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Identification of regulators of the innate immune response to cytosolic DNA and retroviral infection by an integrative approach

Nature Immunology volume 14, pages 179–185 (2013)Cite this article

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Abstract

The innate immune system senses viral DNA that enters mammalian cells, or in aberrant situations self-DNA, and triggers type I interferon production. Here we present an integrative approach that combines quantitative proteomics, genomics and small molecule perturbations to identify genes involved in this pathway. We silenced 809 candidate genes, measured the response to dsDNA and connected resulting hits with the known signaling network. We identified ABCF1 as a critical protein that associates with dsDNA and the DNA-sensing components HMGB2 and IFI204. We also found that CDC37 regulates the stability of the signaling molecule TBK1 and that chemical inhibition of the CDC37-HSP90 interaction and several other pathway regulators potently modulates the innate immune response to DNA and retroviral infection.

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Figure 1: Generation of a candidate gene set by quantitative proteomics and curation.
Figure 2: RNAi screen defines functional components of the ISD-sensing pathway.
Figure 3: Validation of Abcf1 as a regulator of the ISD-sensing response.
Figure 4: Small molecule inhibitors modulate the ISD-sensing response.
Figure 5: ABCF1 interacts with HMGB2, IFI204 and SET complex.
Figure 6: Inhibition of identified regulators by RNAi or small molecules modulates the innate immune response to retroviral infection.

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Acknowledgements

We are grateful to Y.J. Crow for AGS patient fibroblasts and control cells; T. Lindahl for Trex1−/− MEFs and wild-type control MEFs; D. Sabatini and D. Kwiatkowski for p53−/− MEFs; N.A. DeLuca for HSV-1 d109 virus; C. Shamu (Institute of Chemistry and Cell Biology at Harvard Medical School) for the siRNA library and expert advice; The RNAi Consortium at the Broad Institute for assistance with siRNA screening; J. Qiao for assistance with mass spectrometry; M. Rooney and C. Ye for advice about statistical analyses; D. Londono for help with microscopy; J. Kagan, N. Haining, L. Glimcher and M. Brenner for valuable discussions; and W.F. Pendergraft III and other members of the Hacohen laboratory for critical review of the manuscript. This work was supported by the US National Human Genome Research Institute grant HG005062 and US National Institutes of Health Director's New Innovator award DP2OD002230 (N.H.). M.N.L. is supported by a National Institutes of Health Medical Scientist Training Program fellowship; F.D. by a GlaxoSmithKline Immune Disease Institute fellowship; and J.L. by National Institute of Allergy and Infectious Diseases grant AI102816.

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Author notes

  1. Shao-En Ong

    Present address: Present address: Department of Pharmacology, University of Washington, Seattle, Washington, USA.,

Authors and Affiliations

  1. Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA

    Mark N Lee, Matthew Roy, Shao-En Ong, Philipp Mertins, Alexandra-Chloé Villani, Weibo Li, John G Doench, Steven A Carr & Nir Hacohen

  2. Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Charlestown, Massachusetts, USA

    Mark N Lee, Matthew Roy, Weibo Li & Nir Hacohen

  3. Department of Medicine, Harvard Medical School, Boston, Massachusetts, USA

    Mark N Lee, Matthew Roy & Nir Hacohen

  4. Immune Disease Institute and Program in Cellular and Molecular Medicine, Children's Hospital, Boston, Massachusetts, USA

    Farokh Dotiwala & Judy Lieberman

  5. Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA

    Farokh Dotiwala & Judy Lieberman

  6. Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, USA

    Jayita Sen, Megan H Orzalli & David M Knipe

  7. Department of Medicine, Pulmonary Center, National Emerging Infectious Diseases Laboratory, Boston University School of Medicine, Boston, Massachusetts, USA

    Igor Kramnik

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Contributions

M.N.L., M.R., S.-E.O., P.M., W.L., F.D. and J.S. performed experiments and analyzed data; A.-C.V. analyzed data; J.G.D., M.H.O. and I.K. provided reagents; D.M.K., J.L. and S.A.C. supervised experiments; N.H. designed and supervised the study. M.N.L. and N.H. wrote the manuscript.

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Correspondence to Nir Hacohen.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–6 (PDF 2366 kb)

Supplementary Table 1

STING-interacting SILAC. (XLS 229 kb)

Supplementary Table 2

DNA-interacting SILAC. (XLS 187 kb)

Supplementary Table 3

siRNA screen. (XLS 160 kb)

Supplementary Table 4

ABCF1-interacting SILAC. (XLS 300 kb)

Supplementary Table 5

Sequences. (XLSX 35 kb)

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Lee, M., Roy, M., Ong, SE. et al. Identification of regulators of the innate immune response to cytosolic DNA and retroviral infection by an integrative approach. Nat Immunol 14, 179–185 (2013). https://doi.org/10.1038/ni.2509

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