PML4 directly interacts with GATA-1. (A) Reciprocal Co-IP of exogenous HA-PML4 and myc-GATA-1 in 293T. (B) Interaction of GATA-1 and PML isoforms in IP assays. GATA-1 interacts with PML4 specifically. (C) Interaction of PML4 and endogenous GATA-1 in β-estradiol–induced G1E-ER4 cells. IP was performed with rat anti–GATA-1(N6), followed by immunoblotting with anti-PML (PG-M3). (D) Interaction of endogenous PML and GATA-1 in erythroid-differentiated human umbilical CD34⁺ cells. (E) GST-pulldown assay with in vitro expressed GATA-1 (fused to GST) and PML4 (fused to MBP) shows direct interaction between PML4 and GATA-1. CBB staining, Coomassie brilliant blue staining. (F) Colocalization of endogenous PML and GATA-1 in erythroid-differentiated human umbilical cord CD34⁺ cells. Immunofluorescence analyses of endogenous PML (red) and GATA-1 (green) were carried out in the primary erythroid cells at day 6 of EPO induction without (upper) or with (lower) IFNα induction (1000 U/mL, 24 hours) of PML expression. The white arrows in the enlarged panels indicate sites of colocalization. All images were acquired on an Olympus FV1000 Confocal microscope using the Olympus FV1000 Viewer software, version 3.0a; scale bars, 5 μm.