Altered CD8 T-cell transcriptional profile upon gain or loss of expression of miR-17∼92. Effector status of wild-type, CKO, and CTg donor cells was assessed by analyzing the expression of effector molecules and specific transcription factors implicated in effector and memory fate specification. (A) Representative histogram plots of GZMB expression at days 2.75, 7, and 15 after infection are presented. CD8^WT (Thy1,1⁺) and CD8^CKO (Thy1.2⁺) or CD8^CTg (Thy1.2⁺) P14 cells were cotransferred into congenically mismatched naïve B6 mice, which were infected with LCMV. For day 2.75 analysis, 10⁶ naïve donors were adoptively transferred, and for days 7 and 15 analyses, 10⁵ donors were used. MFI of GZMB expression is indicated in black. Adoptively transferred uninfected mice were used as controls to show the level of GZMB expression in naïve CD8 T cells. Bar charts represent mean ± SEM values of MFI of GZMB expression in gated donors at days 7 and 15 after infection. Results are representative of 2 independent experiments, with n ≥ 3 in each group. (B) Bar graphs show relative levels of GZMA and Perforin, mRNA in purified wild-type, CKO, and CTg donor cells at day 7 after infection. Data were normalized using glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA and represent 2 independent experiments. (C) Relative mRNA levels of Blimp, Bcl-6, ID2, ID3, eomesodermin, and T-bet in wild-type, CKO, and CTg CD8 T cells using quantitative reverse-transcriptase PCR are presented at day 7 after infection. Data from two independent experiments are presented as bar graphs of mean ± SEM relative mRNA levels, with wild-type mRNA levels normalized using GAPDH or HPRT controls. (D) T-bet expression in wild-type (black filled), CKO (black line), and CTg (gray filled) CD8 T cells was assessed at days 5 and 7 after infection in spleen. Numbers indicate MFI of T-bet expression and gray line histograms represent naïve controls. Mean ± SEM values of T-bet MFI on days 5 and 7 post infection are presented as bar graphs. Results are representative of 4 independent experiments, with n ≥ 2 to 3 per group. Unpaired Student t test was used for comparison of means.