Specification of the lymphatic lineage is independent of coup-TFII and sox18. (A) Schematic of the Coup-TFII protein indicating the position of the first affected amino acid in the 1 bp insertion allele nr2f2hu10330 (coup-TFIIhu10330 hereafter) which leads to a premature stop codon after an additional four amino acids N-terminal to the nuclear receptor-DNA binding and ligand-binding domain. (B,C) Brightfield images of wild-type (B) and homozygous mutant coup-TFIIhu10330 (C) embryos, with no signs of edema or morphological abnormalities at 6 dpf. (D,E) flt4:mCit;flt1enh:tdTom-positive embryos from a coup-TFIIhu10330 incross. Compared with heterozygous siblings (D), coup-TFII mutants (E) show only marginal TD defects (asterisk) in a small proportion of embryos at 5 dpf. Note the overall normal vascular morphology with proper PCV, DA and ISVs present in homozygous mutants. Arrowheads indicate the presence of TD. (F) The average length of the TD in the first ten segments above the yolk extension is not significantly affected in coup-TFIIhu10330 mutants. (G) The proportion of venous ISVs is unaltered in homozygous coup-TFIIhu10330 mutants at 5 dpf. (H) Schematic overview of the Sox18 protein, indicating the first affected amino acid in the 1-bp insertion allele sox18hu10320, preceding the HMG-box. The resulting frame-shift leads to a premature stop codon after additional 50 amino acids. (I,J) Bright-field images of heterozygous (I) and homozygous (J) mutant sox18hu10320 embryos at 5 dpf. Note the overall wild-type appearance of mutant embryos (J). (K) The average length of the TD in wild-type (green), heterozygous (yellow) and homozygous mutant (red) sox18hu10320 embryos does not differ significantly. (L,M) sox18hu10320 heterozygous (L) and homozygous (M) mutant embryos expressing fli1a:eGFP in all ECs. In a sox18 loss-of-function situation, embryos do not show defects in the formation of the TD (arrowheads). (N) The average number of PLs present in sox18hu10320 mutants is not significantly affected at 54 hpf. Error bars represent s.d. n.s., not statistically significant.