Figure 4. Identification of Sp-1 sites important for TIEG regulation of OPG promoter activity.
A) Four potential Sp-1 binding sites were identified within the first 200 bps of the OPG promoter. B) Site-directed mutagenesis was used to alter each of the identified Sp-1 binding sites. pGL3 basic luciferase constructs harboring the full length OPG promoter with or without individually mutated Sp-1 sites were transfected into TIEG KO calvarial osteoblasts in the presence or absence of a TIEG expression construct. Twenty-four hours following transfection, cells were lysed and analyzed for luciferase activity. The results were repeated in three independent experiments and a representative data set is shown. * indicates significance at the P<0.05 level (ANOVA) compared to vector controls. # indicates significance at the P<0.05 level (ANOVA) compared to the full-length OPG promoter.