An N-terminal mutation of the Foxp3 transcription factor alleviates arthritis but exacerbates diabetes - PubMed (original) (raw)
An N-terminal mutation of the Foxp3 transcription factor alleviates arthritis but exacerbates diabetes
Jaime Darce et al. Immunity. 2012.
Abstract
Maintenance of lymphoid homeostasis in a number of immunological and inflammatory contexts is served by a variety of regulatory T (Treg) cell subtypes and depends on interaction of the transcription factor FoxP3 with specific transcriptional cofactors. We report that a commonly used insertional mutant of FoxP3 (GFP-Foxp3) modified its molecular interactions, blocking HIF-1α but increasing IRF4 interactions. The transcriptional profile of these Treg cells was subtly altered, with an overrepresentation of IRF4-dependent transcripts. In keeping with IRF4-dependent function of Treg cells to preferentially suppress T cell help to B cells and Th2 and Th17 cell-type differentiation, GFP-FoxP3 mice showed a divergent susceptibility to autoimmune disease: protection against antibody-mediated arthritis in the K/BxN model, but greater susceptibility to diabetes on the NOD background. Thus, specific subfunctions of Treg cells and the immune diseases they regulate can be influenced by FoxP3's molecular interactions, which result in divergent immunoregulation.
Copyright © 2012 Elsevier Inc. All rights reserved.
Figures
Figure 1. Foxp3fgfp mice exhibit contrasting phenotypes in different autoimmune models
(A) Schematic representation of FoxP3, GFP-FoxP3, and FoxP3-I-GFP. Protein convertase cleavage sites (RXXR), proline rich regions (Pxxp1 and Pxxp2), leucine zipper (Leu-zip) and forkhead (FKH) domains are represented. (B) Arthritis clinical score and ankle measurements of 8 week-old K/BxN.Foxp3fgfp males, K/BxN.Foxp3fgfp/wt heterozygote females and WT littermate control mice. (C) Anti-GPI IgG reactivity in sera of 8 week-old K/BxN.Foxp3fgfp males, K/BxN.Foxp3fgfp/wt females and WT littermate control mice (mean +/− SD, n=6–8). (D) Arthritis clinical score, ankle thickness, and anti-GPI antibodies in 8 week-old male and female K/BxN.Foxp3igfp and WT controls. (E) Diabetes incidence curves of NOD.Foxp3fgfp males NOD.Foxp3fgfp/wt, females and littermate controls (10 mice/group). Two different cohorts of NOD.Foxp3fgfp/wt heterozygous females were followed, in two different animal facilities.
Figure 2. Foxp3fgfp disease phenotype does not correlate with an alteration in Treg frequency
(A and B) Percentage of FoxP3 positive cells within CD4+ T cells. (A) Spleens and draining lymph nodes (popliteal) of 8 week-old WT and K/BxN.Foxp3fgfp males (each point is an individual mouse). (B) Spleen and pancreas of 9 week-old NOD.Foxp3fgfp males and controls. (C) Proportion of FoxP3+ Treg cells among LN and visceral adipose tissue of 40 week-old B6 males. (D) Representative dot plots showing the percent FoxP3+ Helios− Treg cells in colon lamina propria and spleen of 6 week-old B6.Foxp3fgfp and B6.Foxp3igfp males. Splenocytes from K/BxN.Foxp3fgfp/wt heterozygote females were stained with anti-CD3, -CD4, and –FoxP3 antibodies. (E) Gates and values represent the fraction of GFP positive and negative cells among FoxP3+ population. Summary of data in adjacent plot shows fraction of GFP+ cells among FoxP3+ cells in thymus, spleen and draining lymph nodes of K/BxN.Foxp3fgfp/wt females; the dashed line denotes the 50/50 ratio expected from random X-inactivation (F) Linear regression analysis of serum anti-GPI levels vs the fraction of GFP+ FoxP3+ Treg cells in K/BxN.Foxp3fgfp/wt heterozygote females.
Figure 3. Decrease in Th17 cells in K.BxN.Foxp3fgfp is independent of inherent Th17 differentiation defect
(A) IL17 producing cells in spleen and small intestine lamina propria from 4–5 week old K/BxN and K/BxN.Foxp3fgfp male mice. Gates and values represent fraction of IL17 positive cells in CD4 population. Data is summarized in adjacent graph. Data is representative of 3 independent experiments. (B) In vitro Th17/Treg differentiation of congenically marked naïve cells from WT (CD45.1) and Foxp3fgfp B6 mice. Data is representative of 2 independent experiments.
Figure 4. Elevated FoxP3 expression in Foxp3fgfp Treg cells
(A) Anti-FoxP3 staining profiles of CD3+CD4+ from different organs of K/BxN.Foxp3fgfp male or WT littermates (shaded area is profile from isotype control). Data representative of 3 independent experiments. (B) FoxP3 mean fluorescence intensity (MFI) of splenic Treg cells on 3 different backgrounds. Each dot represents an individual mouse, and values are normalized to the mean FoxP3 MFI of WT mice in each experiment. (C) FoxP3 MFI for LN and visceral fat from 40 week-old B6.Foxp3fgfp mice. (D) Foxp3 mRNA levels in CD4+GFP+ splenic Treg cells from Foxp3fgfp or WT male mice on 2 different backgrounds.
Figure 5. Enhanced Treg function observed in Foxp3fgfp Treg cells
(A) In vitro suppressive ability of CD4+CD25+ Treg cells from B6.Foxp3fgfp or WT littermates, titrated at different ratios in co-cultures with CD4+ Tconv responder cells and APCs. Data represent mean proliferation +/− SD of 3 independent experiments. (B) Th17 differentiation suppressed by GFP-FoxP3 (FGFP) and FoxP3-ires-GFP (IGFP) Tregs. Ifng, Il17a, Il4 mRNA expression level assessed by rt-PCR.
Figure 6. Biased gene expression in Foxp3fgfp Treg cells
Gene expression profiles were generated by microarray of Treg and Tconv cells from B6.Foxp3fgfp males, with parallel profiling from B6.Foxp3igfp as a reference. (A) Comparison of expression values in Treg versus Tconv splenocytes from Foxp3fgfp and Foxp3igfp B6 males. The canonical Treg signature is highlighted in red (Treg up-regulated transcripts) and blue (Treg down-regulated transcripts). (B) Comparison of expression values in Foxp3fgfp and control Foxp3igfp Treg splenocytes, on B6 and NOD backgrounds. Genes concordantly over-expressed in Foxp3fgfp relative to Foxp3igfp in both backgrounds are highlighted in red. Transcription factor genes are represented in green. (C) Heatmap representation of transcripts over-represented in Foxp3fgfp on both backgrounds. (D) Flow cytometric confirmation of CTLA-4, CD103, and KLRG1 over-expression in CD4+FoxP3+ Treg cells from spleens of Foxp3fgfp heterozygote females, on the K/BxN and NOD backgrounds. The values shown are the percent CD103 or KLRG1 positive cells within the GFP-positive or -negative fraction of Treg cells (mean +/− SD of 8 mice). (E) Over-representation of the Irf4 Treg signature: The ratio of expression in Foxp3fgfp vs Foxp3igfp of Irf4-responsive transcripts is shown (log2 scale) for both B6 and NOD datasets. The p-value is calculated with a χ2 test for departure from a null hypothesis of random distribution.
Figure 7. GFP-FoxP3 differentially binds transcriptional co-factors
(A) Association of FoxP3 with FoxP1 determined by co-immunoprecipitation. Anti-FoxP3 was used to immunoprecipitate FoxP3 from nuclear lysates of CD4+CD25− Tconv or CD4+CD25+ Treg cells from B6.Foxp3fgfp or WT mice, and immunoblots were probed for FoxP1 (anti-FoxP3 as control). (B) Nuclear lysates of B6.Foxp3fgfp and B6.Foxp3igfp CD25+ Treg cells were immunoprecipitated with anti-FoxP3 or control IgG, and probed with anti-Hif1a (anti-FoxP3 and anti-β–actin as controls). (C) Anti-Irf4 was used to immunoprecipitate Irf4 from nuclear lysates CD4+CD25+ Treg cells from B6.Foxp3fgfp or WT mice, and immunobloted for FoxP3. (D) Western blot analysis of Irf4 from nuclear lysates of CD4+CD25+ Treg cells from B6.Foxp3fgfp or WT mice, two independent experiments shown.
Comment in
- Foxp3: shades of tolerance.
Chatila TA, Williams CB. Chatila TA, et al. Immunity. 2012 May 25;36(5):693-4. doi: 10.1016/j.immuni.2012.05.011. Immunity. 2012. PMID: 22633453 Free PMC article.
Similar articles
- Loss of epigenetic modification driven by the Foxp3 transcription factor leads to regulatory T cell insufficiency.
Bettini ML, Pan F, Bettini M, Finkelstein D, Rehg JE, Floess S, Bell BD, Ziegler SF, Huehn J, Pardoll DM, Vignali DA. Bettini ML, et al. Immunity. 2012 May 25;36(5):717-30. doi: 10.1016/j.immuni.2012.03.020. Epub 2012 May 10. Immunity. 2012. PMID: 22579476 Free PMC article. - A Mutation in the Transcription Factor Foxp3 Drives T Helper 2 Effector Function in Regulatory T Cells.
Van Gool F, Nguyen MLT, Mumbach MR, Satpathy AT, Rosenthal WL, Giacometti S, Le DT, Liu W, Brusko TM, Anderson MS, Rudensky AY, Marson A, Chang HY, Bluestone JA. Van Gool F, et al. Immunity. 2019 Feb 19;50(2):362-377.e6. doi: 10.1016/j.immuni.2018.12.016. Epub 2019 Jan 29. Immunity. 2019. PMID: 30709738 Free PMC article. - Thymically-derived Foxp3+ regulatory T cells are the primary regulators of type 1 diabetes in the non-obese diabetic mouse model.
Holohan DR, Van Gool F, Bluestone JA. Holohan DR, et al. PLoS One. 2019 Oct 24;14(10):e0217728. doi: 10.1371/journal.pone.0217728. eCollection 2019. PLoS One. 2019. PMID: 31647813 Free PMC article. - Regulatory T cell identity: formation and maintenance.
Li X, Zheng Y. Li X, et al. Trends Immunol. 2015 Jun;36(6):344-53. doi: 10.1016/j.it.2015.04.006. Epub 2015 May 13. Trends Immunol. 2015. PMID: 25981968 Free PMC article. Review. - Control of regulatory T-cell differentiation and function by T-cell receptor signalling and Foxp3 transcription factor complexes.
Ono M. Ono M. Immunology. 2020 May;160(1):24-37. doi: 10.1111/imm.13178. Epub 2020 Mar 9. Immunology. 2020. PMID: 32022254 Free PMC article. Review.
Cited by
- Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells.
Liu Z, Baines KJ, Niessen NM, Heer MK, Clark D, Bishop GA, Trevillian PR. Liu Z, et al. Front Immunol. 2024 Jan 25;15:1292158. doi: 10.3389/fimmu.2024.1292158. eCollection 2024. Front Immunol. 2024. PMID: 38333213 Free PMC article. - Immune changes in hilar tumor draining lymph nodes following node sparing neoadjuvant chemoradiotherapy of localized cN0 non-small cell lung cancer.
Khalifa J, Thébault N, Scarlata CM, Norkowski E, Massabeau C, Brouchet L, Peries Bataille S, Casaroli C, Vaz L, Valle C, Sarot E, Saint-Laurent N, Martin E, Pages PB, Millière A, Mazières J, Cohen-Jonathan Moyal E, Lauzéral-Vizcaïno F, Ayyoub M. Khalifa J, et al. Front Oncol. 2023 Nov 22;13:1269166. doi: 10.3389/fonc.2023.1269166. eCollection 2023. Front Oncol. 2023. PMID: 38074683 Free PMC article. - Regulatory T cells in inflamed liver are dysfunctional in murine primary biliary cholangitis.
Lin CI, Wang YW, Liu CY, Chen HW, Liang PH, Chuang YH. Lin CI, et al. Clin Exp Immunol. 2024 Feb 19;215(3):225-239. doi: 10.1093/cei/uxad117. Clin Exp Immunol. 2024. PMID: 37916967 - Mutations from patients with IPEX ported to mice reveal different patterns of FoxP3 and Treg dysfunction.
Leon J, Chowdhary K, Zhang W, Ramirez RN, André I, Hur S, Mathis D, Benoist C. Leon J, et al. Cell Rep. 2023 Aug 29;42(8):113018. doi: 10.1016/j.celrep.2023.113018. Epub 2023 Aug 21. Cell Rep. 2023. PMID: 37605532 Free PMC article. - Editorial: Regulatory T lymphocytes in cancer immunity.
Adeegbe D, Barbi J, Wing J. Adeegbe D, et al. Front Immunol. 2022 Oct 24;13:1065570. doi: 10.3389/fimmu.2022.1065570. eCollection 2022. Front Immunol. 2022. PMID: 36353629 Free PMC article. No abstract available.
References
- Barnes MJ, Powrie F. Regulatory T cells reinforce intestinal homeostasis. Immunity. 2009;31:401–411. - PubMed
- Bettelli E, Carrier Y, Gao W, Korn T, Strom TB, Oukka M, Weiner HL, Kuchroo VK. Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells. Nature. 2006;441:235–238. - PubMed
Publication types
MeSH terms
Substances
Grants and funding
- AI051530/AI/NIAID NIH HHS/United States
- R37 AI051530/AI/NIAID NIH HHS/United States
- HHMI/Howard Hughes Medical Institute/United States
- R01 DK092541/DK/NIDDK NIH HHS/United States
- DK092541/DK/NIDDK NIH HHS/United States
- R01 AI051530/AI/NIAID NIH HHS/United States
- AI065858/AI/NIAID NIH HHS/United States
- P01 AI065858/AI/NIAID NIH HHS/United States
- T32CA007386/CA/NCI NIH HHS/United States
LinkOut - more resources
Full Text Sources
Other Literature Sources
Medical
Molecular Biology Databases