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Anti-inflammatory activity of IgG1 mediated by Fc galactosylation and association of FcγRIIB and dectin-1

Nature Medicine volume 18, pages 1401–1406 (2012)Cite this article

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Abstract

Complement is an ancient danger-sensing system that contributes to host defense, immune surveillance and homeostasis1. C5a and its G protein–coupled receptor mediate many of the proinflammatory properties of complement2. Despite the key role of C5a in allergic asthma3, autoimmune arthritis4, sepsis5 and cancer6, knowledge about its regulation is limited. Here we demonstrate that IgG1 immune complexes (ICs), the inhibitory IgG receptor FcγRIIB and the C-type lectin–like receptor dectin-1 suppress C5a receptor (C5aR) functions. IgG1 ICs promote the association of FcγRIIB with dectin-1, resulting in phosphorylation of Src homology 2 domain–containing inositol phosphatase (SHIP) downstream of FcγRIIB and spleen tyrosine kinase downstream of dectin-1. This pathway blocks C5aR-mediated ERK1/2 phosphorylation, C5a effector functions in vitro and C5a-dependent inflammatory responses in vivo, including peritonitis and skin blisters in experimental epidermolysis bullosa acquisita. Notably, high galactosylation of IgG N-glycans is crucial for this inhibitory property of IgG1 ICs, as it promotes the association between FcγRIIB and dectin-1. Thus, galactosylated IgG1 and FcγRIIB exert anti-inflammatory properties beyond their impact on activating FcγRs.

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Figure 1: IgG1-ICs inhibit C5a-mediated inflammatory responses in vivo and in vitro by an FcγRIIB-dependent mechanism.
Figure 2: The inhibitory effect of IgG1-ICs on neutrophil migration depends on dectin-1.
Figure 3: IgG1-ICs activate Src kinase and promote phosphorylation of tyrosine and Syk downstream of dectin-1 and of SHIP downstream of FcγRIIB.
Figure 4: High Fc glycan galactosylation is crucial for the inhibitory effect of IgG1-ICs in vivo and promotes the association of FcγRIIB and dectin-1.

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Acknowledgements

This work is supported by the German Research Foundation (Deutsche Forschungsgemeinschaft, GRK1727; project 8 and SFB/TR22; project A21) to J.K., EXC306/1 to D.Z., R.L. and J.K., and by Deutsche Forschungsgemeinschaft EH221-5 to M.E. P.R.T. is a Medical Research Council (UK) Senior Fellow (G0601617). G.D.B. was supported by the Wellcome Trust. We thank T. Köhli for technical assistance with the surface plasmon resonance analysis, T. Gutsmann for help with the fluorescence spectroscopy measurements, T. Peters and L. Wollin for helpful discussions and B. Heyman (Uppsala University, Sweden) for providing the H5 and the 7B4 hybridoma clones.

Author information

Author notes

  1. Christian M Karsten and Manoj K Pandey: These authors contributed equally to this work.

Authors and Affiliations

  1. Institute for Systemic Inflammation Research, University of Lübeck, Lübeck, Germany

    Christian M Karsten, Julia Figge, Regina Kilchenstein, Gabriele Köhl, Marc Ehlers & Jörg Köhl

  2. Division of Cellular and Molecular Immunology, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA

    Manoj K Pandey, Nathaniel L Harris, Fred D Finkelman & Jörg Köhl

  3. Institute of Infection and Immunity, Cardiff University School of Medicine, Heath Park, Cardiff, UK

    Philip R Taylor, Marcela Rosas, Jacqueline U McDonald & Selinda J Orr

  4. Glycodesign and Glycoanalytics, Central Institute of Laboratory Medicine and Pathobiochemistry, Charité Medical University, Berlin, Germany

    Markus Berger, Dominique Petzold & Veroniqué Blanchard

  5. Laboratory of Tolerance and Autoimmunity, German Rheumatism Research Center, Berlin, Germany

    André Winkler, Constanze Hess & Marc Ehlers

  6. Section of Infection and Immunity, University of Aberdeen, Aberdeen, UK

    Delyth M Reid & Gordon D Brown

  7. Institute of Biology, Center for Structural and Cell Biology in Medicine, University of Lübeck, Lübeck, Germany

    Irina V Majoul

  8. Division of Emergency Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA

    Richard T Strait

  9. Department of Respiratory Diseases Research, Boehringer Ingelheim Pharma GmbH, Biberach, Germany

    Eva Wex

  10. Department of Dermatology, University of Lübeck, Lübeck, Germany

    Ralf Ludwig & Detlef Zillikens

  11. Department of Biology, Chair of Genetics, University of Erlangen-Nuremberg, Erlangen, Germany

    Falk Nimmerjahn

  12. Division of Immunology, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA

    Fred D Finkelman

  13. Department of Medicine, Cincinnati Veterans Affairs Medical Center Cincinnati, Ohio, USA

    Fred D Finkelman

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  1. Christian M Karsten
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Contributions

C.M.K. and M.K.P. conducted key studies and analyzed the data. J.F. assessed antigen binding of in vitro–glycosylated IgGs and performed phenotypical characterization of bone marrow cells, and some in vivo studies. R.K. and C.M.K. performed the EBA studies. R.L. and D.Z. provided the rabbit collagen type VII–specific IgG for the EBA studies and helped with EBA-related data analysis. P.R.T., M.R. and J.U.M. provided the dectin-1–transduced neutrophil and macrophage cell lines, and helped with assays using such lines. S.J.O. performed the peptide pull-down experiments. M.B., D.P. and V.B. did the MALDI-TOF analysis. A.W. and C.H. performed the in vitro galactosylation of the H5 antibody. G.D.B. and D.M.R. provided the Clec7a−/− mice and helped with the neutrophil assays using such mice and with ex vivo assays with primary Clec7a−/− cells. I.V.M. provided material and advice for the FRET experiments. R.T.S. and F.D.F. provided TNP-OVA and helped with initial IC studies. N.L.H. and G.K. performed the ERK phosphorylation studies with neutrophils. E.W. provided the conditional Syk-knockout mice and the protocol for in vitro Syk depletion. F.N. provided the recombinant IgG1 7B4 antibody and recombinant mouse FcγRIIB-Fc and assessed HiGalH5-IC and H5-IC binding to FcγRIIB-transfected Chinese hamster ovary cells. M.E. provided scientific input and coordinated the glycan analysis. J.K. designed and coordinated the study, analyzed the data and wrote the manuscript.

Corresponding author

Correspondence to Jörg Köhl.

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Competing interests

E.W. is an employee of Boehringer Ingelheim Pharma.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–23 and Supplementary Table 1 (PDF 4835 kb)

Supplementary Movie 1

Expression of FcγRIIB but absence of Syk phosphorylation in untreated BM neutrophils. This movie shows the expression pattern of FcγRIIB (green, anti-FcγRIIB-FITC Ab) and of phosphorylated Syk (p-Syk, magenta, anti–p-Syk–Alexa568 Ab) in a BM-derived neutrophil 3 min after incubation with PBS, as determined by confocal microscopy in an animated three-dimensional projection. Cell stains positive for FcγRIIB. No phosphorylation of Syk is visible in the absence of 107.3-IC treatment. Similar results were obtained for p-SHIP. (MOV 1966 kb)

Supplementary Movie 2

Expression of dectin-1 but absence of Syk phosphorylation in untreated BM neutrophils. This movie shows the expression pattern of dectin-1 (magenta, anti–dectin-1–APC Ab) and phosphorylated Syk (p-Syk, green, anti–-p-Syk–Alexa568 Ab) in a BM-derived neutrophil 3 min after incubation with PBS, as determined by confocal microscopy in an animated three-dimensional projection. No phosphorylation of Syk occurs in the absence of 107.3-IC treatment. Similar results were obtained for p-SHIP. (MOV 1340 kb)

Supplementary Movie 3

Phosphorylation of Syk and co-localization of p-Syk with FcγRIIB in response to 107.3-IC treatment. This movie shows the co-localization pattern of FcγRIIB (green, anti-FcγRIIB-FITC Ab) and of phosphorylated Syk (p-Syk, magenta, anti-p-Syk-Alexa568 Ab) in a BM-derived neutrophil 3 min following 107.3-IC treatment, as determined by confocal microscopy in an animated three-dimensional projection. 107.3-IC treatment drives phosphorylation of Syk; p-Syk co-localizes with FcγRIIB as indicated by the white spots. Similar results were obtained for p-SHIP. (MOV 2750 kb)

Supplementary Movie 4

Phosphorylation of Syk and co-localization of p-Syk with dectin-1 in response to 107.3-IC treatment. This movie shows the co-localization pattern of dectin-1 (magenta, anti–dectin-1–APC Ab) and phosphorylated Syk (p-Syk, green, anti-p-Syk-Alexa568 Ab) in a BM-derived neutrophil 3 min following 107.3-IC treatment, as determined by confocal microscopy in an animated three-dimensional projection. 107.3-IC treatment drives phosphorylation of Syk; p-Syk co-localizes with dectin-1 as indicated by the white spots. Similar results were obtained for p-SHIP. (MOV 1771 kb)

Supplementary Movie 5

107.3-IC treatment promotes association of FcγRIIB and dectin-1. This movie shows the co-localization pattern of FcγRIIB (green, anti-FcγRIIB-FITC Ab) and of dectin-1 (magenta, anti–dectin-1–APC Ab) in a BM-derived neutrophil 3 min after incubation with IgG1 IC, as determined by confocal microscopy in an animated three-dimensional projection. Co-localization of FcγRIIB and dectin-1 is indicated by the white spots. (MOV 1271 kb)

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Karsten, C., Pandey, M., Figge, J. et al. Anti-inflammatory activity of IgG1 mediated by Fc galactosylation and association of FcγRIIB and dectin-1. Nat Med 18, 1401–1406 (2012). https://doi.org/10.1038/nm.2862

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