Spatiotemporal regulation of MyD88–IRF-7 signalling for robust type-I interferon induction

Abstract

Robust type-I interferon (IFN-α/β) induction in plasmacytoid dendritic cells, through the activation of Toll-like receptor 9 (TLR9), constitutes a critical aspect of immunity1,2,3,4,5,6. It is absolutely dependent on the transcription factor IRF-7, which interacts with and is activated by the adaptor MyD88. How plasmacytoid dendritic cells, but not other cell types (such as conventional dendritic cells), are able to activate the MyD88–IRF-7-dependent IFN induction pathway remains unknown. Here we show that the spatiotemporal regulation of MyD88–IRF-7 signalling is critical for a high-level IFN induction in response to TLR9 activation. The IFN-inducing TLR9 ligand, A/D-type CpG oligodeoxynucleotide (CpG-A)3,4,8,9,10,11, is retained for long periods in the endosomal vesicles of plasmacytoid dendritic cells, together with the MyD88–IRF-7 complex. However, in conventional dendritic cells, CpG-A is rapidly transferred to lysosomal vesicles. We further show that conventional dendritic cells can also mount a robust IFN induction if CpG-A is manipulated for endosomal retention using a cationic lipid. This strategy also allows us to demonstrate endosomal activation of the IFN pathway by the otherwise inactive TLR9 ligand B/K-type oligodeoxynucleotide (CpG-B)3,4,8,9,10,11,12. Thus, our study offers insights into the regulation of TLR9 signalling in space, potentially suggesting a new avenue for therapeutic intervention.

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Figure 1: Trafficking of CpG ODNs in DCs.
Figure 2: Subcellular localization of MyD88 and IRF-7, and trafficking of CpG ODNs in a macrophage cell line.
Figure 3: High production of type-I IFN in BM-derived cDCs achieved by manipulating CpG-A trafficking with DOTAP.
Figure 4: Induction of type-I IFN by manipulating ODN trafficking using DOTAP.

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Acknowledgements

We thank J. Vilcek, H. Rosen, H. Ohno, F. Nakatsu, A. Nakano, M. Lamphier, L. Cantley and T. Curran for advice, S. Akira for TLR9 and MyD88 mutant mice, G. Trinchieri for the pDC-specific antibody, A. Miyawaki for Venus (a variant of YFP), H. Miyoshi for lentivirus vectors, and M. Shishido for technical assistance. This work was supported in part by a grant for Advanced Research on Cancer and a Grant-In-Aid for Scientific Research on Propriety Areas from the Ministry of Education, Culture, Sports, Science and Technology of Japan, the Uehara Memorial Foundation, the Sumitomo Foundation, the Senri Life Science Foundation and the Nakajima Foundation. H.N. was supported by an Ishidu Shun Memorial Scholarship.

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Correspondence to Tadatsugu Taniguchi.

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Supplementary information

Supplementary Figure S1

This file contains Supplementary Figure S1 a, b, c and d. (PDF 655 kb)

Supplementary Figure S2

This file contains Supplementary Figure S2 a, b, c, d and e. (PDF 1107 kb)

Supplementary Figure S3

This file contains Supplementary Figure S3 a, b, c, d and e. (PDF 612 kb)

Supplementary Figure S4

This file contains Supplementary Figure S4 a, b, c, d and e. (PDF 699 kb)

Supplementary Figure S5

This file contains Supplementary Figure S5 a, b, c, d, e, f and g. Modelling and simulation of CpG-dependent IFN induction. (PDF 95 kb)

Supplementary Video S1

Time-lapse analysis of CpG-A trafficking in bone-marrow-derived conventional dendritic cells. (MOV 1176 kb)

Supplementary Video S2

Time-lapse analysis of CpG-B trafficking in bone-marrow-derived conventional dendritic cells. (MOV 1188 kb)

Supplementary Video S3

Time-lapse analysis of CpG-A/DOTAP trafficking in bone-marrow-derived conventional dendritic cells. (MOV 3638 kb)

Supplementary Notes

This file contains the Supplementary Methods, Legends to accompany Supplementary Figures S1-S5, Supplementary Video Legends, Supplementary Discussion and additional References. (DOC 88 kb)

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Honda, K., Ohba, Y., Yanai, H. et al. Spatiotemporal regulation of MyD88–IRF-7 signalling for robust type-I interferon induction. Nature 434, 1035–1040 (2005). https://doi.org/10.1038/nature03547

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