Fig. 6. TBX5 activates the Fgf10 promoter. (A) Schematic representation of the upstream regulatory sequences of mouse and human Fgf10. Regions of homology are indicated in red, and are numbered I, II and III; percent nucleotide identity is indicated between the two sequences. Potential TBX5-binding sites (TBEs) are shown as a, b or c, based on the different types of TBEs (see B). A conserved putative TBE is shown by the asterisk. A conserved putative Lef1/Tcf1-binding site is indicated by `L'. (B) Delineation of the three types of TBEs in the Fgf10 promoter. (C) Transactivation by TBX5 of the Fgf10-luciferase reporter construct in COS-7 cells. The reporter construct was transfected with increasing concentrations (0, 100, 500 or 1000 ng) of a Tbx5 expression construct. Mean fold activation is indicated above each bar. (D) Transactivation of the Fgf10-luciferase reporter construct by an activated ß-catenin (ß-cat) construct alone (black bars) or with a Tbx5 expression construct (white bars) or both (hatched bars). The amount of each plasmid transfected is indicated below the graph. (E) Fgf10 promoter deletion analysis. Deletion constructs were co-transfected with the Tbx5 expression construct (white bars) or the activated ß-catenin expression construct (black bars). Deletion of region II (del II) or a point mutation of TBEa1 (muta1) did not affect activation by activated ß-catenin, but greatly reduced (1.5 times less versus 18 times less) activation by TBX5. Deletion of region III (del III) results in decreased activation by ß-catenin, but did not significantly affect activation by TBX5. Deletion of both regions (del II/III) eliminated activation by either construct. All results are expressed as fold increase in luciferase activity compared with the reporter construct alone. Data are shown as mean±s.d. for one representative experiment performed in triplicate.