Fig. 3. Functional analysis of FoxD3 fusion proteins. (A) Schematic of the structure of FoxD3 and the FoxD3 fusion proteins. FoxD3 contains a conserved winged helix (WH) DNA-binding domain (residues 92-192). The repressor fusion protein (Eng-FoxD3) contains the repressor domain of Drosophila Engrailed (residues 1-298) fused to the FoxD3 WH domain. The activator fusion protein (VP16-FoxD3) contains the activation domain of HSV VP16 (residues 410-490). Embryos were injected with FoxD3 (100 pg), Eng-FoxD3 (100 pg) or VP16-FoxD3 (250 pg) and animal explants were analyzed at the tailbud stage (stage 25) for morphogenesis (B-E) and by RT-PCR for the expression of Muscle Actin (M. Actin) and Collagen Type II (Col II) (F). Like FoxD3, Eng-FoxD3 induced convergent extension movements and mesodermal gene expression, whereas VP16-FoxD3 did not. (G) Co-expression of VP16-FoxD3 and FoxD3 blocked induction of mesodermal genes by FoxD3. PCR controls are as described in Fig. 2.