USP2a sustains pY701-STAT1 levels in cells.

(A) 293T cells transfected with or without HA-USP2a were treated with IFNα (500 IU/ml) for 30 min. The levels of pY701-STAT1, HA-USP2a and β-actin were immunoblotted as indicated. (B) HT1080 cells transfected with or without Flag-USP2a were treated with IFNα (500 IU/ml) for 30 min and 60 min, and immunoblotting was performed as indicated. (C) 293T cells transfected with shCON or shUSP2a were treated with IFNα (500 IU/ml) for 30 min and 60 min, and immunoblotting was performed as (B). (D) 293T cells were transfected with Flag-STAT1 and (or) HA-USP2a as indicated. Flag-STAT1 proteins were immunoprecipitated by Flag antibody, and pY701-STAT1-Flag levels were analyzed using indicated antibody. (E) 293T cells transfected with empty vector or Flag-USP2a were stimulated with IFNα (1,000 IU/ml) for 30 min. IFNα was removed and then cells were incubated for indicated times. The levels of pY701-STAT1, Flag-USP2a and β-actin were immunoblotted as indicated. (F) 293T cells transfected with shCON or shUSP2a were treated with IFNα for 30 min. The nuclear proteins were separated, and then were subjected to native-PAGE analysis by indicated antibodies.