Activation of the Jak2/EpoR pathway by Jak2V617F is inhibited by the Y114A mutation. Parental BaF3.EpoR cells or cells expressing various mutants of Jak2 (as indicated) were used. (A,B) Parental BaF3.EpoR cells and cells infected to express Jak2 or various mutants were growth factor deprived. The phosphorylation and protein expression of Jak2 (A) or the Epo receptor (B) were detected after specific immunoprecipitations by immunoblotting as indicated. (C) Intracellular ROS levels were measured by DCF-DA staining (n = 3) in the absence of growth factors. Parental BaF3.EpoR cells were used as a control.