Induction of γ-catenin by X-RARα (A) Sca1+/lin- stem cells transduced with PML/RARα, PLZF/RARα, or γ-catenin were transplanted into sublethally irradiated recipient mice. At day 10 after transplantation MNCs from the BM and the spleen were analyzed by Western blotting for the expression of X-RARα and γ-catenin. Another PML/RARα-positive animal that showed disease symptoms at day 12 after transplantation (PML/RARα-2) was included. The blots were first incubated with an anti-RARα antibody and after stripping with an anti-γ-catenin antibody. Equal protein loading was confirmed after further stripping of the membrane and staining with an anti-β-tubulin antibody. The corresponding bands are indicated. (B) Transactivation of the γ-catenin promoter by X-RARα and AML-1/ETO. The γ-catenin promoter construct was transfected by electroporation into U937 cells expressing the respective transgenes under the control of an Zn²⁺-inducible methallothionein 1 (MT-1) promoter. The pGL3basic was used as control. Twelve hours after transfection the transgene expression was induced by the exposure to 100 μM Zn₂SO⁴, and luciferase expression was measured 24 hours later and normalized with Renilla activity. The average of triplicates of one representative of 3 independent experiments is given. (C) Effect of t-RA on the transactivation of the γ-catenin promoter by X-RARα and AML-1/ETO. Twelve hours after transfection the transgene expression was induced by the exposure to 100 μM Zn2SO4 for 4 hours prior to adding t-RA at a concentration of 10-6 M, and luciferase expression was measured 24 hours later. The average of triplicates of one representative of 3 independent experiments is given.