Extended Data Fig. 8: PGE2-mediated changes in TCF1+CD8+ T cells, effect on γc expression and proliferation.
From: PGE2 limits effector expansion of tumour-infiltrating stem-like CD8+ T cells
a-d, scRNA-seq based analysis of TF activity alterations in TCF1+CD8+ TILs and their TIM-3+CXCR6+ effector progeny between control BRAFV600E melanoma tumours in Cd4crePtger2−/−Ptger4fl/fl and GzmbcrePtger2−/−Ptger4fl/fl mice compared to Ptger2−/−Ptger4fl/fl mice. a,b, Alterations of TF network activity in TCF1+CD8+ TILs from BRAFV600E melanoma tumours in a, Cd4crePtger2−/−Ptger4fl/fl mice compared to Ptger2−/−Ptger4fl/fl mice or b, GzmbcrePtger2−/−Ptger4fl/fl mice compared to Ptger2−/−Ptger4fl/fl mice. c, Heatmap visualisation, d, Correlation of TF activity alterations for TIL populations from Cd4crePtger2−/−Ptger4fl/fl and GzmbBcrePtger2−/−Ptger4fl/fl mice. e, Flow cytometry plots showing the sorting strategy for TCF1+ TILs (identified as TIM-3−CXCR6− cells) from Ptgs1/Ptgs2−/− BRAFV600E tumours. f, Volcano plot showing the effect of PGE2 exposure on gene expression in TCF1+CD8+ T cells from in vitro T cell cultures, based on RNA-seq analysis. g, Effect of PGE2 treatment on IL-2Rγc protein expression (n = 3). h, WT or Cd4crePtger2−/−Ptger4fl/fl TCF1+CD8+ T cells from in vitro T cell cultures were stimulated with anti-CD3/CD28 and high-dose IL-2 in presence or absence of PGE2. After 24 h cells were analysed for T cell proliferation by EdU incorporation. i, WT, Ptger2−/−Ptger4fl/fl or Cd4crePtger2−/−Ptger4fl/fl TCF1+CD8+ T cells were treated or not with tumour cell conditioned medium (CM) or PGE2 and stimulated as indicated. After 72 h, T cell expansion was analysed by flow cytometry. n = 3. Data in a-d, f are from one experiment. Data in g,i show data for one representative of two independent experiments with n = 3 biological replicates. In f), DEGs (P < 0.05; fold change ≥ 2) were identified by Wald test with multiple testing by the Benjamini-Hochberg method. Horizontal lines and error bars in (g,i) indicate mean ± s.d. P-values are from d, Fitted linear regression models or g, and i, unpaired t-test. P ≥ 0.05, not significant (NS). Plots in (h) show data for one sample representative for n = 3 samples.
