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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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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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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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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DOI: https://doi.org/10.1038/ni.2509
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