Identification of a Novel Alternatively Spliced Form of Inflammatory Regulator SWAP-70-Like Adapter of T Cells

Figure 3

SLAT2 enhances TCR-induced NFAT activity and IL-4 expression. (a and b) Jurkat-TAg cells (5 × 10⁶) were cotransfected with the indicated plasmid cDNA amounts of empty vector or SLAT and/or SLAT2 together with NFAT-Luc (5 μg) and β-galactosidase (β-gal) (5 μg) reporter plasmids. The cells were left unstimulated or were stimulated with 2 μg/mL anti-CD3 Ab (OKT3) for 6 h at 37°C. Luciferase activity was normalized to the activity of β-gal and data are displayed as % NFAT activity relative to basal activity in unstimulated cells (=100%). Data represent means of triplicates, and graphs show means ± SD. Statistical analysis was performed using Student’s t-test (; ). Expression of transfected SLAT and SLAT2 or endogenous actin proteins was detected by Western blotting (bottom panels). Data are representative of three independent experiments. (c) Schematic diagram of retroviral SLAT and SLAT2 constructs in the pMIG vector. (d) Five-day differentiated Th2 cells were restimulated with anti-CD3 plus -CD28 mAbs and infected with the indicated retroviruses. Cells were harvested on day 9 (4 days after second stimulation), restimulated with anti-CD3 plus -CD28 mAbs, and IL-4-producing cells were enumerated by intracellular cytokine staining. Data are representative of two independent experiments.

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