Figure 4. Nur77-GFP expression in naive CD8⁺ T cells during steady-state conditions correlates with gene expression changes.

(A) MA plot of DEGs between GFP^LO and GFP^HI naive OT-I CD8⁺ T cells. DEGs were defined as genes with an FDR < 0.05. Selected genes have been highlighted. The number of upregulated and downregulated genes in GFP^HI relative to GFP^LO cells are indicated in red and blue, respectively. (B) GSEA of genes downregulated in naive compared to effector CD8⁺ T cells (left panel) and genes upregulated in effector compared to resting memory CD8⁺ T cells (right panel) (Luckey et al., 2006). FDR values were derived from running GSEA on the c7_Immunesigdb.v2022.1 database. (C) Curated heatmaps of normalized expression of DEGs in indicated categories. (D) Histograms show expression of the indicated markers by GFP^LO and GFP^HI cells. The cells were gated on naive, polyclonal CD8⁺ T cells. Bar graphs depict gMFI of indicated proteins. (E) Flow cytometry plots of CD127 and CD200 expression in naive GFP^LO and GFP^HI naive, polyclonal CD8⁺ T cells as in D. (F) Flow cytometry plots (left, middle) show the gating scheme to identify CD127^HI CD200^LO and CD127^LO CD200^HI populations. Histogram (right) shows the GFP fluorescence intensity for CD127^HI CD200^LO and CD127^LO CD200^HI populations. Plots depict naive, polyclonal Nur77-GFP CD8⁺ T cells. Data represent two (D) to three (E and F) independent experiments with n = 6 mice (D) or n = 3 mice (E and F). Bars in D depict the mean, and error bars show ± s.d. Statistical testing was performed by unpaired two-tailed Student’s t test. NES, normalized enrichment score.