Fig. 2

Regulation of PDE4D activity at IFNAR2 by TPL2. (a) T24 cells were treated with IFN-α (1 × 10⁴ U/mL) for specific time points. The levels of phosphorylated and total TPL2 bound to IFNAR2 or IFNAR1 were detected by performing western blotting after co-immunoprecipitation using IFNAR2 or IFNAR1 antibodies. (b) T24 cells were treated with IFN-α (1 × 10⁴ U/mL), TPL2i (2 μM), and PD98059 (40 μM) for 4 h. The levels of phosphorylated and total TPL2 bound to IFNAR2 or IFNAR1 were detected by performing western blotting after co-immunoprecipitation using IFNAR2 or IFNAR1 antibodies. (c) T24 cells were treated with IFN-α (1 × 10⁴ U/mL) for specific time points. The levels of RACK1 and PDE4D that interacted with IFNAR2 or IFNAR1 were detected after co-immunoprecipitation using IFNAR2 or IFNAR1 antibodies. (d) T24 cells were treated with IFN-α (1 × 10⁴ U/mL) for specific time points. The intracellular cAMP level, activity of total PDE4D, and activity of PDE4D that interacted with IFNAR2 were detected after co-immunoprecipitation using IFNAR2 or PDE4D antibodies. (e) T24 cells were treated with IFN-α (1 × 10⁴ U/mL), TPL2i (2 μM), and PD98059 (40 μM) for 4 h. The activity of total PDE4D was detected after immunoprecipitation using PDE4D antibody. (f) T24 cells were treated with IFN-α (1 × 10⁴ U/mL), TPL2i (2 μM), and PD98059 (40 μM) for 4 h. The PDE4D that interacted with IFNAR2 and their activity were detected after co-immunoprecipitation using IFNAR2 antibody. Data represent the results of five independent experiments. Error bars indicate mean ± SD. *, P < 0.05; **, P < 0.01 (t-test)