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Bacterial recognition by TLR7 in the lysosomes of conventional dendritic cells

Abstract

Little is known of how and where bacterial recognition triggers the induction of type I interferon. Whether the type of recognition receptor used in these responses is determined by the subcellular location of bacteria is not understood. Here we show that phagosomal bacteria such as group B streptococcus, but not cytosolic bacteria, potently induced interferon in conventional dendritic cells by a mechanism that required Toll-like receptor 7, the adaptor MyD88 and the transcription factor IRF1, all of which localized together with bacterial products in degradative vacuoles bearing lysosomal markers. Thus, this cell type–specific recognition pathway links lysosomal recognition of bacterial RNA with a robust, host-protective interferon response.

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Figure 1: Production of high concentrations of IFN-β by cDCs in response to GBS or L. monocytogenes.
Figure 2: Type I interferon responses to various bacterial stimuli.
Figure 3: Phagocytosis is required for IFN-β production.
Figure 4: Signaling requirements for bacteria-induced production of IFN-β.
Figure 5: IFN-β induction by phagosomal bacteria.
Figure 6: Subcellular localization of GBS in cDCs.
Figure 7: Subcellular localization of TLR7, MyD88 and IRF1 in GBS-infected cDCs.
Figure 8: Neonatal mice lacking TLR7 or IRF1 are highly susceptible to GBS infection.
Figure 9: IFN-β responses to GBS extracts and RNA preparations.

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Acknowledgements

We thank T. Taniguchi (University of Tokyo) for permission to use Irf1−/−, Irf3−/− and Irf7−/− mice; D. Golenbock, K. Fitzgerald and E. Lien (University of Massachusetts Medical School) for Irf1−/−, Irf3−/− and Irf7−/− mice; T. Leanderson (University of Lund) for Ifnb−/− mice; G. Grandi (Novartis Vaccines) and D. Portnoy (University of California) for bacterial strains; and F. Ioppolo (Carl Zeiss) for help with fluorescence microscopy. Supported by the Ministero dell'Università e della Ricerca of Italy (Project 'Innovative vaccine against group A streptococcus').

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Authors and Affiliations

Authors

Contributions

G.M. and A.M., design and performance of in vivo and in vitro infection experiments; M.G., design and performance of microscopy studies; S.P., generation of reagents; S.A., provision of experimental models and discussions; G.M., C.Bi., G.T. and C.Be., data analysis and manuscript preparation.

Corresponding author

Correspondence to Giuseppe Teti.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4 and Supplementary Table 1 (PDF 547 kb)

Supplementary Movie 1

Z-stack of optical sections obtained by structured illumination microscopy (confront Fig. 7a). GBS-infected cDC, were incubated with anti-TLR7 and anti-MyD88 primary antibodies, and with FITC- and Texas Red-labeled secondary antibodies, respectively. (MOV 1549 kb)

Supplementary Movie 2

Z-stack of optical sections obtained by structured illumination microscopy (confront Fig. 7b). GBS-infected cDC, were incubated with anti-TLR7 and anti-IRF1 primary antibodies, and FITC- and Alexa 594-labeled secondary antibodies, respectively. (MOV 1575 kb)

Supplementary Movie 3

Z-stack of optical sections obtained by structured illumination microscopy (confront Fig. 7c). GBS-infected cDC, were incubated with anti-IRF1 and anti-MyD88 primary antibodies and with Alexa 488- and Alexa 594-labeled secondary antibodies, respectively. Arrows indicate colocalization of IRF1 with MyD88 in two DNA-negative areas. (MOV 3094 kb)

Supplementary Movie 4

Z-stack of optical sections obtained by structured illumination microscopy (confront supplementary Fig. 3a on line). GBS-infected cDC, were incubated with anti-GBS and anti-IRF1 primary antibodies and with FITC- and Alexa 594-labeled secondary antibodies, respectively. Images show extensive colocalization of GBS antigen with IRF1 in clusters of coalescing, DNA-negative structures. (MOV 1235 kb)

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Mancuso, G., Gambuzza, M., Midiri, A. et al. Bacterial recognition by TLR7 in the lysosomes of conventional dendritic cells. Nat Immunol 10, 587–594 (2009). https://doi.org/10.1038/ni.1733

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