Fig 1: AFAP1L2 is upregulated in chronically activated T cells and blunts their survival(A) FACS-purified naive CD8+ T cells from four healthy donors were either analyzed directly by LC-MS or after increasing times of activation with antibodies to CD3 and CD28. Heatmap shows the abundance of proteins that were significantly upregulated in tumor-infiltrating CD8+ T cells with a high percentage of PD-1+ cells (see Figure 4). Each column represents a different donor.(B) Schematic of the activation conditions of naive CD8+ T cells and flow cytometry plots showing the expression of PD-1, TIM-3, CD39, and LAG-3. See also Figure S4B for quantifications.(C) Naive CD8+ T cells were activated as indicated in the schematic in (B) and then analyzed by LC-MS. Heatmap shows the abundance of indicated proteins, where each column represents a different donor.(D) Plot shows AFAP1L2 protein copy numbers at different time points following activation. n = 4 for resting T cells, and n = 3 for other time points.(E) AFAP1L2 protein copy numbers in CD8+ T cells following different activation schemes. n = 3. Two-tailed t test.(F) Same as in (E) but T cells were analyzed by RNA-seq, and AFAP1L2 mRNA counts are shown.(G) NTC and AFAP1L2-edited CD8+ T cells were labeled with CTV and then stimulated with ImmunoCult CD3/CD28/CD2 Activator (1:333). Proliferation (CTV intensity) was followed by flow cytometry over the time course of 5 days. n = 9 from three different donors.(H) NTC and AFAP1L2-edited CD8+ T cells were stimulated with plate-bound anti-CD3/CD28 antibodies for 8 and 12 days, and the number of surviving T cells was quantified by flow cytometry. n = 5 from five donors. Two-tailed t test.