumatoid arthritis (RA), multiple sclerosis (MS) and psoriasis. Despite some homology with the toll-like receptors, IL-17 receptor subunits (IL-17RA and IL-17RC) do not recruit adaptors such as MyD88, but instead associate with an adaptor protein, Act1. Upon recruitment, Act1 activates Traf6, leading to a subsequent activation of a variety of transcription factors such as NF- κ B, C/EBP β and MAPKs. Despite advances in identifying molecular events regulating IL-17 signal transduction, the details of the inhibitory mechanisms of IL-17 signaling are still not well understood. Given the implication that unregulated IL-17 signaling results in enhanced autoimmune disease susceptibility, it is extremely important to understand the inhibitory mechanisms of the pathway. Recent studies have identified that post-translational modifications such as phosphorylation and ubiquitination are central to the regulation of IL-17 signal transduction. Ubiquitination is a stepwise process of linking ubiquitin chains on substrate proteins through any of the seven lysine residues on ubiquitin. Of the three enzymes, E1, E2 and E3 ubiquitin ligases, E3 ligases provide substrate specificity to the process. Interestingly, both Act1 and Traf6 have E3 ubiquitin ligase activity and lysine-63 linked ubiquitination of Traf6 is key in activating IL-17 pathway. Therefore, targeting ubiquitination as a mechanism for downregulating these pathways is a plausible inhibitory strategy. Genome analysis has identified over 100 deubiquitinases (DUBs) in humans but their role in IL-17 signaling is unknown. Mutations in the enzyme A20 (Tnfaip3) are associated with several autoimmune diseases including RA and psoriasis. Moreover, A20 inhibits a variety of proinflammatory signaling pathways such as TNF-alpha and IL-1, making it a compelling target. Methods Function of A20 in IL-17 signaling was tested by either siRNA-mediated knockdown of A20 or overexpressing A20 and measuring subsequent effect on IL-17 target gene expression by ELISA and quantitative PCR. Immunoprecipitation of A20 with different IL-17RA truncations was performed to map the association domain on IL-17RA. Results We demonstrate a functional role of A20 in the inhibition of IL-17 signaling. We observed an IL-17-dependent increase in A20 expression, and transient knockdown of A20 resulted in increased IL-17-dependent proinflammatory gene expression. Consistently, overexpression of A20 inhibited IL-17 target gene expression, which was associated with reduced NF- κ B activity. Interestingly, A20 associates with IL-17RA at its distal C-terminal domain, a motif that has been previously linked to negative regulation of the IL-17 signaling pathway. Conclusion We show for the first time that A20 can inhibit IL-17 signaling, which provides an intriguing mechanistic approach for inhibiting this pathway and could be a potential target for therapeutic intervention in IL-17-mediated autoimmune diseases.