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Direct regulation of interleukin-6 expression by Notch signaling in macrophages

Cellular & Molecular Immunology volume 9, pages 155–162 (2012)Cite this article

Abstract

Interleukin-6 (IL-6) is a pleiotropic, pro-inflammatory cytokine produced by various types of cells, including macrophages. Within the IL-6 gene promoter region, the signature binding motif of CBF1/Su(H)/Lag-1 (CSL), a key DNA-binding protein in the Notch signaling pathway, was identified and found to overlap with a consensus nuclear factor (NF)-κB-binding site. Notch signaling is highly conserved and is involved in the regulation of biological functions in immune cells. In this study, we investigated the role of Notch signaling in the regulation of the IL-6 transcript in murine macrophages. The upregulation of Notch1 protein levels and the appearance of cleaved Notch1 (Val1744) correlated well with the increased IL-6 mRNA expression levels in murine primary bone marrow-derived macrophages (BMMφ) after activation by lipopolysaccharide (LPS) together with interferon-gamma (IFN-γ). Treatment of BMMφ with the γ-secretase inhibitor IL-CHO to suppress the transduction of Notch signaling resulted in a partial decrease in the level of IL-6 mRNA and the amount of IL-6 protein produced. In contrast, the overexpression of a constitutively activated intracellular Notch1 protein (NIC) in the RAW264.7 macrophage-like cell line resulted in significantly higher IL-6 transcript expression levels than in cells transfected with the empty vector control. The NF-κB inhibitor completely abrogated IL-6 mRNA expression induced by the overexpression of NIC. Chromatin immunoprecipitation (ChIP) using an anti-Notch1 antibody demonstrated that Notch1 is associated with the IL-6 promoter in RAW264.7 cells activated by LPS/IFN-γ but not in unstimulated cells. Taken together, these results strongly suggest that Notch1 positively regulates IL-6 expression via NF-κB in activated macrophages.

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Acknowledgements

This work was partly supported by the Thailand Research Fund (TRF) Grant No. RSA5280014 and by the Thai Government Stimulus Package 2 (TKK2555), under the Project for Establishment of Comprehensive Center for Innovative Food, Health Products and Agriculture. Financial supports from the TRF through the Royal Golden Jubilee Ph.D. Program (PHD/0337/2551) and Research Foundation Enhancement (Ratchadaphiseksomphot Endowment Fund to authors are also acknowledged.

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  1. Graduate Program in Biotechnology, Faculty of Science, Chulalongkorn University, Bangkok, Thailand

    Wipawee Wongchana & Tanapat Palaga

  2. Department of Microbiology, Faculty of Science, Chulalongkorn University, Bangkok, Thailand

    Tanapat Palaga

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  1. Wipawee Wongchana
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  2. Tanapat Palaga
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Correspondence to Tanapat Palaga.

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Wongchana, W., Palaga, T. Direct regulation of interleukin-6 expression by Notch signaling in macrophages. Cell Mol Immunol 9, 155–162 (2012). https://doi.org/10.1038/cmi.2011.36

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