Figure - PMC

Skip to main content

View full-text article in PMC

. Author manuscript; available in PMC: 2007 Jun 15.

Published in final edited form as: Arch Biochem Biophys. 2006 Oct 16;460(2):285–292. doi: 10.1016/j.abb.2006.09.027

The mutant VDR fails to bind coactivators but interacts with the corepressor hairless. Panel A. VDRs were synthesized by in vitro coupled transcription-translation and incubated with GST-SRC-1 or GST-DRIP205 in the presence of vehicle or 100 nM 1,25(OH)₂D₃. The samples were then subjected to GST-pull down assays. Bands were detected by western blot. No interactions were observed between the truncated VDR and the coactivators while the WT VDR exhibited a ligand-dependent interaction with both coactivators. Panel B. Mutant and WT VDRs were co-expressed with myc-tagged rat HR in COS-7 cells. Cell extracts were prepared and proteins immunoprecipitated with VDR or non-specific IgG antibodies. HR was detected by western blot using α-c-myc antibodies. HR was immunoprecipitated by both the WT VDR and Y401X mutant VDR demonstrating that the mutant VDR interacts with HR in vivo. A small amount of binding was observed with the control IgG. In, input; α-VDR, immunoprecipitating antibody; IgG, non-specific control immunoprecipitating antibody.

The mutant VDR fails to bind coactivators but interacts with the corepressor hairless. Panel A. VDRs were synthesized by in vitro coupled transcription-translation and incubated with GST-SRC-1 or GST-DRIP205 in the presence of vehicle or 100 nM 1,25(OH)₂D₃. The samples were then subjected to GST-pull down assays. Bands were detected by western blot. No interactions were observed between the truncated VDR and the coactivators while the WT VDR exhibited a ligand-dependent interaction with both coactivators. Panel B. Mutant and WT VDRs were co-expressed with myc-tagged rat HR in COS-7 cells. Cell extracts were prepared and proteins immunoprecipitated with VDR or non-specific IgG antibodies. HR was detected by western blot using α-c-myc antibodies. HR was immunoprecipitated by both the WT VDR and Y401X mutant VDR demonstrating that the mutant VDR interacts with HR in vivo. A small amount of binding was observed with the control IgG. In, input; α-VDR, immunoprecipitating antibody; IgG, non-specific control immunoprecipitating antibody.