Cylindromatosis lysine 63 deubiquitinase (CYLD) suppress TLR3-mediated CCL5 expression in human renal proximal tubular epithelial cells.

backgroundOne of the causes of tubulointerstitial nephritis is viral infection, with innate immune responses affecting its pathogenesis. Toll-like receptor 3 (TLR3) recognizes viral infections and acts antivirally by activating signaling to produce inflammatory cytokines/chemokines, including C-C motif chemokine ligand 5 (CCL5) and interferon-β (IFN-β). Although cylindromatosis lysine 63 deubiquitinase (CYLD) is known to be associated with tubulointerstitial nephritis and renal function, its role in the antiviral innate immune response in tubular epithelial cells remains unknown. In this study, we investigated the association between CYLD and TLR3-mediated CCL5 production in cultured human renal proximal tubular epithelial cells (hRPTECs). METHODS AND

resultsPolyinosinic-polycytidylic acid (poly IC), a synthetic TLR3 ligand, was used to stimulate hRPTECs. mRNA expression was measured using reverse transcription-quantitative polymerase chain reaction. Protein expression was assayed using western blotting or an enzyme-linked immunosorbent assay. Knockdown of IFN-β, nuclear factor-kappa B (NF-κB) p65, and CYLD was performed by transfecting cells with specific small interfering RNAs. The intracellular localization of CYLD in hRPTECs was analyzed using immunofluorescence. Poly IC induced CCL5 expression in a time- and concentration-dependent manner, and knockdown of either IFN-β or p65 reduced poly IC-induced CCL5 expression. CYLD knockdown increased the poly IC-induced CCL5, phosphorylated IκB kinase α/β (IKK complex), and phosphorylated p65 expression. The CYLD protein was localized in the cytoplasm, and poly IC did not alter its expression.

conclusionCYLD may prevent excessive inflammation due to an antiviral innate immune response by suppressing IKK complex and NF-κB activation downstream of TLR3 in hRPTECs.