Figure 4.

Activation of Xenopus tropicalis MBD3 promoter by liganded TR in vivo is dependent on the TRE2 within the first intron. (A) Schematic diagram of the MBD3 promoter construct with the putative intronic TRE2 (MBD3) or a mutant TRE2 (MBD3_mTRE). The wild type and mutant promoter fragments were cloned in front of the firefly luciferase coding region. (B) The MBD3 promoter is activated by liganded TR in frog oocytes. The wild type or mutant promoter construct was co-injected with the control Renilla luciferase construct phRG-TK into the nuclei of Xenopus laevis oocytes with prior cytoplasmic injection of GFP mRNA or Xenopus tropicalis TRα and RXRβ mRNAs. The oocytes were incubated at 18 °C overnight in the absence or presence of 100 nM T3 and then lysed for dual luciferase assay. The relative activities of the firefly luciferase to Renilla luciferase were plotted. Error bars indicate S.E.M. Note that mutations in the intronic TRE abolished the liganded TR-mediated activation of the promoter. One-way ANOVA followed by Boferroni’s test was done to compare the samples (* p<0.05, ns: not significant).