FIG. 5.

BTG2 activates BMP-dependent transcription and associates with Smad1 or Smad8 in vivo. (a and b) Effect of Btg2 on TGF-β1-induced transcription of SBE₄-Luc (a) and ARE-Luc reporters (b). C2C12 cells were transiently transfected with a control vector (pCMV-β-gal) or Btg2 (1 μg/well) along with either SBE₄-Luc (a) or ARE reporter plus FAST1 (b). (c and d) Effect of Btg2 on BMP-induced transcription of BRE-Luc reporter. (c) C2C12 cells were transiently transfected with the control vector or Btg2 (1 μg/well) along with BRE-Luc. The cells were treated with 50 ng of BMP-2/ml (black bars) or left untreated (white bars). (d) C2C12 cells were transiently transfected with the control vector or increasing amounts of Btg2 along with BRE-Luc. Luciferase activity was normalized to β-galactosidase activity and plotted as the mean and standard deviation for triplicates from a representative experiment. (e) GST-BTG2 was transfected into 293T cells with the Flag-tagged Smad1 or Smad5 or Myc-tagged Smad8 construct. Cell extracts were subjected to GST pull-down assay using glutathione-Sepharose 4B beads, followed by immunoblotting with anti-Flag or anti-Myc antibody. Expression of GST, GST-BTG2, and Smads was monitored as indicated.