Proliferation of 32D-IL-2Rβ cells reconstituted with similar expression levels of WT and KI IL-15Rα. The coding region of WT IL-15Rα or an IL-15Rα^ext/IL-2Rα^int chimeric construct was subcloned into pRV-GFP vector. Different doses of the plasmids (pRV-GFP-WT or pRV-GFP-KI) were transfected into 32D-IL-2Rβ cells. One day after the transfection, cells were rested in cytokine-free medium for 4 hours, then incubated in medium alone, or with IL-15 (0.01, 0.1, 1.0, or 10 ng/mL), IL-3 (50 ng/mL), or IL-2 (100 U/mL). At the end of the incubation, IL-15Rα expression on transfected cells was measured by flow cytometry. We chose 2 groups of transfected cells with similar transfection efficiency (percentage of GFP⁺ cells, indicated on left panels) and expression levels of WT and KI IL-15Rα (mean fluorescence intensity, indicated on right panels; A) and examined their proliferation (B) by measuring [³H]-thymidine uptake in triplicate as described in “Methods.” Means plus or minus SEM are shown. The results shown are representative of 3 separate experiments.