Abstract
Despite their importance, the molecular circuits that control the differentiation of naive T cells remain largely unknown. Recent studies that reconstructed regulatory networks in mammalian cells have focused on short-term responses and relied on perturbation-based approaches that cannot be readily applied to primary T cells. Here we combine transcriptional profiling at high temporal resolution, novel computational algorithms, and innovative nanowire-based perturbation tools to systematically derive and experimentally validate a model of the dynamic regulatory network that controls the differentiation of mouse TH17 cells, a proinflammatory T-cell subset that has been implicated in the pathogenesis of multiple autoimmune diseases. The TH17 transcriptional network consists of two self-reinforcing, but mutually antagonistic, modules, with 12 novel regulators, the coupled action of which may be essential for maintaining the balance between TH17 and other CD4+ T-cell subsets. Our study identifies and validates 39 regulatory factors, embeds them within a comprehensive temporal network and reveals its organizational principles; it also highlights novel drug targets for controlling TH17 cell differentiation.
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References
Bettelli, E., Oukka, M. & Kuchroo, V. K. TH-17 cells in the circle of immunity and autoimmunity. Nature Immunol. 8, 345–350 (2007)
Zhou, L. et al. TGF-β-induced Foxp3 inhibits TH17 cell differentiation by antagonizing RORγt function. Nature 453, 236–240 (2008)
O’Shea, J. et al. Signal transduction and Th17 cell differentiation. Microbes Infect. 11, 599–611 (2009)
Zhou, L. & Littman, D. Transcriptional regulatory networks in Th17 cell differentiation. Curr. Opin. Immunol. 21, 146–152 (2009)
Korn, T., Bettelli, E., Oukka, M. & Kuchroo, V. K. IL-17 and Th17 cells. Annu. Rev. Immunol. 27, 485–517 (2009)
Amit, I. et al. Unbiased reconstruction of a mammalian transcriptional network mediating pathogen responses. Science 326, 257–263 (2009)
Novershtern, N. et al. Densely interconnected transcriptional circuits control cell states in human hematopoiesis. Cell 144, 296–309 (2011)
Litvak, V. et al. Function of C/EBPδ in a regulatory circuit that discriminates between transient and persistent TLR4-induced signals. Nature Immunol. 10, 437–443 (2009)
Suzuki, H. et al. The transcriptional network that controls growth arrest and differentiation in a human myeloid leukemia cell line. Nature Genet. 41, 553–562 (2009)
Shalek, A. K. et al. Vertical silicon nanowires as a universal platform for delivering biomolecules into living cells. Proc. Natl Acad. Sci. USA 107, 1870–1875 (2010)
Lee, Y. et al. Induction and molecular signature of pathogenic TH17 cells. Nature Immunol. 13, 991–999 (2012)
Linhart, C., Halperin, Y. & Shamir, R. Transcription factor and microRNA motif discovery: the Amadeus platform and a compendium of metazoan target sets. Genome Res. 18, 1180–1189 (2008)
Zheng, G. et al. ITFP: an integrated platform of mammalian transcription factors. Bioinformatics 24, 2416–2417 (2008)
Wilson, N. K. et al. Combinatorial transcriptional control in blood stem/progenitor cells: genome-wide analysis of ten major transcriptional regulators. Cell Stem Cell 7, 532–544 (2010)
Lachmann, A. et al. ChEA: transcription factor regulation inferred from integrating genome-wide ChIP-X experiments. Bioinformatics 26, 2438–2444 (2010)
Liberzon, A. et al. Molecular signatures database (MSigDB) 3.0. Bioinformatics 27, 1739–1740 (2011)
Jiang, C., Xuan, Z., Zhao, F. & Zhang, M. TRED: a transcriptional regulatory element database, new entries and other development. Nucleic Acids Res. 35, D137–D140 (2007)
Elkon, R., Linhart, C., Sharan, R., Shamir, R. & Shiloh, Y. Genome-wide in silico identification of transcriptional regulators controlling the cell cycle in human cells. Genome Res. 13, 773–780 (2003)
Heng, T. S. & Painter, M. W. The Immunological Genome Project: networks of gene expression in immune cells. Nature Immunol. 9, 1091–1094 (2008)
Schwanhäusser, B. et al. Global quantification of mammalian gene expression control. Nature 473, 337–342 (2011)
Dardalhon, V. et al. Lentivirus-mediated gene transfer in primary T cells is enhanced by a central DNA flap. Gene Ther. 8, 190–198 (2001)
McManus, M. et al. Small interfering RNA-mediated gene silencing in T lymphocytes. J. Immunol. 169, 5754 (2002)
Shalek, A. K. et al. Nanowire-mediated delivery enables functional interrogation of primary immune cells: application to the analysis of chronic lymphocytic leukemia. Nano Lett. 12, 6498–6504 (2012)
Segrè, D., Deluna, A., Church, G. M. & Kishony, R. Modular epistasis in yeast metabolism. Nature Genet. 37, 77–83 (2005)
Peleg, T., Yosef, N., Ruppin, E. & Sharan, R. Network-free inference of knockout effects in yeast. PLOS Comput. Biol. 6, e1000635 (2010)
Marson, A. et al. Foxp3 occupancy and regulation of key target genes during T-cell stimulation. Nature 445, 931–935 (2007)
Zheng, Y. et al. Genome-wide analysis of Foxp3 target genes in developing and mature regulatory T cells. Nature 445, 936–940 (2007)
Glasmacher, E. et al. A genomic regulatory element that directs assembly and function of immune-specific AP-1-IRF complexes. Science 338, 975–980 (2012)
Ciofani, M. et al. A validated regulatory network for Th17 cell specification. Cell 151, 289–303 (2012)
Glasmacher, E. et al. A genomic regulatory element that directs assembly and function of immune-specific AP-1-IRF complexes. Science 338, 975–980 (2012)
Okamoto, M. et al. Mina, an Il4 repressor, controls T helper type 2 bias. Nature Immunol. 10, 872–879 (2009)
Hill, J. A. et al. Foxp3 transcription-factor-dependent and -independent regulation of the regulatory T cell transcriptional signature. Immunity 27, 786–800 (2007)
Korn, T. et al. IL-21 initiates an alternative pathway to induce proinflammatory TH17 cells. Nature 448, 484–487 (2007)
Waldner, H., Sobel, R. A., Howard, E. & Kuchroo, V. K. Fas- and FasL-deficient mice are resistant to induction of autoimmune encephalomyelitis. J. Immunol. 159, 3100–3103 (1997)
Teitell, M. A. OCA-B regulation of B-cell development and function. Trends Immunol. 24, 546–553 (2003)
Laurence, A. et al. Interleukin-2 signaling via STAT5 constrains T helper 17 cell generation. Immunity 26, 371–381 (2007)
Choi, S.-J. et al. Tsc-22 enhances TGF-β signaling by associating with Smad4 and induces erythroid cell differentiation. Mol. Cell. Biochem. 271, 23–28 (2005)
Peters, A., Lee, Y. & Kuchroo, V. K. The many faces of Th17 cells. Curr. Opin. Immunol. 23, 702–706 (2011)
Chaudhry, A. et al. Interleukin-10 signaling in regulatory T cells is required for suppression of Th17 cell-mediated inflammation. Immunity 34, 566–578 (2011)
Zielinski, C. E. et al. Pathogen-induced human TH17 cells produce IFN-γ or IL-10 and are regulated by IL-1β. Nature 484, 514–518 (2012)
Jing, Y. et al. A mechanistic study on the effect of dexamethasone in moderating cell death in Chinese Hamster Ovary cell cultures. Biotechnol. Prog. 28, 490–496 (2012)
Hu, S. M., Luo, Y. L., Lai, W. Y. & Chen, P. F. Effects of dexamethasone on intracellular expression of Th17 cytokine interleukin 17 in asthmatic mice [in Chinese]. Nan Fang Yi Ke Da Xue Xue Bao 29, 1185–1188 (2009)
Yang, X. P. et al. Opposing regulation of the locus encoding IL-17 through direct, reciprocal actions of STAT3 and STAT5. Nature Immunol. 12, 247–254 (2011)
Jostins, L. et al. Host–microbe interactions have shaped the genetic architecture of inflammatory bowel disease. Nature 491, 119–124 (2012)
Wu, C. et al. Induction of pathogenic TH17 cells by inducible salt-sensing kinase SGK1. Nature http://dx.doi.org/10.1038/nature11984 (2013)
Kim, U. et al. The B-cell-specific transcription coactivator OCA-B/OBF-1/Bob-1 is essential for normal production of immunoglobulin isotypes. Nature 383, 542–547 (1996)
Wang, V. E., Tantin, D., Chen, J. & Sharp, P. A. B cell development and immunoglobulin transcription in Oct-1-deficient mice. Proc. Natl Acad. Sci. USA 101, 2005–2010 (2004)
Bettelli, E. et al. Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells. Nature 441, 235–238 (2006)
Awasthi, A. et al. A dominant function for interleukin 27 in generating interleukin 10-producing anti-inflammatory T cells. Nature Immunol. 8, 1380–1389 (2007)
Awasthi, A. et al. Cutting edge: IL-23 receptor gfp reporter mice reveal distinct populations of IL-17-producing cells. J. Immunol. 182, 5904–5908 (2009)
Wei, G. et al. Global mapping of H3K4me3 and H3K27me3 reveals specificity and plasticity in lineage fate determination of differentiating CD4+ T cells. Immunity 30, 155–167 (2009)
Reich, M. et al. GenePattern 2.0. Nature Genet. 38, 500–501 (2006)
Storey, J., Xiao, W., Leek, J., Tompkins, R. & Davis, R. Significance analysis of time course microarray experiments. Proc. Natl Acad. Sci. USA 102, 12837–12482 (2005)
Leek, J. T., Monsen, E., Dabney, A. R. & Storey, J. D. EDGE: extraction and analysis of differential gene expression. Bioinformatics 22, 507–508 (2006)
Chechik, G. & Koller, D. Timing of gene expression responses to environmental changes. J. Comput. Biol. 16, 279–290 (2009)
Schraml, B. U. et al. The AP-1 transcription factor Batf controls TH17 differentiation. Nature 460, 405–409 (2009)
Shi, L. Z. et al. HIF1α-dependent glycolytic pathway orchestrates a metabolic checkpoint for the differentiation of TH17 and Treg cells. J. Exp. Med. 208, 1367–1376 (2011)
Durant, L. et al. Diverse targets of the transcription factor STAT3 contribute to T cell pathogenicity and homeostasis. Immunity 32, 605–615 (2010)
Jux, B., Kadow, S. & Esser, C. Langerhans cell maturation and contact hypersensitivity are impaired in aryl hydrocarbon receptor-null mice. J. Immunol. 182, 6709–6717 (2009)
Xiao, S. et al. Retinoic acid increases Foxp3+ regulatory T cells and inhibits development of Th17 cells by enhancing TGF-β-driven Smad3 signaling and inhibiting IL-6 and IL-23 receptor expression. J. Immunol. 181, 2277–2284 (2008)
Wilkins, M. R. et al. Protein identification and analysis tools in the ExPASy server. Methods Mol. Biol. 112, 531–552 (1999)
Durant, L. et al. Diverse targets of the transcription factor STAT3 contribute to T cell pathogenicity and homeostasis. Immunity 32, 605–615 (2010)
Huh, J. R. et al. Digoxin and its derivatives suppress TH17 cell differentiation by antagonizing RORγt activity. Nature 472, 486–490 (2011)
Sundrud, M. S. et al. Halofuginone inhibits TH17 cell differentiation by activating the amino acid starvation response. Science 324, 1334–1338 (2009)
Chevrier, N. et al. Systematic discovery of TLR signaling components delineates viral-sensing circuits. Cell 147, 853–867 (2011)
Geiss, G. K. et al. Direct multiplexed measurement of gene expression with color-coded probe pairs. Nature Biotechnol. 26, 317–325 (2008)
Ram, O. et al. Combinatorial patterning of chromatin regulators uncovered by genome-wide location analysis in human cells. Cell 147, 1628–1639 (2011)
Acknowledgements
We thank L. Gaffney and L. Solomon for artwork, the Broad’s Genomics Platform for sequencing, and D. Kozoriz for cell sorting. Work was supported by NHGRI (1P50HG006193-01 to H.P. and A.R.), NIH Pioneer Awards (5DP1OD003893-03 to H.P., DP1OD003958-01 to A.R.), NIH (NS 30843, NS045937, AI073748 and AI45757 to V.K.K.), National MS Society (RG2571 to V.K.K.), HHMI (A.R.), and the Klarman Cell Observatory (A.R.).
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N.Y., A.K.S., J.T.G., H.P., V.K.K. and A.R. conceived the study and designed experiments. N.Y. developed computational methods. N.Y., A.K.S. and J.T.G. analysed the data. A.K.S., J.T.G., H.J., Y.L., A.A., C.W., K.K., S.X., M.J., D.G., R.S., D.Y.L. and J.J.T. conducted the experiments. A.S., M.R.P., P.J.R., M.L.C., M.B. and D.T. provided knockout mice. N.Y., A.K.S., J.T.G., V.K.K., H.P. and A.R. wrote the paper with input from all the authors.
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Supplementary information
Supplementary Information
This file contains Supplementary Notes, Supplementary Methods, Supplementary Figures 1-12, legends for Supplementary Tables 1-8 (see separate excel files) and Supplementary References. (PDF 1823 kb)
Supplementary Table 1
This file contains a list of microarray probesets that were differentially expressed in the TGF-β1+Il6 microarray data - see Supplementary Information file for full legend. (XLS 323 kb)
Supplementary Table 2
This file shows the functional enrichments for expression clusters -see Supplementary Information file for full legend. (XLS 1034 kb)
Supplementary Table 3
This file shows the regulatory interactions in the three canonical temporal networks (Early, Intermediate, and Late) - see Supplementary Information file for full legend. (XLS 2258 kb)
Supplementary Table 4
This file contains a table of ranked regulators of Th17 differentiation - see Supplementary Information file for full legend. (XLS 80 kb)
Supplementary Table 5
This file contains the results of Nanostring nCounter and Fluidigm analysis - see Supplementary Information file for full legend. (XLS 191 kb)
Supplementary Table 6
This file contains the Primers for Nanostring STA and qRT-PCR/Fluidigm and siRNA sequences - see Supplementary Information file for full legend. (XLS 153 kb)
Supplementary Table 7
This file shows the RNA-seq data analysis - see Supplementary Information file for full legend. (XLS 1539 kb)
Supplementary Table 8
This file shows the Tsc22d3 ChIP-seq data analysis - see Supplementary Information file for full legend. (XLS 108 kb)
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Yosef, N., Shalek, A., Gaublomme, J. et al. Dynamic regulatory network controlling TH17 cell differentiation. Nature 496, 461–468 (2013). https://doi.org/10.1038/nature11981
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DOI: https://doi.org/10.1038/nature11981
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