Fig. 2: RASA2 ablation promotes T cell activation, antigen sensitivity and effector function.
From: RASA2 ablation in T cells boosts antigen sensitivity and long-term function
a, RAS signaling and downstream transcriptional programmes in T cells. Drawing is adapted from ref. 29. IKK, inhibitor of NF-κB kinase. b, Western blot showing RASA2 protein expression in Jurkat cells and GTP-bound active RAS after TCR stimulation. c, Flow cytometry-based analysis of phospho-ERK kinetics in stimulated primary human T cells. d, Scaled phosphoprotein mean fluorescence intensity (MFI) in MAPK and AKT–mTOR pathways. e, Effector cytokine levels in stimulated T cells. f,g, pERK levels 10 min after TCR stimulation with anti-CD3/CD28 (f) or T2 cells preloaded with cognate peptide (g). h, Left, CD19 expression on engineered Nalm6 cancer target cells (green) compared with unstained cells (grey). Right, CAR T cell killing of Nalm6 cells expressing varying CD19 levels, measured by annexin staining. Data are mean ± s.d. of technical triplicates from one representative donor out of two. WT, wild type. i, Percentage of Jurkat cells positive for transcription factor-responsive mCherry reporters. j, GSEA of differentially expressed genes between RASA2-KO and control cells after TCR stimulation. Dot size represents adjusted P-value (Padj; two-sided permutation test). NES, normalized enrichment score; phospho, phosphorylation; resp., response. k, Differentially expressed genes in stimulated RASA2-KO T cells with perturbation of the indicated target genes13. Colour indicates mean expression level and size shows the percentage of cells with detectable expression (n = 2 donors). l–o, RASA2 expression in a mouse model of Listeria infection38 (l; n = 3 mice; mean ± s.e.m.), in vitro activated human T cells20 (m; n = 91 donors; two-sided Wilcoxon test), a mouse model of tumour-infiltrating T cells38 (TIL) (n, showing days after T cell transfer; n = 3 mice; mean ± s.e.m.) and human tumour-infiltrating T cells (orange) or peripheral T cells (green) (o). o, Box limits show quartiles, the horizontal line is the median (n = 12 donors for colorectal cancer40 (CRC) and n = 14 donors for non-small cell lung carcinoma41 (NSCLC); two-sided Wilcoxon test). c–e, Lines show mean; n = 2 donors in triplicate; two-sided Wilcoxon test. f,g, n = 2 donors in triplicate; fitted 4-parameter dose–response curves; two-sample Kolmogorov–Smirnov test. *P < 0.05, **P < 0.01, ****P < 0.0001.
