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Author Notes:

Address correspondence to: William A. Faubion Jr, MD, Division of Gastroenterology and Hepatology, Mayo Clinic, 200 First Street SW, Guggenheim Building, Room 10-24, Rochester, Minnesota 55905. fax: (507) 255-6318. faubion.william@mayo.edu

Adebowale O. Bamidele was responsible for the study concept and design, data acquisition, data analysis and interpretation, and manuscript preparation; Phyllis A. Svingen was responsible for data acquisition and manuscript preparation and revision; Michelle Gonzalez, Mary R. Sagstetter, Olga F. Sarmento, and Manuel B. Braga Neto acquired and interpreted data; Raul A. Urrutia, Subra Kugathasan, and Gwen Lomberk interpreted data and prepared the manuscript; and William A. Faubion Jr was responsible for study supervision, the study concept and design, data analysis and interpretation, manuscript preparation, and acquired funding.

The authors thank the Mayo Clinic Flow Cytometry Core Facility for expert and technical assistance and Dr Laura Raffals and Jessica Friton for assistance in obtaining human biopsy specimens.

The authors disclose no conflicts.


Research Funding:

This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases training grant T32DK007198 and the Pilot and Feasibility Award by the Center for Cell Signaling in Gastroenterology (P30DK084567) (A.O.B.), the National Institutes of HealthNational Institute of Diabetes and Digestive and Kidney Diseases RO1 grant DK52913 (R.A.U.), and the National Institute of Allergy and Infectious Diseases RO1 grant AI089714 and the Leona Helmsley Charitable Trust (W.A.F.).


  • C232, cysteine 232
  • CD, Crohn’s disease
  • ChIP, chromatin-immunoprecipitation
  • Crohn’s Disease
  • EED, embryonic ectoderm development
  • EZH2, enhancer of zeste homolog 2
  • Epigenetics
  • FCS, fetal calf serum
  • FOXP3, forkhead domain-containing X-chromosome–encoded protein
  • H3K27me3, trimethylated histone H3 at lysine 27
  • IBD, inflammatory bowel disease
  • IL, interleukin
  • IPEX, immune dysregulation, polyendocrinopathy, enteropathy, X-linked
  • JAK, Janus kinase
  • LZ, leucine zipper
  • PBMC, peripheral blood mononuclear cell
  • PBS, phosphate-buffered saline
  • PLA, proximity ligation assay
  • PMA, phorbol 12-myristate 13-acetate
  • PRC2, polycomb repressive complex 2
  • Proinflammatory Cytokine
  • Regulatory T Cells
  • STAT, signal transducer and activator of transcription
  • SUZ12, suppressor of zeste
  • Th, T helper
  • Treg, regulatory T cell
  • WT, wild-type
  • co-IP, co-immunoprecipitation

Disruption of FOXP3–EZH2 Interaction Represents a Pathobiological Mechanism in Intestinal Inflammation


Journal Title:

Cellular and Molecular Gastroenterology and Hepatology


Volume 7, Number 1


, Pages 55-71

Type of Work:

Article | Final Publisher PDF


Background & Aims: Forkhead box protein 3 (FOXP3)+ regulatory T cell (Treg) dysfunction is associated with autoimmune diseases; however, the mechanisms responsible for inflammatory bowel disease pathophysiology are poorly understood. Here, we tested the hypothesis that a physical interaction between transcription factor FOXP3 and the epigenetic enzyme enhancer of zeste homolog 2 (EZH2) is essential for gene co-repressive function. Methods: Human FOXP3 mutations clinically relevant to intestinal inflammation were generated by site-directed mutagenesis. T lymphocytes were isolated from mice, human blood, and lamina propria of Crohn's disease (CD) patients and non-CD controls. We performed proximity ligation or a co-immunoprecipitation assay in FOXP3-mutant+, interleukin 6 (IL6)-treated or CD-CD4+ T cells to assess FOXP3–EZH2 protein interaction. We studied IL2 promoter activity and chromatin state of the interferon γ locus via luciferase reporter and chromatin-immunoprecipitation assays, respectively, in cells expressing FOXP3 mutants. Results: EZH2 binding was abrogated by inflammatory bowel disease–associated FOXP3 cysteine 232 (C232) mutation. The C232 mutant showed impaired repression of IL2 and diminished EZH2-mediated trimethylation of histone 3 at lysine 27 on interferon γ, indicative of compromised Treg physiologic function. Generalizing this mechanism, IL6 impaired FOXP3–EZH2 interaction. IL6-induced effects were reversed by Janus kinase 1/2 inhibition. In lamina propria–derived CD4+T cells from CD patients, we observed decreased FOXP3–EZH2 interaction. Conclusions: FOXP3–C232 mutation disrupts EZH2 recruitment and gene co-repressive function. The proinflammatory cytokine IL6 abrogates FOXP3–EZH2 interaction. Studies in lesion-derived CD4+ T cells have shown that reduced FOXP3–EZH2 interaction is a molecular feature of CD patients. Destabilized FOXP3–EZH2 protein interaction via diverse mechanisms and consequent Treg abnormality may drive gastrointestinal inflammation.

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© 2018 The Authors

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