Eomes broadens the scope of CD8 T-cell memory by inhibiting apoptosis in cells of low affinity
Fig 2
Eomes promotes memory precursor formation early after activation.
(A) WT (CD45.1+) and EomesCKO (CD45.2+) OT-1 cells were mixed in a 1:1 ratio and stimulated with the indicated peptides. The ratio between cell populations was followed over time by flow cytometry. (B-D) OT-1 (CD45.1/2+) and EomesCKO OT-1 cells (CD45.2+) were mixed in a 1:1 ratio, and 104 cells were injected into WT recipients (CD45.1+). Two days later, recipients were infected with LM-N4, LM-Q4, or LM-V4. (B) Ratio between WT and EomesCKO cells in blood over time. (C) Fraction of CD127+KLRG1- cells among donor cells. (D) Representative FACS plots for day 14 were gated for CD8+CD45.2+ cells. (E) OT-1 (CD45.1/2+) and EomesCKO OT-1 cells (CD45.2+) were mixed in a 1:1 ratio, and 104 cells were injected into WT recipients (CD45.1+). One day later, recipients were infected with mCMV-N4, mCMV-Y3, or mCMV-Q4. Seven days after infection, Ki67 expression was determined in splenic donor cells by flow cytometry. (F) EomesiCKO cells were transferred in WT recipients. Mice were infected with LM-N4, LM-Q4, or LM-V4 and injected with Poly(I:C) 0 or 6 days after infection. After 4 weeks, mice were reinfected with mCMV-N4. The fraction of eYFP+ donor cells was followed in the blood. Experiments show representative data from at least two (B-F) or four (A) independent experiments using 3–5 mice per group. In (A,B,E), ANOVA followed by Bonferroni posttesting was used; in (C,F), Student t test was used to analyze difference between groups. Shown are means ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.001. Values for each data point can be found in S1 Data. A2, SAINFEKL; EomesCKO, Eomesflox/floxCD4Cre; EomesiCKO, EomesFL/FLOT-1+/-RosaStop-EYFPMx1Cre/+; eYFP, enhanced yellow fluorescent protein; FACS, fluorescence-activated cell sorting; LM, L. monocytogenes; mCMV, murine cytomegalovirus; N4, SIINFEKL; Poly(I:C), polyinosine-polycytidylic acid; Q4, SIIQFEKL; T4, SIITFEKL; V4, SIIVFEKL; WT, wild-type; Y3, SIYNFEKL.