(A) IL-2Rβ per β-actin mRNA expression (mean ± SEM) in thymic CD4⁺ Tregs from WT and KO mice (n = 4/group). (B) Baseline levels of surface IL-2Rβ in WT (considered as 100%) and KO (relative to WT) thymic CD4⁺ Tregs and time kinetics in surface IL-2Rβ remaining after IL-2 stimulation (20 nM) as detected by flow cytometry. (C) Thymic CD4⁺ Tregs from KO and WT mice were cultured overnight in full medium alone or in α-CD3–precoated plates in the presence of IL-2 or (D) in the presence of the PP2A inhibitor okadaic acid or the PP2A activator FTY720, and thereafter the levels of sIL-2Rβ were determined in the supernatants using ELISA. Statistical analysis is relative to unstimulated thymic CD4⁺ Tregs from WT mice. (E) Schematic of primer design and alignment for mouse Il2rb mRNA expression. Total RNA from thymic CD4⁺ Tregs from KO and WT mice (n = 3–4/group) was assessed for mRNA transcripts of the TM domain of IL-2Rβ using quantitative real-time PCR. Cumulative data (mean ± SEM) from 3 experiments are shown. (F) Surface IL-2Rβ (relative to WT) and (G) p-STAT5 expression in thymic CD4⁺ Tregs from KO mice following stimulation with IL-2 (20 nM, 20 minutes) and the pan-sheddase inhibitor TAPI-2. (H) Surface IL-2Rβ (relative to baseline) in thymic WT Tregs after PP2A inhibition with okadaic acid (OA) and in the presence of the pan-sheddase inhibitor TAPI-2 (left end) and p-STAT5 expression relative to OA effect (represented by the dashed line) in response to IL-2 (right end). (I) Surface IL-2Rβ (left end) and p-STAT5 expression (right end) relative to baseline levels (represented by the dashed line) in thymic WT Tregs following IL-2 stimulation and pan-sheddase inhibition with TAPI-2. *P < 0.05; **P < 0.005; ***P < 0.001.