Science Immunology

Supplementary Materials

The PDF file includes:

  • Materials and Methods
  • Fig. S1. Gating strategy and purity assessment for sorted naïve and Treg cells.
  • Fig. S2. Definition of the Treg transcriptional signature.
  • Fig. S3. Assessing the enrichment of Treg up-regulated or down-regulated genes in fetal and adult iTreg populations.
  • Fig. S4. Fetal iTreg cells have increased sensitivity to TGF-β signaling.
  • Fig. S5. Identification of Treg-accessible and -inaccessible enhancers.
  • Fig. S6. Binding motifs for downstream effectors of Treg differentiation are enriched within shared Treg-accessible peaks in fetal naïve T cells.
  • Fig. S7. The highest ranked SEs shared across all cell populations are associated with T cell development and function.
  • Fig. S8. Chromatin accessibility and H3K27ac enrichment at the Helios locus in fetal naïve T cells correlate with increased RNA and protein expression.
  • Fig. S9. Fetal naïve T cells do not have an increased proportion of CD31+ cells relative to adult naïve T cells.
  • Fig. S10. A fraction of fetal naïve T cells are highly proliferative.
  • Fig. S11. Fetal naïve T cells do not have demethylation at the FOXP3 conserved noncoding sequence 2 TSDR.
  • Fig. S12. Fetal naïve T cells up-regulate Helios during Treg induction.
  • Fig. S13. Validation of CRISPR-Cas9 editing at the Helios locus.
  • Fig. S14. The effect of CRISPR-Cas9 knockout of Helios on protein expression of Treg functional markers is variable.
  • Fig. S15. Fetal, but not adult, iTreg cells have suppressed IL-2 production after restimulation.
  • Fig. S16. Helios knockout in fetal iTreg cells result in a subtle shift in the underlying transcriptome.
  • Table S8. Experimental setup for Treg induction time course carried out for adult and fetal naïve T cells.
  • Table S9. Experimental setup for Helios CRISPR-Cas9–mediated editing for subsequent Treg induction carried out for fetal naïve T cells.
  • References (95105)

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Other Supplementary Material for this manuscript includes the following:

  • Table S1. RNA-seq Treg up-regulated and down-regulated signature genes (Excel spreadsheet).
  • Table S2. Genes up-regulated/down-regulated within iTreg populations with cluster assignment (Excel spreadsheet).
  • Table S3. Statistics from pre-ranked GSEA for fetal versus adult iTreg populations (Excel spreadsheet).
  • Table S4. Treg-accessible enhancers (Excel spreadsheet).
  • Table S5. Treg-inaccessible enhancers (Excel spreadsheet).
  • Table S6. De novo motif enrichment in Treg-accessible ATAC peaks shared between fetal naïve and adult Treg cells (Excel spreadsheet).
  • Table S7. Top 25 ranked SEs in adult naïve, fetal naïve, and adult Treg cells (Excel spreadsheet).
  • Table S10. Up-regulated and down-regulated differentially expressed genes defined in Helios knockout versus Helios wild-type fetal iTreg cells (Excel spreadsheet).
  • Table S11. Raw data file (Excel spreadsheet).

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