Abstract

The endogenous metabolite itaconate has recently emerged as a regulator of macrophage function, but its precise mechanism of action remains poorly understood1,2,3. Here we show that itaconate is required for the activation of the anti-inflammatory transcription factor Nrf2 (also known as NFE2L2) by lipopolysaccharide in mouse and human macrophages. We find that itaconate directly modifies proteins via alkylation of cysteine residues. Itaconate alkylates cysteine residues 151, 257, 288, 273 and 297 on the protein KEAP1, enabling Nrf2 to increase the expression of downstream genes with anti-oxidant and anti-inflammatory capacities. The activation of Nrf2 is required for the anti-inflammatory action of itaconate. We describe the use of a new cell-permeable itaconate derivative, 4-octyl itaconate, which is protective against lipopolysaccharide-induced lethality in vivo and decreases cytokine production. We show that type I interferons boost the expression of Irg1 (also known as Acod1) and itaconate production. Furthermore, we find that itaconate production limits the type I interferon response, indicating a negative feedback loop that involves interferons and itaconate. Our findings demonstrate that itaconate is a crucial anti-inflammatory metabolite that acts via Nrf2 to limit inflammation and modulate type I interferons.

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Acknowledgements

We thank M. McMahon and J. D. Hayes for plasmids, and Cancer Research UK (C20953/A18644) and the BBSRC (BB/L01923X/1) for financial support for ATDK. This work was supported by a Wellcome Trust Investigator award to R.C.H. (110158/Z/15/Z), a grant to M.P.M. from the Medical Research Council UK (MC_U105663142), a Wellcome Trust Investigator award to MPM (110159/Z/15/Z), and a grant to E.R.S.K. and M.S.K. from the Medical Research Council UK (MC_U105663139). B.M.K. and R.F. are supported by the Kennedy Trust Fund. We acknowledge Metabolon for their assistance with the metabolic work and analysis. The O’Neill laboratory acknowledges the following grant support: European Research Council (ECFP7-ERC-MICROINNATE), Science Foundation Ireland Investigator Award (SFI 12/IA/1531), GlaxoSmithKline Visiting Scientist Programme and The Wellcome Trust (oneill-wellcometrust-metabolic, grant number 205455). E.T.C. is supported by the Claudia Adams Barr Program.

Author information

Author notes

    • Evanna L. Mills
    •  & Dylan G. Ryan

    These authors contributed equally to this work.

    • Michael P. Murphy
    •  & Luke A. O’Neill

    These authors jointly supervised this work.

Affiliations

  1. School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland

    • Evanna L. Mills
    • , Dylan G. Ryan
    • , Deepthi Menon
    • , Zbigniew Zaslona
    • , Marah C. Runtsch
    • , Anne F. McGettrick
    • , Mark M. Hughes
    • , Richard G. Carroll
    •  & Luke A. O’Neill
  2. Department of Cancer Biology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Evanna L. Mills
    • , Mark P. Jedrychowski
    •  & Edward T. Chouchani
  3. Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Evanna L. Mills
    • , Mark P. Jedrychowski
    • , John Szpyt
    •  & Edward T. Chouchani
  4. GlaxoSmithKline, Gunnelswood Road, Stevenage, Hertfordshire, UK

    • Evanna L. Mills
    • , Richard G. Carroll
    • , Lee M. Booty
    • , Louise K. Modis
    •  & Luke A. O’Neill
  5. MRC Mitochondrial Biology Unit, University of Cambridge, Cambridge CB2 0XY, UK

    • Hiran A. Prag
    • , Martin S. King
    • , Lee M. Booty
    • , Edmund R. S. Kunji
    •  & Michael P. Murphy
  6. Jacqui Wood Cancer Centre, Division of Cancer Research, School of Medicine, University of Dundee, Dundee DD1 9SY, UK

    • Dina Dikovskaya
    • , Maureen Higgins
    • , Elena V. Knatko
    •  & Albena T. Dinkova-Kostova
  7. MRC Cancer Unit, University of Cambridge, Hutchison/MRC Research Centre, Box 197, Cambridge Biomedical Campus, Cambridge CB2 0XZ, UK

    • Ana S. H. Costa
    •  & Christian Frezza
  8. School of Medicine, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland

    • Emily Hams
    •  & Padraic G. Fallon
  9. School of Chemistry, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland

    • Joanna F. McGouran
  10. Nuffield Department of Medicine, Target Discovery Institute, University of Oxford, Oxford OX3 7FZ, UK

    • Roman Fischer
    •  & Benedikt M. Kessler
  11. Division of Molecular and Clinical Medicine, School of Medicine, University of Dundee, Dundee DD1 9SY, UK

    • Paul J. Meakin
    •  & Michael L. J. Ashford
  12. GlaxoSmithKline, Park Road, Ware, Hertfordshire, UK

    • Gino Brunori
  13. Cellzome, GlaxoSmithKline R&D, Heidelberg, Germany

    • Daniel C. Sévin
  14. WestCHEM School of Chemistry, University of Glasgow, Glasgow G12 8QQ, UK

    • Stuart T. Caldwell
    •  & Richard C. Hartley
  15. Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA

    • Albena T. Dinkova-Kostova

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Contributions

E.L.M. and D.G.R. designed and performed experiments and analysed the data. E.L.M. wrote the manuscript with assistance from all other authors. D.M., M.M.H., M.C.R. and A.F.M. performed in vitro experiments using OI. R.G.C., D.C.S., A.S.H.C. and C.F. assisted with the metabolomics analysis. Z.Z., P.G.F. and E.H. assisted with the in vivo mouse LPS trials. S.T.C. and R.C.H. were responsible for the design and synthesis of octyl esters. H.A.P., E.R.S.K., M.S.K. and L.M.B. assessed the effect of OI and itaconate on mitochondrial parameters and itaconate transport. D.D., M.H. and A.T.D.-K. performed the NQO1 assay and KEAP1 wild-type and Cys151Ser mutant experiments. J.F.M., R.F., B.M.K., E.T.C., M.P.J. and J.S. assisted with mass spectrometry experiments. L.K.M. and G.B. provided guidance and advice. E.V.K., P.J.M. and M.L.J.A. assisted with experiments in Nrf2-deficient mice. L.A.O’N. conceived ideas and oversaw the research programme. M.P.M. provided advice, reagents and oversaw a portion of the work.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Luke A. O’Neill.

Reviewer Information Nature thanks N. S. Chandel, R. Rossignol, S. Werner and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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    This file contains Supplementary Figure 1, the uncropped scans with size marker indications, Supplementary Methods and Supplementary References.

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DOI

https://doi.org/10.1038/nature25986

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