Extended Data Fig. 9: Effects of STAT5 activation on T cell exhaustion and stemness. | Nature

Extended Data Fig. 9: Effects of STAT5 activation on T cell exhaustion and stemness.

From: An engineered IL-2 partial agonist promotes CD8+ T cell stemness

Extended Data Fig. 9

a, ChIP-seq analysis of STAT5A- and STAT5B-binding sites at the Havcr2 locus from previously published datasets (GSE36890). b, c, TIM-3 expression on CD8+ T cells from Stat5a (b) and Stat5b (c) knock-out mice versus wild-type littermate controls. Cells were expanded for 8 days with 10 nM of IL-2 as described above, and TIM-3 expression analysed by flow cytometry. Data are mean ± SEM, n = 4 mice, two-sided t-test. Data are from two independent repeats. d, ChIP–seq analysis of STAT5-binding sites at HAVCR, GZMB, TCF7, SLC2A1 and SLC2A3 loci. Human CD8+ T cells were pre-activated with anti-CD3/anti-CD28 beads for 2 days, rested overnight, incubated with 10 nM IL-2 or H9T for 2 h, and then fixed and lysed for ChIP–seq. Data are from two independent experiments. eh, Three days after retroviral transduction, mouse CD8+ T cells expressing empty vector (EV)-GFP or STAT5A-1*6 vector-GFP were sorted and cultured in H9T-containing medium for an additional four days prior to staining with the indicated antibodies. Data are representative of two independent experiments. im, RNA-seq analysis of the effects of STAT5A-1*6 expression. Mouse CD8+ T cells were treated as above, and cells were collected for RNA-seq. PCA analysis (i) and expression of selected genes (j) are shown. GSEA analysis of IL-2-STAT5 signalling (k), PI3K-AKT-mTOR signalling (l) and exhaustion versus memory (m) are also shown, Kolmogorov–Smirnov test. Data are from two biological repeats. np, ATAC-seq analysis of the effect of STAT5A-1*6 expression. Mouse CD8+ T cells were treated as described above, and cells were collected for ATAC-seq. Shown are PCA analysis to compare IL-2-, H9-, and H9T-expanded cells (n), and ATAC-seq data at the Havcr2 (o) and Tcf7 (p) loci

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