Fig. 6
Bmi1 represses Na,K-ATPase β1-subunit. a. β1-subunit expression in 293 T cells transiently transfected with increasing amounts of pEGFPC1 or pEGFPC1-Bmi1. Immunoblots for β1-subunit, GFP, and Bmi1 are shown. α-tubulin serves as the loading control. b. Bmi1 decreased β1-subunit promoter activity in 293 T cells transfected with pEGFPC1 or pEGFPC1-Bmi1 together with β1-subunit promoter-Luc and Renilla control vector. 48 h after transfection, dual-luciferase assays were performed and Firefly luciferase activity was normalized to the Renilla luciferase signal to control for transfection efficiency. Data shown are from three experiments (* = p < 0.05). c. Knockdown of Bmi1 increased β1-subunit promoter activity in 293 T cells. 293 T cells were co-transfected with empty vector or human Bmi1 shRNA construct together with β1-subunit promoter-Luc and Renilla control vector. 48 h after transfection, dual-luciferase assays were performed as in b. Data shown are representative of two independent experiments (** = p < 0.01) d. Schematic model depicting Shh-mediated repression of Na,K-ATPase β1-subunit resulting in increased proliferation and tumorigenicity