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Select autophagy genes maintain quiescence of tissue-resident macrophages and increase susceptibility to Listeria monocytogenes

Nature Microbiology volume 5, pages 272–281 (2020)Cite this article

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An Author Correction to this article was published on 25 May 2021

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Abstract

Innate and adaptive immune responses that prime myeloid cells, such as macrophages, protect against pathogens1,2. However, if left uncontrolled, these responses may lead to detrimental inflammation3. Macrophages, particularly those resident in tissues, must therefore remain quiescent between infections despite chronic stimulation by commensal microorganisms. The genes required for quiescence of tissue-resident macrophages are not well understood. Autophagy, an evolutionarily conserved cellular process by which cytoplasmic contents are targeted for lysosomal digestion, has homeostatic functions including maintenance of protein and organelle integrity and regulation of metabolism4. Recent research has shown that degradative autophagy, as well as various combinations of autophagy genes, regulate immunity and inflammation5,6,7,8,9,10,11,12. Here, we delineate a function of the autophagy proteins Beclin 1 and FIP200—but not of other essential autophagy components ATG5, ATG16L1 or ATG7—in mediating quiescence of tissue-resident macrophages by limiting the effects of systemic interferon-γ. The perturbation of quiescence in mice that lack Beclin 1 or FIP200 in myeloid cells results in spontaneous immune activation and resistance to Listeria monocytogenes infection. While antibiotic-treated wild-type mice display diminished macrophage responses to inflammatory stimuli, this is not observed in mice that lack Beclin 1 in myeloid cells, establishing the dominance of this gene over effects of the bacterial microbiota. Thus, select autophagy genes, but not all genes essential for degradative autophagy, have a key function in maintaining immune quiescence of tissue-resident macrophages, resulting in genetically programmed susceptibility to bacterial infection.

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Fig. 1: Mice with deficiencies of certain autophagy genes in myeloid cells display enhanced resistance to L. monocytogenes.
Fig. 2: Alterations of peritoneal tissue-resident macrophages in mice with select autophagy gene deficiency.
Fig. 3: Disrupted immune cell homeostasis associated with Becn1 myeloid deficiency.
Fig. 4: IFN-γ signalling is necessary for macrophage activation and has a dominant effect over antibiotic-mediated immune quiescence in Becn1 deficiency.

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Data availability

The data that support the findings of this study are available from the corresponding authors on reasonable request. RNA-seq data are available at the European Nucleotide Archive (PRJEB29191). Single-cell RNA-seq data are available at the GEO database (GSE121521).

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References

  1. Barton, E. S. et al. Herpesvirus latency confers symbiotic protection from bacterial infection. Nature 447, 326–329 (2007).

    Article  CAS  PubMed  Google Scholar 

  2. Netea, M. G. et al. Trained immunity: a program of innate immune memory in health and disease. Science 352, aaf1098 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  3. Medzhitov, R. Origin and physiological roles of inflammation. Nature 454, 428–435 (2008).

    Article  CAS  PubMed  Google Scholar 

  4. Mizushima, N. & Komatsu, M. Autophagy: renovation of cells and tissues. Cell 147, 728–741 (2011).

    Article  CAS  PubMed  Google Scholar 

  5. Cadwell, K. et al. A key role for autophagy and the autophagy gene Atg16l1 in mouse and human intestinal Paneth cells. Nature 456, 259–263 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Saitoh, T. et al. Loss of the autophagy protein Atg16L1 enhances endotoxin- induced IL-1β production. Nature 456, 264–268 (2008).

    Article  CAS  PubMed  Google Scholar 

  7. Levine, B., Mizushima, N. & Virgin, H. W. Autophagy in immunity and inflammation. Nature 469, 323–335 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Kimmey, J. M. et al. Unique role for ATG5 in neutrophil-mediated immunopathology during M. tuberculosis infection. Nature 528, 565–569 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Lu, Q. et al. Homeostatic control of innate lung inflammation by Vici syndrome gene Epg5 and additional autophagy genes promotes influenza pathogenesis. Cell Host Microbe 19, 102–113 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Park, S. et al. Autophagy genes enhance murine gammaherpesvirus 68 reactivation from latency by preventing virus-induced systemic inflammation. Cell Host Microbe 19, 91–101 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Martinez, J. et al. Noncanonical autophagy inhibits the autoinflammatory, lupus-like response to dying cells. Nature 533, 115–119 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Cunha, L. D. et al. LC3-associated phagocytosis in myeloid cells promotes tumor immune tolerance. Cell 175, 429–441 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Huang, S. et al. Immune response in mice that lack the interferon-gamma receptor. Science 259, 1742–1745 (1993).

    Article  CAS  PubMed  Google Scholar 

  14. Unanue, E. R. Inter-relationship among macrophages, natural killer cells and neutrophils in early stages of Listeria resistance. Curr. Opin. Immunol. 9, 35–43 (1997).

    Article  CAS  PubMed  Google Scholar 

  15. Yoshikawa, Y. et al. Listeria monocytogenes ActA-mediated escape from autophagic recognition. Nat. Cell Biol. 11, 1233–1240 (2009).

    Article  CAS  PubMed  Google Scholar 

  16. Yano, T. et al. Autophagic control of Listeria through intracellular innate immune recognition in Drosophila. Nat. Immunol. 9, 908–916 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Gluschko, A. et al. The β2 integrin mac-1 induces protective LC3-associated phagocytosis of Listeria monocytogenes. Cell Host Microbe 23, 324–337 (2018).

    Article  CAS  PubMed  Google Scholar 

  18. Zhao, Z. et al. Autophagosome-independent essential function for the autophagy protein Atg5 in cellular immunity to intracellular pathogens. Cell Host Microbe 4, 458–469 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Meyer-Morse, N. et al. Listeriolysin O is necessary and sufficient to induce autophagy during Listeria monocytogenes infection. PLoS ONE 5, e8610 (2010).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  20. Huang, J. & Brumell, J. H. Bacteria-autophagy interplay: a battle for survival. Nat. Rev. Microbiol. 12, 101–114 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Liang, X. H. et al. Induction of autophagy and inhibition of tumorigenesis by beclin 1. Nature 402, 672–676 (1999).

    Article  CAS  PubMed  Google Scholar 

  22. Lee, H. K. et al. In vivo requirement for Atg5 in antigen presentation by dendritic cells. Immunity 32, 227–239 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Choi, J. et al. The parasitophorous vacuole membrane of Toxoplasma gondii is targeted for disruption by ubiquitin-like conjugation systems of autophagy. Immunity 40, 924–935 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Mitchell, G. et al. Listeria monocytogenes triggers noncanonical autophagy upon phagocytosis, but avoids subsequent growth-restricting xenophagy. Proc. Natl Acad. Sci. USA 115, E210–E217 (2018).

    Article  CAS  PubMed  Google Scholar 

  25. Cain, D. W. et al. Identification of a tissue-specific, C/EBPβ-dependent pathway of differentiation for murine peritoneal macrophages. J. Immunol. 191, 4665–4675 (2013).

    Article  CAS  PubMed  Google Scholar 

  26. Galluzzi, L. et al. Autophagy in malignant transformation and cancer progression. EMBO J. 34, 856–880 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Martinez, J. et al. Molecular characterization of LC3-associated phagocytosis reveals distinct roles for Rubicon, NOX2 and autophagy proteins. Nat. Cell Biol. 17, 893–906 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Hartlova, A. et al. DNA damage primes the type I interferon system via the cytosolic DNA sensor STING to promote anti-microbial innate immunity. Immunity 42, 332–343 (2015).

    Article  PubMed  CAS  Google Scholar 

  29. Liang, Q. et al. Crosstalk between the cGAS DNA sensor and Beclin-1 autophagy protein shapes innate antimicrobial immune responses. Cell Host Microbe 15, 228–238 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Rodier, F. et al. Persistent DNA damage signalling triggers senescence-associated inflammatory cytokine secretion. Nat. Cell Biol. 11, 973–979 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. West, A. P. et al. Mitochondrial DNA stress primes the antiviral innate immune response. Nature 520, 553–557 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  32. Jenkins, S. J. et al. Local macrophage proliferation, rather than recruitment from the blood, is a signature of TH2 inflammation. Science 332, 1284–1288 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Gautier, E. L. et al. Gata6 regulates aspartoacylase expression in resident peritoneal macrophages and controls their survival. J. Exp. Med. 211, 1525–1531 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Rosas, M. et al. The transcription factor Gata6 links tissue macrophage phenotype and proliferative renewal. Science 344, 645–648 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Okabe, Y. & Medzhitov, R. Tissue-specific signals control reversible program of localization and functional polarization of macrophages. Cell 157, 832–844 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Bain, C. C. et al. Long-lived self-renewing bone marrow-derived macrophages displace embryo-derived cells to inhabit adult serous cavities. Nat. Commun. 7, ncomms11852 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Mabbott, N. A. & Gray, D. Identification of co-expressed gene signatures in mouse B1, marginal zone and B2 B-cell populations. Immunology 141, 79–95 (2014).

    Article  CAS  PubMed  Google Scholar 

  38. Ansel, K. M., Harris, R. B. & Cyster, J. G. CXCL13 is required for B1 cell homing, natural antibody production, and body cavity immunity. Immunity 16, 67–76 (2002).

    Article  CAS  PubMed  Google Scholar 

  39. Shi, C. S. et al. Activation of autophagy by inflammatory signals limits IL-1β production by targeting ubiquitinated inflammasomes for destruction. Nat. Immunol. 13, 255–263 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Nakahira, K. et al. Autophagy proteins regulate innate immune responses by inhibiting the release of mitochondrial DNA mediated by the NALP3 inflammasome. Nat. Immunol. 12, 222–230 (2011).

    Article  CAS  PubMed  Google Scholar 

  41. Santeford, A. et al. Impaired autophagy in macrophages promotes inflammatory eye disease. Autophagy 12, 1876–1885 (2016).

  42. Auerbuch, V., Brockstedt, D. G., Meyer-Morse, N., O’Riordan, M. & Portnoy, D. Mice lacking the type I interferon receptor are resistant to Listeria monocytogenes. J. Exp. Med. 200, 527–533 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Carrero, J. A., Calderon, B. & Unanue, E. R. Type I interferon sensitizes lymphocytes to apoptosis and reduces resistance to Listeria infection. J. Exp. Med. 200, 535–540 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. O’Connell, R. M. et al. Type I interferon production enhances susceptibility to Listeria monocytogenes infection. J. Exp. Med. 200, 437–445 (2004).

    Article  PubMed  PubMed Central  Google Scholar 

  45. Pitts, M. G., Myers-Morales, T., D’Orazio, S. E. & Type, I. IFN does not promote susceptibility to foodborne Listeria monocytogenes. J. Immunol. 196, 3109–3116 (2016).

    Article  CAS  PubMed  Google Scholar 

  46. Martin, P. K. et al. Autophagy proteins suppress protective type I interferon signalling in response to the murine gut microbiota. Nat. Microbiol. 3, 1131–1141 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Abt, M. C. et al. Commensal bacteria calibrate the activation threshold of innate antiviral immunity. Immunity 37, 158–170 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Ichinohe, T. et al. Microbiota regulates immune defense against respiratory tract influenza A virus infection. Proc. Natl Acad. Sci. USA 108, 5354–5359 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Steed, A. L. et al. The microbial metabolite desaminotyrosine protects from influenza through type I interferon. Science 357, 498–502 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Kim, K. W. et al. MHC II+ resident peritoneal and pleural macrophages rely on IRF4 for development from circulating monocytes. J. Exp. Med. 213, 1951–1959 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Kanayama, M., He, Y. W. & Shinohara, M. L. The lung is protected from spontaneous inflammation by autophagy in myeloid cells. J. Immunol. 194, 5465–5471 (2015).

    Article  CAS  PubMed  Google Scholar 

  52. Biering, S. B. et al. Viral replication complexes are targeted by LC3-guided interferon-inducible GTPases. Cell Host Microbe 22, 74–85 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. DeSelm, C. J. et al. Autophagy proteins regulate the secretory component of osteoclastic bone resorption. Dev. Cell 21, 966–974 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Medina, D. L. et al. Transcriptional activation of lysosomal exocytosis promotes cellular clearance. Dev. Cell 21, 421–430 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Tsuboyama, K. et al. The ATG conjugation systems are important for degradation of the inner autophagosomal membrane. Science 354, 1036–1041 (2016).

  56. Kuma, A., Komatsu, M. & Mizushima, N. Autophagy-monitoring and autophagy-deficient mice. Autophagy 13, 1619–1628 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Fernandez, A. F. et al. Disruption of the beclin 1-BCL2 autophagy regulatory complex promotes longevity in mice. Nature 558, 136–140 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Miller, B. C. et al. The autophagy gene ATG5 plays an essential role in B lymphocyte development. Autophagy 4, 309–314 (2008).

    Article  CAS  PubMed  Google Scholar 

  59. Sanjuan, M. A. et al. Toll-like receptor signaling in macrophages links the autophagy pathway to phagocytosis. Nature 450, 1253–1257 (2007).

    Article  CAS  PubMed  Google Scholar 

  60. Gan, B. et al. Role of FIP200 in cardiac and liver development and its regulation of TNFα and TSC-mTOR signaling pathways. J. Cell Biol. 175, 121–133 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Hwang, S. et al. Nondegradative role of Atg5-Atg12/ Atg16L1 autophagy protein complex in antiviral activity of interferon gamma. Cell Host Microbe 11, 397–409 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. MacDuff, D. A. et al. Phenotypic complementation of genetic immunodeficiency by chronic herpesvirus infection. eLife 4, e04494 (2015).

  63. Jarjour, N. N. et al. Bhlhe40 mediates tissue-specific control of macrophage proliferation in homeostasis and type 2 immunity. Nat. Immunol. 20, 687–700 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  65. Butler, A., Hoffman, P., Smibert, P., Papalexi, E. & Satija, R. Integrating single-cell transcriptomic data across different conditions, technologies, and species. Nat. Biotechnol. 36, 411–420 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Finak, G. et al. MAST: a flexible statistical framework for assessing transcriptional changes and characterizing heterogeneity in single-cell RNA sequencing data. Genome Biol. 16, 278 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  67. Baldridge, M. T. et al. Commensal microbes and interferon-λ determine persistence of enteric murine norovirus infection. Science 347, 266–269 (2015).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank G. J. Randolph, B. T. Edelson, D. Bhattacharya and the former members of the Virgin laboratory for discussion; J. R. Brestoff and S. Piersma for reading the manuscript; C. Yokoyama and H. Deng for technical assistance; and staff at the McDonnell Genome Institute, Genome Technology Access Center, Flow Cytometry Core Facility, Molecular Microbiology Imaging Facility, Pulmonary Morphology Core, and Center for Human Immunology & Immunotherapy Programs at Washington University School of Medicine for technical support. The research was supported by NIH grant U19 AI109725 and the Crohn’s and Colitis Foundation grant no. 326556 (to H.W.V.), NIH grant U19 AI42784 (to H.W.V. and C.L.S) and NIH grant R01 AI132697 (to C.L.S.). Authors receive support from MES of Russia (project 2.3300.2017/4.6; to K.Z.); NIH K08 (A128043; to C.B.W.); Pediatric Infectious Diseases Society/St Jude Children’s Research Hospital fellowship in basic research (to A.O.); Young Investigators Grant for Probiotics Research from the Global Probiotics Council (to M.T.B.); and Burroughs Wellcome Fund Investigators in the Pathogenesis of Infectious Disease (to C.L.S.).

Author information

Author notes

  1. Qun Lu

    Present address: Center for Life Sciences, School of Life Sciences, Yunnan University, Kunming, China

  2. W. Timothy Schaiff, Dale R. Balce & Herbert W. Virgin

    Present address: Vir Biotechnology, San Francisco, CA, USA

  3. Craig B. Wilen

    Present address: Yale School of Medicine, New Haven, CT, USA

  4. Robert C. Orchard

    Present address: University of Texas Southwestern Medical Center, Dallas, TX, USA

Authors and Affiliations

  1. Department of Pathology and Immunology, Washington University School of Medicine, St Louis, MO, USA

    Ya-Ting Wang, Konstantin Zaitsev, Qun Lu, W. Timothy Schaiff, Ki-Wook Kim, Lindsay Droit, Craig B. Wilen, Chandni Desai, Dale R. Balce, Robert C. Orchard, Sunmin Park, Darren Kreamalmeyer, Scott A. Handley, Maxim N. Artyomov & Herbert W. Virgin

  2. Department of Molecular Microbiology, Washington University School of Medicine, St Louis, MO, USA

    Ya-Ting Wang & Christina L. Stallings

  3. Computer Technologies Department, ITMO University, St Petersburg, Russia

    Konstantin Zaitsev

  4. Department of Cell Biology and Physiology, Washington University School of Medicine, St Louis, MO, USA

    Shan Li

  5. Department of Pharmacology, University of Illinois College of Medicine, Chicago, IL, USA

    Ki-Wook Kim

  6. Department of Pediatrics, Division of Infectious Diseases, Washington University School of Medicine, St Louis, MO, USA

    Anthony Orvedahl

  7. Lauren V. Ackerman Laboratory of Surgical Pathology, Division of Anatomic and Molecular Pathology, Department of Pathology and Immunology, Washington University Medical Center, St Louis, MO, USA

    John D. Pfeifer

  8. Department of Medicine, Division of Infectious Diseases, Washington University School of Medicine, St Louis, MO, USA

    Megan T. Baldridge

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  1. Ya-Ting Wang
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Contributions

Y.-T.W. designed the project, performed experiments, analysed the data and wrote the manuscript. H.W.V. supervised project design and edited the manuscript. C.L.S. assisted with project design and edited the manuscript. Q.L., S.L., W.T.S., L.D. and C.B.W. performed experiments. K.-W.K., D.R.B., R.C.O., A.O., S.P., D.K. and M.T.B. assisted with experiments or project design. C.D. and S.A.H. helped to design RNA-seq experiments and analyse the data. K.Z. and M.N.A. analysed RNA-seq and single-cell RNA-seq data. J.D.P. analysed the histology. All of the authors read and edited the manuscript.

Corresponding authors

Correspondence to Ya-Ting Wang, Christina L. Stallings or Herbert W. Virgin.

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Competing interests

H.W.V. is a founder of Casma Therapeutics and PierianDx. The work reported here was not funded by either company.

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Extended data

Extended Data Fig. 1 Mice with deficiencies of certain autophagy genes in myeloids cells display enhanced resistance to L. monocytogenes.

a, L. monocytogenes CFU in spleen or liver 3 days after infection of mice harboring myeloid deficiency (myeΔ) in multiple autophagy genes (data pooled from 2 experiments, Atg7f/f, n=9; Atg7mye∆, n=13; Atg16l1f/f, n=8; Atg16l1mye∆, n=10; Atg14f/f, n=8; Atg14mye∆, n=11 mice; mean ± s.e.m.; P by 2-tailed t test). b, Western blot analysis of p62, LC3 and GAPDH in peritoneal macrophages from naïve mice (Representative of n≥3 replicates). c, Ex vivo phagocytosis activity of peritoneal macrophages at 0 hour (data pooled from 2 experiments, Becn1f/f, n=12; Becn1mye∆, n=11; mean ± SEM; ns=not significant by 2-tailed t test).

Source data

Extended Data Fig. 2 Mice with Beclin 1 deletion in DCs or neutrophils do not display L. monocytogenes resistance or macrophage activation phenotype.

a, Survival of mice harboring Beclin 1 deletion in CD11c+ and MRP8+ cells vs littermate controls, after i.p. inoculation with 4~5x105 CFUs of L. monocytogenes (Data pooled from 3-4 experiments; not significantly different by Log-rank Mantel-Cox test). b, Flow cytometry of ICAM2+ macrophage subsets in peritoneum of adult naïve mice (Data represents 2 experiments, Becn1f/f, n=3 vs Becn1f/f-Mrp8-cre, n=3; Becn1f/f, n=5 vs Becn1f/f-CD11c-cre, n=4; mean ± SEM; not significant by 2way ANOVA Sidak’s multiple comparisons).

Extended Data Fig. 3 Alterations of peritoneal tissue resident macrophages in mice with central autophagy gene deficiency.

a, b, c, d, Quantification of number of total cells, total ICAM2+ macrophages, and numbers of MHC-IIhigh and MHC-IIlow fractions of ICAM2+ macrophages from peritoneum lavage of mice harboring myeloid deficiency (myeΔ) in multiple autophagy genes and total deficiency (Δ) of Rubicon. (Data pooled from ≥3 independent experiments: Fip200f/f, n=14; Fip200mye∆, n=16; Atg5f/f, n=14; Atg5mye∆, n=12; Atg7f/f, n=8; Atg7mye∆, n=8; Atg16l1f/f, n=18; Atg16l1mye∆, n=13; Rubicon WT, n=10; Rubicon KO, n=9; Atg14f/f, n=16; Atg14mye∆, n=15 mice; mean ± SEM; P, or Padj for multiple comparison with 2-tailed t test). e, Violin plot showing percent of MHC-IIhighICAM2+ macrophages in total peritoneal immune cells (Atg14f/f, n=42; Atg14mye∆, n=38; Becn1f/f, n=12; Becn1mye∆, n=16; Fip200f/f, n = 15; Fip200myeΔ, n = 15; mean ± SEM; Padj by Kruskal–Wallis Dunn’s multiple comparison test).

Extended Data Fig. 4 Beclin 1 deficiency augmented baseline macrophage IFN signaling.

a, Volcano plot shows genes upregulated in macrophages from Becn1mye∆ mice on the left and downregulated on the right in RNA-seq data set (Becn1f/f, n=4; Becn1mye∆, n=4). b, Gene set enrichment analysis of Becn1 dependent signature. (The green curve represents the density of the genes identified in the RNAseq with Normalized Enrichment Score (NES), P value and False Discovery Rate (FDR) listed.) c, Transcript levels of the indicated genes in naïve peritoneal macrophages. (3 independent experiments, Becn1f/f, n=9; Becn1mye∆, n=8; mean ± SEM; P by 2-tailed t test.).

Extended Data Fig. 5 DNA damage response and cell proliferation of Beclin 1 deficient peritoneal macrophages.

a, The presence of DNA double-strand break were revealed by immunofluorescence for γ-H2AX (red) in peritoneal macrophages treated with Bleomycin for 6 hours or untreated p62 is stained in green and nuclei were labeled by DAPI (blue). Cells displaying ≥10 γ-H2AX foci were counted as positive. (Data represents 2 independent experiments; n=4, mean ±SEM; P by 2-tailed t test.). b, Flow cytometry of BrdU incorporation by WT and Beclin 1- deficient ICAM2+ macrophages. (2 independent experiments; Becn1f/f, n=6 vs. Becn1mye∆, n=8; mean ±SEM; not significant by 2-tailed t test.), c, WT and Beclin 1-deficient ICAM2+ macrophages were enumerated after IL-4c injections (3 independent experiments; Becn1f/f+PBS, n=11; Becn1mye∆+PBS, n=16; Becn1f/f+IL4c, n=11; Becn1mye∆+ IL4c, n=19), and analyzed for frequency of BrdU+ and RELMα level (2 independent experiments; Becn1f/f+PBS, n=6; Becn1mye∆+PBS, n=9; Becn1f/f+IL4c, n=11; Becn1mye∆ + IL4c, n = 11; mean ± SEM; Padj by Tukey’s multiple comparisons test.).

Extended Data Fig. 6 Peritoneal lymphocytes changes revealed by Single-cell RNA sequencing.

a and c, Violin plots showing the expression of marker genes of B (a) and T (c) cells clusters by single cell RNAseq. Bar graph comparing fraction size of clusters. b and d Flow cytometry validation on naïve mice (Becn1f/f, n=9; Becn1myeΔ, n = 10; mean ± SEM; P and Padj for multiple comparison, by 2-tailed t test). e, Ifng transcript level among clusters revealed by single cell RNAseq.

Extended Data Fig. 7 Peritoneal macrophage activation in Becn1mye∆ mice is independent of inflammasome and adaptive immune response.

a and b, Peritoneal macrophages obtained from naïve mice were analyzed for total cells (a), total ICAM2+ macrophages, MHC-IIhigh and MHC-IIlow fractions of ICAM2+ macrophages (b) by flow cytometry (Becn1f/fCasp1/11∆, n=6; Becn1mye∆Casp1/11∆, n=6 ; Becn1f/fRag1∆, n=7; Becn1mye∆Rag1∆, n = 8 mice, mean ± SEM, P and Padj by unpaired 2-tailed t test). c and d, Survival of mice after i.p. inoculation of 5x105 CFUs (c) or 5x104 CFU (d) of L. monocytogenes (Data pooled from 3-4 experiments, P by Log-rank Mantel-Cox test).

Extended Data Fig. 8 Ifngr∆ rescues peritoneal immune cell homeostasis in Becn1mye∆ mice.

a–e, Flow cytometry analysis of total cells (a), B cells (b), T cells (c), SPM and monocytes (d), and peritoneal neutrophils (e) and obtained from naïve mice of the indicated genotypes. (Data are from 2 independent experiments; Becn1f/f, n=8 vs. Becn1mye∆, n=9; mean ± SEM; not significant by 2-tailed Mann-Whitney test.). f, Blood neutrophils were analyzed by flow cytometry. (n=6; mean ± SEM; P by 2-tailed Mann-Whitney test.).

Extended Data Fig. 9 Peritoneal macrophage activation in Becn1mye∆ mice is independent of the presence of microbiota.

a and c, Quantification of total peritoneal cells and ICAM2+ macrophages (a) and numbers of ICAM2- macrophages and CD226- fraction of ICAM2- macrophages (c) (Padj by Dunn’s multiple comparisons test). Becn1f/f(Kool-Aid), n=7; Becn1f/f(abx), n=9; Becn1mye∆(Kool-Aid), n=9; Becn1mye∆(abx), n = 10; mean ± SEM, Padj by one-way ANOVA with Dunn’s multiple comparisons test). b, Quantification of 16S copy number from stool samples of mice. (Becn1f/f, n=7 each for kool-aid and abx vs. Becn1mye∆, n=5 each for kool-aid and abx; mean ± SEM, Padj analyzed by Tukey’s multiple comparisons test).

Extended Data Fig. 10 Becn1mye∆ mice did not exhibit enhanced resistance to pulmonary influenza infection.

Mice were infected intranasally with 250 TCID50 influenza A PR8 and monitored for weight loss. (Data pooled from 4 independent experiments, mean ± SEM, not significant by 2way ANOVO for the whole curve or by 2-tailed t test for each time point).

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Supplementary Figs. 1–6 and legends for Supplementary Tables 1 and 2.

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Unprocessed western blots.

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Wang, YT., Zaitsev, K., Lu, Q. et al. Select autophagy genes maintain quiescence of tissue-resident macrophages and increase susceptibility to Listeria monocytogenes. Nat Microbiol 5, 272–281 (2020). https://doi.org/10.1038/s41564-019-0633-0

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