Ccl3 organ-specific posttranscriptional fine-tuning during mCMV. (A) Scheme of the experiment. CCL3-EASER mice were infected with 5 × 105 PFU of mCMV at the indicated time points and analyzed together at time “0.” Isolated leukocytes were stimulated in vitro in the presence of monensin and brefeldin A for 4 h. (B) Representative contour plots from CCL3-EASER liver NK cells at 0 (naive), 4, 8, and 16 h after mCMV infection. The red shaded boxes indicate tdTomato-positive cells. (C) Frequency of tdTomato-positive cells from total NK cells. (D) Kinetic of Ccl3-promotor activity evaluated with tdTomato MFI. (E) Frequency of Venus-positive cells from total NK cells. (F) Dynamics of Ccl3-protein expression evaluated with Venus MFI. (G) Representative XY plots of Venus and tdTomato channel intensity from liver NK cells gated on tdTomato-positive cells (as shown in A) at 0 (naive), 4, 8, and 16 h after mCMV infection. Dotted lines represent the simple linear regression of the correlation between Venus and tdTomato channel intensity. Linear regression is color coded according to the time point of infection: gray = 0 h (naive), blue = 4 h, green = 8 h, and red = 16 h. (H) Comparison of simple linear regression from liver, spleen, and lung. Color code as described in G. Black dotted line indicates tdT50: point of half of tdTomato channel intensity from the control group (naive). Data is presented as mean ± SD; n = 3 mice per group; and representative of two independent experiments. In C–F, data is shown as mean ± SD. Ordinary one-way ANOVA with multiple comparisons to naive group was used: *P < 0.05; **P < 0.01; ***P < 0.001. In G, correlation test was used, two-tailed P value, Pearson r with 95% confidence interval.
