The kinase DYRK1A reciprocally regulates the differentiation of Th17 and regulatory T cells
- Massachusetts General Hospital, Harvard Medical School, United States
- Broad Institute of MIT and Harvard, United States
- Massachusetts General Hospital, United States
- Harvard University, United States
- Harvard Medical School, United States
- National Institutes of Health, United States
Figures
Figure 1 with 8 supplements
Chemical biology approach to identify novel Treg enhancers.
All data representative of at least 2 independent experiments. (A) Overview of our approach, including key methods applied. (B) Dose-response curves showing fractional enhancement (Fr enhance) of …
Figure 1—figure supplement 1
Titrating Th differentiation conditions.
Titrating cytokines for Treg, Th1, and Th17 conditions identifies sub-maximal (blue arrowheads) and near-maximal (red arrowheads) lineage-promoting conditions.
Figure 1—figure supplement 2
Culture cellularity affects Treg differentiation.
Treglow (blue) stimulation with varying initial cell numbers recapitulates the inverse relationship between final culture cellularity and percentage of FOXP3-expressing cells. Treghi (red) …
Figure 1—figure supplement 3
Schematic of analytic and hit-calling pipeline.
https://doi.org/10.7554/eLife.05920.006
Figure 1—figure supplement 4
Effect of compounds (LD50/EC50 < 2) on Th differentiation.
Dose-response curves showing compound effect on fractional enhancement of Treg (blue), Th1 (orange), and Th17 (red) lineages.
Figure 1—figure supplement 5
Modeling analyses to calculate EC50 values, indicated in parentheses, for all 21 Treg-specific enhancers.
https://doi.org/10.7554/eLife.05920.008
Figure 1—figure supplement 6
Modeling analyses to calculate LD50 values, indicated in parentheses, for all 21 Treg-specific enhancers.
https://doi.org/10.7554/eLife.05920.009
Figure 1—figure supplement 7
Similarity clustering analysis of combined Treg, Th1 and Th17 phenotypic data.
https://doi.org/10.7554/eLife.05920.010
Figure 1—figure supplement 8
Euclidean distance clustering analysis of gene expression data from cell lines treated with Treg enhancers first analyzed by principal component analysis.
https://doi.org/10.7554/eLife.05920.011
Figure 2 with 2 supplements
Harmine-enhanced Treg cells and harmine attenuate inflammation.
(A–E) Experiments comparing the suppressive activity of Treg cells generated under either TregHAR (blue) or Treghi (red) conditions; conditions without Treg cells are shown in black. All data …
Figure 2—figure supplement 1
Effects of harmine treatment in vivo on T cell populations.
Mice were administered either intranasal harmine HCl (blue) or water (gray). T cell populations in thoracic lymph nodes were quantitated as shown, demonstrating relative percentages (above) and …
Figure 2—figure supplement 2
Effects of harmine treatment in vivo on antigen-presenting cell populations.
Mice were administered either intranasal harmine HCl (blue) or water (gray). Migratory and classical dendritic cells (mDC and cDC, respectively) in thoracic lymph nodes were quantitated (top right), …
Figure 3 with 5 supplements
Harmine's effects on canonical Treg/Th17 pathways.
All data representative of at least 2 independent experiments. (A) Effect of harmine on murine Treg, Th1 and Th17 differentiation. (B) Effect of harmine on absolute numbers of total live and Treg …
Figure 3—figure supplement 1
Effect of harmine on cellular proliferation.
CFSE-labelled CD4+ T cells were stimulated under Treglow conditions and proliferation assessed daily. The effect of adding harmine, rapamycin, or high TGFβ (Treghi) was compared as shown.
Figure 3—figure supplement 2
Effect of harmine on absolute numbers of Th17 cells.
***p < 0.001, Student's t-test.
Figure 3—figure supplement 3
Effect of addition or removal of harmine at different times on Th17 differentiation.
https://doi.org/10.7554/eLife.05920.018
Figure 3—figure supplement 4
Correlation between gene expression in Treghi-Treg cells and TregHAR-Treg cells, relative to naïve CD4+ T cells.
https://doi.org/10.7554/eLife.05920.019
Figure 3—figure supplement 5
Qualitative analyses of genomewide expression in TregHAR-Treg cells.
Previously described methods were used to identify similarities between TregHAR-Treg cells and other specialized Treg subsets (Joller et al., 2014). Volcano plots compare relative expression of …
Figure 4 with 3 supplements
Mechanistic dissection of harmine.
(A) Comparison of expression profiles suggesting a harmine-relevant Treg signature. The black bar separates 2 independently row-normalized experiments. (B) Harmine signature genes are enriched for …
Figure 4—figure supplement 1
Effect of harmine on NFAT1 nuclear localization with time.
Time-course Amnis analyses showing effect of harmine (blue) relative to control Treglow conditions (black).
Figure 4—figure supplement 2
Comparing effects of DYRK1A deficiency to harmine treatment.
RNAseq experiments in primary CD4+ T cells comparing the effects of transduction with Dyrk1a shRNA (shDyrk1a) to harmine treatment in control shRNA-transduced cells (left). Further statistical …
Figure 4—figure supplement 3
Secondary analyses of effects of DYRK1A deficiency compared to harmine treatment.
Volcano plots comparing p value vs fold change in gene expression in harmine-treated (left panels) and Dyrk1a shRNA-treated (right panels) cells. Previously reported signature genes for Treg cells …
Additional files
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Supplementary file 1
(A) Staining reagents used. (B) Chemicals used.
- https://doi.org/10.7554/eLife.05920.025
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Supplementary file 2
Th conditions used.
- https://doi.org/10.7554/eLife.05920.026
