Extended Data Fig. 6: Pcif1 KO enhances CD8+ T cell activation. | Nature Immunology

Extended Data Fig. 6: Pcif1 KO enhances CD8+ T cell activation.

From: The epitranscriptional factor PCIF1 orchestrates CD8+ T cell ferroptosis and activation to control antitumor immunity

Extended Data Fig. 6

a, CD69+ percentages and MFI were quantified in WT or Pcif1 KO CD4+ T cells stimulated with anti-CD3/CD28 for 5 hours. n = 3 mice/each group. b, WB were showed the protein levels of CD69 in WT or Pcif1 KO CD8+ T cells stimulated with anti-CD3/CD28 for 5 hours. c-h, The percentages and MFI of cytokines in WT or Pcif1 KO CD8+ T cells were assessed after stimulation with anti-CD3/CD28 for 48 hours. n = 4 mice/each group. i,j, The secreted cytokines in the culture media of WT and Pcif1 KO CD8+ T cells were quantified using ELISAs. n = 3 mice/each group. k,l, The Ki67+ percentages were quantified in WT or Pcif1 KO CD8+ (k) or CD4+ (l) T cells stimulated with anti-CD3/CD28 for 72 hours. n = 4 mice/each group. m,n, EdU staining (m) and CellTrace dilution (n) in WT and Pcif1 KO CD8+ T cells. n = 4 mice/each group. o, WB showing the expression of p-Lck(Y394), Lck, p-Lat(Y220), and Lat in splenic naïve CD8+ T cells derived from the mice with subcutaneous LLC tumors. Total Lck, Lat and Vinculin proteins were used as loading controls. Right: Quantification of p-Lck(Y394)/Lck or p-Lat(Y220)/Lat. n = 3 mice/each group. p-r, MFI, RT-qPCR, and ELISA analysis of cytokines in Pcif1 KO CD8+ T cells retrovirally transduced with GFP, Pcif1 WT-GFP, or Pcif1 N552A-GFP restimulated with 2 μg/mL anti-CD3/CD28 for 48 hours. n = 3 mice/each group. s, Flow cytometry analysis of retrovirus-mediated expression of GFP, Pcif1-WT-GFP and Pcif1-N552A-GFP in Pcif1 KO CD8+ T cells. t-y, The percentages, MFI, RT-qPCR and ELISA analysis of cytokines in Pcif1 KO CD8+ T cells retrovirally transduced with GFP, Pcif1 WT-GFP, or Pcif1 N552A-GFP restimulated with 2 μg/mL anti-CD3/CD28 for 72 hours. n = 3 mice/each group. Each symbol represents an individual mouse. Two-tailed unpaired Student’s t-test (c-j,o) or two-way ANOVA followed by Sidak’s multiple comparisons test (a,k-n), two-way ANOVA followed by Tukey’s multiple comparisons test (p-r,t-y) were performed. Data are shown as the mean ± SEM.

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