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Transcription factor Foxp3 and its protein partners form a complex regulatory network

Nature Immunology volume 13, pages 1010–1019 (2012)Cite this article

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Abstract

The transcription factor Foxp3 is indispensible for the differentiation and function of regulatory T cells (Treg cells). To gain insights into the molecular mechanisms of Foxp3-mediated gene expression, we purified Foxp3 complexes and explored their composition. Biochemical and mass-spectrometric analyses revealed that Foxp3 forms multiprotein complexes of 400–800 kDa or larger and identified 361 associated proteins, ∼30% of which were transcription related. Foxp3 directly regulated expression of a large proportion of the genes encoding its cofactors. Some transcription factor partners of Foxp3 facilitated its expression. Functional analysis of the cooperation of Foxp3 with one such partner, GATA-3, provided additional evidence for a network of transcriptional regulation afforded by Foxp3 and its associates to control distinct aspects of Treg cell biology.

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Figure 1: Strategy for purification of Foxp3-associated proteins.
Figure 2: Foxp3 forms large protein complexes with its partners.
Figure 3: Functional annotation of Foxp3-associated proteins.
Figure 4: A large proportion of Foxp3-associated factors are also transcriptional targets of Foxp3.
Figure 5: Foxp3-Gata3 gene regulatory module in Treg cells.
Figure 6: A subset of genes regulated by Foxp3 and GATA-3 in Treg cells.

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Acknowledgements

We thank S. Lee, A. Bravo, J. Herlihy and J. Gerard for help with the mouse colony management and technical assistance; I.-C. Ho (Harvard Medical School) for Gata3fl/fl mice, D. Littman (New York University) for Runx1 antibody, I. Taniuchi (RIKEN Research Center for Allergy and Immunology) for Cbfβ andtibody and K. Georgopoulos (Massachusetts General Hospital) for Ikzf1 and Ikzf3 antibodies. This work was supported by US National Institutes of Health R37 AI034206 grant and the Howard Hughes Medical Institute (A.Y.R.). D.R. was supported by Arthritis Foundation postdoctoral fellowship. A.C. is supported by the Irvington Institute Fellowship Program of the Cancer Research Institute. R.E.N. is supported by National Institutes of Health Medical Scientist Training Program grant GM07739 and National Institute of Neurological Disorders and Stroke grant 1F31NS073203-01. R.M.S. is supported by National Institutes of Health DK091968 and Medical Scientist Training Program GM07739 grants.

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  1. Scott A Shaffer

    Present address: Present address: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.,

Authors and Affiliations

  1. Howard Hughes Medical Institute and Immunology Program, Sloan-Kettering Institute, New York, New York, USA

    Dipayan Rudra, Paul deRoos, Ashutosh Chaudhry, Rachel E Niec, Aaron Arvey, Robert M Samstein & Alexander Y Rudensky

  2. Computational Biology Program, Sloan-Kettering Institute, New York, New York, USA

    Aaron Arvey & Christina Leslie

  3. Department of Medicinal Chemistry, University of Washington, Seattle, Washington, USA

    Scott A Shaffer & David R Goodlett

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Contributions

D.R. designed and performed the majority of experiments, analyzed data and wrote the manuscript; P.d. performed chromatography and protein purifications, and was involved in mass-spectrometric analysis; A.C. was involved in co-immunoprecipitation studies; R.E.N. was involved in functional analysis of Gata3fl/fl Foxp3-YFP-Cre mice; R.M.S. performed Foxp3 ChIP-seq and gene-expression analyses in Treg and TFN cells; A.A. and C.L. performed computational and statistical analysis of ChIP-Seq and gene expression data sets; S.A.S. and D.R.G. assisted with mass-spectrometric analyses; A.Y.R. directed the project, was involved in design of experiments, data analysis and interpretation, and wrote the manuscript.

Corresponding author

Correspondence to Alexander Y Rudensky.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8, Supplementary Table 5 (PDF 1450 kb)

Supplementary Table 1

Foxp3-associated proteins in TCli cells and ex vivo isolated Treg cells and cumulative abundance of of Foxp3 associated proteins. (XLSX 67 kb)

Supplementary Table 2

GO term enrichment of the “molecular function” category of Foxp3-associated proteins and GO term enrichment of the “biological process” category of Foxp3-associated proteins. (XLSX 34 kb)

Supplementary Table 3

Foxp3 occupied regions within the gene loci encoding transcription-related Foxp3 interacting proteins. (XLSX 23 kb)

Supplementary Table 4

Genomic regions co-occupied by Foxp3 and Gata3. (XLSX 53 kb)

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Rudra, D., deRoos, P., Chaudhry, A. et al. Transcription factor Foxp3 and its protein partners form a complex regulatory network. Nat Immunol 13, 1010–1019 (2012). https://doi.org/10.1038/ni.2402

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