AGK facilitates the JAK2/Stat3 signaling pathway in platelets and megakaryocytes by binding to JAK2. (A) Immunofluorescence image of HEK293T cells transfected with AGK-Flag. HEK293T cells were stained with Flag antibodies (Alexa Fluor 488), DAPI, and a mitochondria-targeting dye (MitoTracker Deep Red). The scale bars represent 30 μm. (B) Coimmunoprecipitation of the lysates of HEK293T cells transfected with JAK2-HA and AGK-Flag/AGK-G126E-Flag using HA beads or Flag beads. (C) Immunoprecipitation of platelet lysates with anti-AGK or anti- JAK2 antibodies. (D) The JAK2 and Stat3 phosphorylation levels of WT and AGK-null platelets in response to 25 IU/mL TPO were tested by western blot analysis. (E) The JAK2 and Stat3 phosphorylation levels of the platelets of WT and AGK-G126E PM mice in response to 25 IU/mL TPO were tested by western blot analysis. (F) Immunofluorescence images of undifferentiated or differentiated megakaryocytes derived from the fetal livers of Agkf/f and Agk−/− mice. Megakaryocytes were stained with tubulin antibodies (Alexa Fluor 488), DAPI, and Stat3 antibodies (rhodamine). Scale bars, 50 μm. IgG, immunoglobulin G; IP, immunoprecipitation.