Fig. 1

Global FGF signaling perturbation using the fgfr1‐dn‐cargo transgenic line. A: Schematic showing the crosses and a representative treatment scheme for ubiquitous genetic FGF signaling perturbation with the fgfr1‐dn‐cargo transgenic line. Note that fgfr1‐dn‐cargo contains a 2A‐linked Cerulean‐CAAX ORF. B,B’: Ubiquitous fgfr1‐dn‐cargo activation in 4‐OHT– and heatshock‐treated ubi:creERT2;fgfr1‐dn‐cargo transgenic embryos as shown in schematic (A) leads to ubiquitous but mosaic Cerulean‐CAAX (Cerulean) expression. C: CreERT2‐mediated ubiquitous excision of the loxP‐flanked STOP cassette in ubi:creERT2;fgfr1‐dn‐cargo transgenics 4‐OHT induced during early somitogenesis as detected by PCR (166 bp after recombination, 1129 bp when unrecombined). Excision of the STOP cassette occurs within 1 hr and gradually increases up to 4 hr after 4‐OHT treatment. Shown are PCR on recombined and unrecombined transgene insertions. D–G: Heatshock (hs) controls, ubi:creERT2;fgfr1‐dn‐cargo embryos induced with 4‐OHT at shield stage and heatshocked at 15 ss, and wild‐type embryos treated with 2 or 10 µM SU5402 at 15 ss were fixed at 19 hpf and stained for etv4 expression via mRNA in situ hybridization. Ubiquitous fgfr1‐dn‐cargo activation and concentration‐dependent SU5402 treatment abolish etv4 expression as a readout for FGF signaling activity. H,I: Overlay of EGFP expression on a brightfield (BF) image of a 36‐hpf heatshock control and ubi:creERT2;fgfr1‐dn‐cargo embryo induced with 4‐OHT at shield stage and heatshock‐treated at 20 ss. Embryos with ubiquitous expression of Fgfr1‐dn during late somitogenesis have a mis‐looped heart (asterisk) as well as head defects and malformations in posterior tail structures (arrowheads). The heart malformations also manifest in blood pooling in front of the inflow tract of the heart, leading to a visible edema on top of the yolk (arrow). H’,I’: Cerulean‐CAAX expression in control and signaling‐perturbed embryos. J: 7x magnification of the heart of a 4‐OHT–induced and heatshock‐treated ubi:creERT2;fgfr1‐dn‐cargo double transgenic (I) showing heart defects with a large atrium (A) and diminished ventricle (V); green background fluorescence is caused by bleedthrough of Cerulean fluorescence (see also I’). K: Quantification of phenotypes resulting from global FGF signaling perturbation in genetically perturbed double‐transgenic ubi:creERT;fgfr‐dn‐cargo embryos and single‐transgenic heatshock controls treated as indicated (I).