259 : Lipopolysaccharide decreases SIGIRR/TIR8 gene expression by suppressing Sp1 via TLR4-p38 mapkinase pathway in monocytes and neutrophils

Abstract

Single Immunoglobulin IL-1R-related molecule (SIGIRR), also known as Toll/IL-1R (TIR) 8, is one of the immunoglobulin (Ig)-like membrane proteins that are crucial for negative regulation of toll-like receptors (TLRs) such as TLR4 and the Ig domain-containing receptors such as IL-1R. Despite the importance of understanding the expression and function of the SIGIRR, knowledge is limited on the regulatory mechanism in the epithelial tissues, such as the liver, lung and gut, where its predominant expression is originally described. Here, we confirmed the expression of SIGIRR in non-epithelial innate immune cells, including neutrophilic-differentiated HL-60 (dHL60) cells and monocytic RAW264 cells, and the negative regulatory role of SIGIRR in LPS-mediated signaling. Consistent with previous findings in epithelial tissues, SIGIRR gene and protein expression were also down-regulated by LPS treatment in both a time- and a dose-dependent manner in dHL60 and RAW264 cells. A reduction was also observed in primary peripheral blood monocytes (MC) and polymorphonuclear neutrophils (PMN). Notably, exogenous introduction of dominant negative form of TLR4 and siRNA of p38 MAPK resulted in inhibition of LPS-induced SIGIRR down-regulation, while treatment with p38 activator anisomycin showed a dose-dependent decrease in SIGIRR expression, suggesting TLR4-p38 MAPK signal as a critical pathway for LPS-induced SIGIRR down-regulation. Finally, reporter gene and chromatin immunoprecipitation assay demonstrated that Sp1 is a key factor that directly binds to the proximal promoter of SIGIRR gene and, consequently, regulates basal SIGIRR expression, which is negatively regulated by LPS-dependent TLR4-p38 MAPK pathway. In summary, the data demonstrate precisely, for the fist time, how LPS down-regulates SIGIRR expression in non-epithelial innate immune cells, and provide a role of LPS signal that counteracts Sp1-dependent basal promoter activation of SIGIRR gene via TLR4-p38 MAPK pathway.