Direct Regulation of Interleukin-6 Expression by Notch Signaling in Macrophages (172.36)

Abstract Notch signaling is required for immune cell development. The intracellular domain of Notch (NIC) induces expression of target genes by binding to CSL, a DNA-binding protein. In macrophages, Notch signaling regulates inflammatory cytokines TNFα and IL-6. IL-6 is secreted by various cell types, including activated macrophages and functions in immune regulation and inflammation. The consensus CSL binding site in the promoter region of IL-6 was shown to negatively regulate IL-6 mRNA, but the role of Notch signaling in IL-6 expression in macrophages remained unclear. We investigated the role of Notch signaling in the regulation of the IL-6 transcript in macrophages. Our results showed that bone marrow derived macrophages (BMMΦ) and RAW264.7 cell lines activated with lipopolysaccharide (LPS) and interferon gamma (IFNγ) induced IL-6 mRNA expression and upregulation of Notch1. Inhibition of Notch signaling in BMMΦ using a gamma secretase inhibitor partially decreases the level of IL-6 mRNA and protein. In contrast, RAW264.7, transiently transfected with constitutively active Notch1, displayed a dramatic increase in IL-6 mRNA. Inhibition of NF-κB completely abrogated IL-6 mRNA expression induced by the over expression of NIC. Chromatin immunoprecipitation assay demonstrated that Notch1 binds to the promoter of IL-6 in activated RAW264.7 but not in unstimulated cells. These results strongly suggest that Notch1 positively regulates IL-6 expression via NF-κB in activated macrophages.