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Guanylate-binding proteins promote activation of the AIM2 inflammasome during infection with Francisella novicida

Nature Immunology volume 16, pages 476–484 (2015)Cite this article

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Abstract

The AIM2 inflammasome detects double-stranded DNA in the cytosol and induces caspase-1-dependent pyroptosis as well as release of the inflammatory cytokines interleukin 1β (IL-1β) and IL-18. AIM2 is critical for host defense against DNA viruses and bacteria that replicate in the cytosol, such as Francisella tularensis subspecies novicida (F. novicida). The activation of AIM2 by F. novicida requires bacteriolysis, yet whether this process is accidental or is a host-driven immunological mechanism has remained unclear. By screening nearly 500 interferon-stimulated genes (ISGs) through the use of small interfering RNA (siRNA), we identified guanylate-binding proteins GBP2 and GBP5 as key activators of AIM2 during infection with F. novicida. We confirmed their prominent role in vitro and in a mouse model of tularemia. Mechanistically, these two GBPs targeted cytosolic F. novicida and promoted bacteriolysis. Thus, in addition to their role in host defense against vacuolar pathogens, GBPs also facilitate the presentation of ligands by directly attacking cytosolic bacteria.

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Figure 1: Products of ISGs are required for AIM2 activation during infection with F. novicida.
Figure 2: Screening by RNA-mediated interference identifies members of the GBP family as activators of AIM2.
Figure 3: Macrophages from Gbpchr3-deficient mice have deficient activation of AIM2 in response to F. novicida.
Figure 4: GBP2 and GBP5 independently control activation of AIM2 during infection with F. novicida.
Figure 5: The escape of F. novicida from phagosomes is GBP independent.
Figure 6: GBPs promote the activation of AIM2 by inducing bacteriolysis.
Figure 7: GBPs restrict the intracellular replication of F. novicida.
Figure 8: GBPs control host defense against F. novicida in vivo.

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References

  1. Paludan, S.R. & Bowie, A.G. Immune sensing of DNA. Immunity 38, 870–880 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Ishikawa, H., Ma, Z. & Barber, G.N. STING regulates intracellular DNA-mediated, type I interferon-dependent innate immunity. Nature 461, 788–792 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Sauer, J.D. et al. The N-ethyl-N-nitrosourea-induced Goldenticket mouse mutant reveals an essential function of Sting in the in vivo interferon response to Listeria monocytogenes and cyclic dinucleotides. Infect. Immun. 79, 688–694 (2011).

    CAS  PubMed  Google Scholar 

  4. Hornung, V. et al. AIM2 recognizes cytosolic dsDNA and forms a caspase-1-activating inflammasome with ASC. Nature 458, 514–518 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Fernandes-Alnemri, T., Yu, J.W., Datta, P., Wu, J. & Alnemri, E.S. AIM2 activates the inflammasome and cell death in response to cytoplasmic DNA. Nature 458, 509–513 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Roberts, T.L. et al. HIN-200 proteins regulate caspase activation in response to foreign cytoplasmic DNA. Science 323, 1057–1060 (2009).

    CAS  PubMed  Google Scholar 

  7. Bürckstümmer, T. et al. An orthogonal proteomic-genomic screen identifies AIM2 as a cytoplasmic DNA sensor for the inflammasome. Nat. Immunol. 10, 266–272 (2009).

    PubMed  Google Scholar 

  8. Ge, J., Gong, Y.N., Xu, Y. & Shao, F. Preventing bacterial DNA release and absent in melanoma 2 inflammasome activation by a Legionella effector functioning in membrane trafficking. Proc. Natl. Acad. Sci. USA 109, 6193–6198 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Fernandes-Alnemri, T. et al. The AIM2 inflammasome is critical for innate immunity to Francisella tularensis. Nat. Immunol. 11, 385–393 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Jones, J.W. et al. Absent in melanoma 2 is required for innate immune recognition of Francisella tularensis. Proc. Natl. Acad. Sci. USA 107, 9771–9776 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Kim, S. et al. Listeria monocytogenes is sensed by the NLRP3 and AIM2 inflammasome. Eur. J. Immunol. 40, 1545–1551 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Sauer, J.D. et al. Listeria monocytogenes triggers AIM2-mediated pyroptosis upon infrequent bacteriolysis in the macrophage cytosol. Cell Host Microbe 7, 412–419 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Rathinam, V.A. et al. The AIM2 inflammasome is essential for host defense against cytosolic bacteria and DNA viruses. Nat. Immunol. 11, 395–402 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Briken, V., Ahlbrand, S.E. & Shah, S. Mycobacterium tuberculosis and the host cell inflammasome: a complex relationship. Front. Cell. Infect. Microbiol. 3, 62 (2013).

    PubMed  PubMed Central  Google Scholar 

  15. Peng, K., Broz, P., Jones, J., Joubert, L.M. & Monack, D. Elevated AIM2-mediated pyroptosis triggered by hypercytotoxic Francisella mutant strains is attributed to increased intracellular bacteriolysis. Cell. Microbiol. 13, 1586–1600 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Muruve, D.A. et al. The inflammasome recognizes cytosolic microbial and host DNA and triggers an innate immune response. Nature 452, 103–107 (2008).

    CAS  PubMed  Google Scholar 

  17. Henry, T., Brotcke, A., Weiss, D.S., Thompson, L.J. & Monack, D.M. Type I interferon signaling is required for activation of the inflammasome during Francisella infection. J. Exp. Med. 204, 987–994 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Cole, L.E. et al. Macrophage proinflammatory response to Francisella tularensis live vaccine strain requires coordination of multiple signaling pathways. J. Immunol. 180, 6885–6891 (2008).

    CAS  PubMed  Google Scholar 

  19. Cole, L.E. et al. Toll-like receptor 2-mediated signaling requirements for Francisella tularensis live vaccine strain infection of murine macrophages. Infect. Immun. 75, 4127–4137 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Jones, J.W., Broz, P. & Monack, D.M. Innate immune recognition of Francisella tularensis: activation of type-I interferons and the inflammasome. Front Microbiol 2, 16 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Kim, B.H., Shenoy, A.R., Kumar, P., Bradfield, C.J. & MacMicking, J.D. IFN-inducible GTPases in host cell defense. Cell Host Microbe 12, 432–444 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Howard, J.C., Hunn, J.P. & Steinfeldt, T. The IRG protein-based resistance mechanism in mice and its relation to virulence in Toxoplasma gondii. Curr. Opin. Microbiol. 14, 414–421 (2011).

    CAS  PubMed  Google Scholar 

  23. Yamamoto, M. et al. A cluster of interferon-γ-inducible p65 GTPases plays a critical role in host defense against Toxoplasma gondii. Immunity 37, 302–313 (2012).

    CAS  PubMed  Google Scholar 

  24. Kim, B.H. et al. A family of IFN-gamma-inducible 65-kD GTPases protects against bacterial infection. Science 332, 717–721 (2011).

    CAS  PubMed  Google Scholar 

  25. Degrandi, D. et al. Extensive characterization of IFN-induced GTPases mGBP1 to mGBP10 involved in host defense. J. Immunol. 179, 7729–7740 (2007).

    CAS  PubMed  Google Scholar 

  26. Kresse, A. et al. Analyses of murine GBP homology clusters based on in silico, in vitro and in vivo studies. BMC Genomics 9, 158 (2008).

    PubMed  PubMed Central  Google Scholar 

  27. Degrandi, D. et al. Murine guanylate binding protein 2 (mGBP2) controls Toxoplasma gondii replication. Proc. Natl. Acad. Sci. USA 110, 294–299 (2013).

    CAS  PubMed  Google Scholar 

  28. Kravets, E. et al. The GTPase activity of murine guanylate-binding protein 2 (mGBP2) controls the intracellular localization and recruitment to the parasitophorous vacuole of Toxoplasma gondii. J. Biol. Chem. 287, 27452–27466 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Meunier, E. et al. Caspase-11 activation requires lysis of pathogen-containing vacuoles by IFN-induced GTPases. Nature 509, 366–370 (2014).

    CAS  PubMed  Google Scholar 

  30. Checroun, C., Wehrly, T.D., Fischer, E.R., Hayes, S.F. & Celli, J. Autophagy-mediated reentry of Francisella tularensis into the endocytic compartment after cytoplasmic replication. Proc. Natl. Acad. Sci. USA 103, 14578–14583 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Nothelfer, K., Dias Rodrigues, C., Bobard, A., Phalipon, A. & Enninga, J. Monitoring Shigella flexneri vacuolar escape by flow cytometry. Virulence 2, 54–57 (2011).

    PubMed  Google Scholar 

  32. Juruj, C. et al. Caspase-1 activity affects AIM2 speck formation/stability through a negative feedback loop. Front. Cell. Infect. Microbiol. 3, 1–11 (2013).

    Google Scholar 

  33. Chong, A. et al. Cytosolic clearance of replication-deficient mutants reveals Francisella tularensis interactions with the autophagic pathway. Autophagy 8, 1342–1356 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Miao, E.A. et al. Caspase-1-induced pyroptosis is an innate immune effector mechanism against intracellular bacteria. Nat. Immunol. 11, 1136–1142 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  35. Zhou, H. et al. Genome-wide RNAi screen in IFN-γ-treated human macrophages identifies genes mediating resistance to the intracellular pathogen Francisella tularensis. PLoS ONE 7, e31752 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  36. Edwards, J.A., Rockx-Brouwer, D., Nair, V. & Celli, J. Restricted cytosolic growth of Francisella tularensis subsp. tularensis by IFN-γ activation of macrophages. Microbiology 156, 327–339 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Mariathasan, S., Weiss, D.S., Dixit, V.M. & Monack, D.M. Innate immunity against Francisella tularensis is dependent on the ASC/caspase-1 axis. J. Exp. Med. 202, 1043–1049 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Pilla, D.M. et al. Guanylate binding proteins promote caspase-11-dependent pyroptosis in response to cytoplasmic LPS. Proc. Natl. Acad. Sci. USA 111, 6046–6051 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Hagar, J.A., Powell, D.A., Aachoui, Y., Ernst, R.K. & Miao, E.A. Cytoplasmic LPS activates caspase-11: implications in TLR4-independent endotoxic shock. Science 341, 1250–1253 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Woodward, J.J., Iavarone, A.T. & Portnoy, D.A. c-di-AMP secreted by intracellular Listeria monocytogenes activates a host type I interferon response. Science 328, 1703–1705 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Sun, L., Wu, J., Du, F., Chen, X. & Chen, Z.J. Cyclic GMP-AMP synthase is a cytosolic DNA sensor that activates the type I interferon pathway. Science 339, 786–791 (2013).

    CAS  PubMed  Google Scholar 

  42. Li, X.D. et al. Pivotal roles of cGAS-cGAMP signaling in antiviral defense and immune adjuvant effects. Science 341, 1390–1394 (2013).

    CAS  PubMed  Google Scholar 

  43. Manzanillo, P.S., Shiloh, M.U., Portnoy, D.A. & Cox, J.S. Mycobacterium tuberculosis activates the DNA-dependent cytosolic surveillance pathway within macrophages. Cell Host Microbe 11, 469–480 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Broz, P. et al. Caspase-11 increases susceptibility to Salmonella infection in the absence of caspase-1. Nature 490, 288–291 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Rathinam, V.A. et al. TRIF licenses caspase-11-dependent NLRP3 inflammasome activation by gram-negative bacteria. Cell 150, 606–619 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Martens, S. et al. Disruption of Toxoplasma gondii parasitophorous vacuoles by the mouse p47-resistance GTPases. PLoS Pathog. 1, e24 (2005).

    PubMed  PubMed Central  Google Scholar 

  47. Howard, J.C., Hunn, J.P. & Steinfeldt, T. The IRG protein-based resistance mechanism in mice and its relation to virulence in Toxoplasma gondii. Curr. Opin. Microbiol. 14, 414–421 (2011).

    CAS  PubMed  Google Scholar 

  48. Bekpen, C. et al. The interferon-inducible p47 (IRG) GTPases in vertebrates: loss of the cell autonomous resistance mechanism in the human lineage. Genome Biol. 6, R92 (2005).

    PubMed  PubMed Central  Google Scholar 

  49. Haldar, A.K. et al. IRG and GBP host resistance factors target aberrant, “non-self” vacuoles characterized by the missing of “self” IRGM proteins. PLoS Pathog. 9, e1003414 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank N. Gekara (Umea University) for StingGt/Gt mice; M. Roth and S. Hofer for support with mouse experiments; D. Monack (Stanford University) for chicken antibody to F. novicida; N. Kayagaki (Genentech) and V. Dixit (Genentech) for Gbp5−/− and Aim2−/− mice, rat antibody to ASC and rat antibody to caspase-1; L. Gallagher and C. Manoil (University of Washington) for plasmid pFFlp; O. Allatif for statistical analysis; the Imaging and FACS Core Facilities of the Biozentrum, University of Basel, for technical assistance; and the Plateau de Biologie Expérimentale de la Souris and the flow cytometry platform of SFR Biosciences Gerland-Lyon Sud. Supported by Swiss National Science Foundation (PP00P3_139120/1 to P.B.), the University of Basel (ID2153162 to P.B.), the European Research Council (311542 to T.H.) and the Déléguation Générale de l'Armement (M.R.).

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  1. Etienne Meunier and Pierre Wallet: These authors contributed equally to this work.

Authors and Affiliations

  1. Focal Area Infection Biology, Biozentrum, University of Basel, Basel, Switzerland

    Etienne Meunier, Roland F Dreier, Leonie Anton, Sebastian Rühl, Mathias S Dick, Anne Kistner & Petr Broz

  2. Centre International de Recherche en Infectiologie, Inserm U1111, CNRS UMR 5308, Université Claude Bernard Lyon-1, Ecole Normale Supérieure, Lyon, France

    Pierre Wallet, Stéphanie Costanzo, Mélanie Rigard & Thomas Henry

  3. Structure Fédérative de Recherche Biosciences, UMS344/US8, Inserm, CNRS, Université Claude Bernard Lyon-1, Ecole Normale Supérieure, Lyon, France

    Sébastien Dussurgey

  4. Institute of Medical Microbiology and Hospital Hygiene, Heinrich-Heine-University Duesseldorf, Duesseldorf, Germany

    Daniel Degrandi & Klaus Pfeffer

  5. Department of Microbiology and Immunology, Osaka University, Osaka, Japan

    Masahiro Yamamoto

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Contributions

E.M., P.W., T.H. and P.B. conceived of the research; E.M., P.W., R.F.D., S.C., L.A., S.R., S.D., M.S.D., A.K., M.R., T.H. and P.B. performed experiments; D.D., K.P. and M.Y. provided reagents; and T.H. and P.B. wrote the manuscript.

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Integrated supplementary information

Supplementary Figure 1 Induction of Ifnb, Aim2 or Gbp mRNA in dependence of STING and IFNAR signaling.

(a) Induction of Ifnb expression from unprimed wild-type (WT), Tlr2–/–, Myd88–/–, Trif–/– and Stinggt/gt bone-marrow derived macrophages (BMDMs) following infection with wild-type F. novicida U112 for 6 h. (b–e) Induction of Gbp2, Gbp3, Gbp5 or Aim2 expression from unprimed wild-type, Tlr2–/–, Myd88–/–, Ifnar1–/– and Stinggt/gt BMDMs following infection with wild-type F. novicida U112 for 6 h. Graphs show mean and s.d. of quadruplicate assays and data are representative of two (b-d) or three (a) independent experiments.

Supplementary Figure 2 Gbp knockdown efficiency and real-time cell death after knock-down of Gbps.

(a) Induction of the expression of individual Gbps from LPS/IFNγ-primed wild-type BMDMs treated with Non-Targeting (NT) or the indicated gene-specific siRNA for 22 h. Graph shows mean and s.d. of quadruplicate wells. Gbp1, 4, 6/10, 11 were not tested due to their low expression (see Fig. 2b). (b) Cell death as measured by propidium iodide (PI) influx in real-time in unprimed wild-type BMDMs infected with wild-type F. novicida U112. BMDMs were treated with Non-Targeting (NT) or indicated gene-specific siRNA for 48 h before infection. Graphs show mean and s.d. of triplicate assays and data are representative of three
 independent experiments. *, p<0.05; **, p<0.01; NS, not significant (two-tailed unpaired t-test).

Supplementary Figure 3 Aim2 induction and real-time cell death assay in Gbp-deficient and Ifnar1-deficient cells.

(a) Induction of Aim2 expression from unprimed wild-type, Gbpchr3-deleted and Ifnar1–/– BMDMs infected with wild-type F. novicida U112 for 6 h. *, p<0.01; NS, not significant (two-tailed unpaired t-test). (b) Cell death as measured by propidium iodide influx in real-time in unprimed wild-type, Gbpchr3-deleted and Ifnar1–/– BMDMs left uninfected (UI) or infected with wild-type F. novicida U112. Graphs show mean and s.d. of triplicate assays and data are representative of two (a) and three (b) independent experiments.

Supplementary Figure 4 Ectopic expression of GBPs in Ifnar1–/– cells and efficiency of Gbp2 and Gbp5 knockdown.

(a–c) Ectopic expression of GBP2 or GBP5 do not complement type-I-IFN receptor deficiency. Ifnar1–/– macrophages were transduced with lentivirus encoding either GFP only (EV = empty vector) and GFP-GBP2 and GFP-GBP5. 48 h post transduction, macrophages were infected with F. novicida at the indicated MOI. IL-1β concentration in the supernatant was determined at 7 h and 10.5 h post infection (a). Specific ectopic expression was verified by quantifying the Gbp3 (control), Gbp2 and Gbp5 transcript levels. Results are expressed as fold induction relative to the transcript level in Ifnar1–/– macrophages transduced with empty vector control (b). Graphs show mean and s.d. of triplicate assays. The percentage of transduced cells was determined by flow cytometry based on GFP expression (c). (d) RT-PCR for Gbp2 and Gbp5 expression from unprimed wild-type, Gbp2–/– and Gbp5–/– BMDMs treated with Non-Targeting or the indicated gene-specific siRNA for 22 h and infected for 8 h with wild-type F. novicida U112. Graphs show mean and s.d. of quadruplicate assays and data are representative of independent two experiments. *, p<0.05; **, p<0.01; NS, not significant (two-tailed unpaired t-test). ND, not detected.

Supplementary Figure 5 Phagosomal rupture assay using the CCF4/β-lactamase system.

Wild-type, Ifnar1–/– and Gbpchr3-deleted BMDMs were primed for 16 h with IFN-β (500 units/ml), infected for 1 h with wild-type F. novicida (FN) U112, a β-lactamase-deficient mutant (Δbla) or a ΔFPI mutant and loaded with CCF4-AM for 1 h before analysis by flow cytometry. Phagosomal rupture is associated with β-lactamase (encoded by FTN_1072) release into the cytosol and cleavage of the CCF4 substrate (maximum emission at 520 nm) into a product which emits with a maximum of fluorescence at 447 nm. FACS plots show pooled data from three independent samples and representative of three independent experiments. Live cells (propidium iodide negative) are shown.

Supplementary Figure 6 GBPs co-localize with irregularly shaped bacteria next to ASC specks.

Wild-type BMDMs infected with wild-type F. novicida for 8 h, stained for DNA (DAPI), GBP2, F. novicida and ASC. Scale bars: 10 μm. Data are representative of three
 independent experiments.

Supplementary Figure 7 Analysis of bacterial replication in infected macrophages.

(a) Quantification of bacterial loads in single cells by high-resolution microscopy in flow over time. Wild-type BMDMs were left uninfected or infected with GFP+-wild-type F. novicida or a ΔFPI mutant at an MOI of 10 for 0-12 h, fixed and analyzed by ImageStreamTM microscopy in flow. Each bar corresponds to the number of cells with the indicated numbers of bacteria per cell grouped by increments of 5... Wild-type UI vs. wild-type MOI 10 3h p>0.9999, wild-type UI vs. wild-type MOI 10 6h p>0.9999, wild-type UI vs. wild-type MOI 10 9h p<0.0001, wild-type UI vs. wild-type MOI 10 12h p<0.0001, wild-type MOI 10 12h vs. ΔFPI MOI 10 12h p<0.0001 Kolmogorov-Smirnov test with Bonferroni correction). (b) Quantification of bacterial loads as determined by microscopy. Wild-type, Aim2–/–, Gbpchr3-deleted and Ifnar1–/– BMDMs were infected with GFP+-wild-type F. novicida at an MOI of 10 for 16 h, fixed and analyzed by confocal microscopy. Graph show pooled data from 2 independent experiments (n>1000 bacteria counted). *, p<0.05; **, p<0.01; ***, p<0.001 (two-tailed unpaired t-test).

Supplementary Figure 8 Bacteriolysis and cell death during F. novicida infection are independent of ROS or NO production.

(a) Quantification of lysed (propidium iodide+ F. novicida) in IFN-γ-primed wild-type and Nos2–/–/Cybb–/–BMDMs infected for 8 h with wild-type F. novicida. Imaging of lysed (propidium iodide+) F. novicida in IFN-γ-primed wild-type and Nos2–/–/Cybb–/–BMDMs infected for 8 h with wild-type F. novicida. Arrowheads indicate region in insets. Scale bars 10 μm. (b) LDH release from naïve or IFN-γ-primed wild-type and Nos2–/–/Cybb–/– BMDMs infected for 8 h with wild-type F. novicida U112. Graphs show mean and s.d. of quadruplicate wells and data are representative of three
 independent experiments. NS, not significant (two-tailed unpaired t-test).

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Supplementary Table 1

List of siRNA used for screening (XLS 282 kb)

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Meunier, E., Wallet, P., Dreier, R. et al. Guanylate-binding proteins promote activation of the AIM2 inflammasome during infection with Francisella novicida. Nat Immunol 16, 476–484 (2015). https://doi.org/10.1038/ni.3119

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