Figure 3: CIC negatively regulates T-cell activation.

(a,b) Increased CIC expression in activated CD4⁺ T cells. Western blot analysis for CIC levels in CD4⁺ T cells during T-cell activation by α-CD3/CD28 antibodies (a) and in sorted naïve (CD44^loCD62L^hi) and effector/memory (CD44^hiCD62L^lo) CD4⁺ T cells (b). (c) ELISA of IL-2. WT and Cic-deficient (cKO) CD4⁺CD25⁻CD44^loCD62L^hi naïve T cells were stimulated with plate-bound anti-CD3 (1.0 μg ml⁻¹) in the presence (right) or absence (left) of plate-bound anti-CD28 (2.0 μg ml⁻¹). The supernatants were taken 48 h and 72 h after stimulation and subjected to ELISA for IL-2 concentration. n=4 per each sample. Error bars indicate s.e.m. *P<0.05, **P<0.01 and ***P<0.001 (two-tailed two-sample unequal variance Student t-test). (d) In vitro T-cell proliferation assay. Naïve CD4⁺ T cells purified from pooled spleens and lymph nodes of Cic^f/f and Cic^f/fVav1-Cre mice were labelled with CTV dye and stimulated with plate-bound anti-CD3 (1.0 μg ml⁻¹) in the presence (right) or absence (left) of plate-bound anti-CD28 (2.0 μg ml⁻¹). The cells were analysed 72 h after stimulation. Data are representative of three independent experiments. Shaded area: unstimulated control, blue line: Cic^f/f, red line: Cic^f/fVav1-Cre.