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Fig. 1. Activation of the LEF-1 promoter by PITX2. (A) Human LEF-1 promoter elements used in the transfection assays. PITX2 binding sites (Bicoid sites) are denoted by an asterisk (*). (B) CHO cells were transfected with 5 µg of the appropriate human LEF-1 luciferase reporter constructs. The cells were co-transfected with 2.5 µg of either the CMV-PITX2A, or the CMV plasmid without PITX2 (vector control). CHO cell lysates transfected with empty vector were used as a control to show lack of endogenous PITX2 protein in CHO cells. To control for transfection efficiency, all transfections included the SV-40 ß-galactosidase reporter. Cells were incubated for 24 hours, and then assayed for luciferase and ß-galactosidase activities. The activities are shown as mean fold activation compared with the LEF-1 promoter plasmids without PITX2 expression and normalized to ß-galactosidase activity (± s.e.m. from four independent experiments). PITX2 expression did not change the levels of ß-galactosidase activity in the transfected cells.
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