Figure 2

Tbx3 represses endogenous PTEN mRNA levels. (A) Different amounts of plasmid DNA (0.2, 0.5 and 1.0 μg) that expresses Tbx3 was transfected into HeLa and HEK cell lines. RNA from untransfected and transfected cell lines was isolated and PTEN mRNA levels were determined using TaqMan probes. Results were normalized to β-Actin mRNA levels. Spearman’s Correlation test revealed a significant decrease in PTEN mRNA levels with increasing amounts of Tbx3 expressing plasmid (r= −0.858 and p<0.05). (B) Cells transfected with 1μg of Tbx3 expressing plasmid and untransfected cells were analysed by flow cytometry using differentially labelled antibodies against Tbx3 (Labelled with DyLight 488) and PTEN (labelled with PE-A) for determination of Tbx3 and PTEN protein levels. Left panels (untransfected cells, a and c) display the endogenous Tbx3 and PTEN protein levels, which are shown in P6 and P3, respectively. Upon transfection, Tbx3 protein is increased (b) within the cells. In cells, where Tbx3 is over expressed, PTEN protein is decreased (d) with respect to that seen in untransfected cells (c).