Figure 6

Asymmetric spaw expression in vangl2^m209 mutants cannot be rescued by VANGL2 variants. (A–A″) Representative images of mRNA in situ hybridization for southpaw (spaw) the zebrafish homologue of Nodal at 20 hpf. In wild-type siblings (A), spaw is restricted to the left lateral plate mesoderm (LPM, arrowhead), alternatively, spaw may be abnormally expressed in the right LPM (A′, arrowhead) or bilaterally (A″, arrowheads). (B) Quantification of abnormal spaw expression at 20 hpf to examine efficacy of WT VANGL2 to rescue homozygous vangl2^m209 mutant phenotype. WT VANGL2 mRNA injection completely rescues spaw expression. Colour-coded regions defined by vangl2^m209 mutant data range: within/no rescue is grey, below/rescue is green and above/worsens is magenta. Each dot denotes a clutch, consisting of 17–30 embryos. (C) Quantification of abnormal spaw expression at 20 hpf in homozygous vangl2^m209 mutants injected with mRNA encoding different VANGL2 VUS. Variants are ranked and grouped based on result of statistical test when compared to uninjected homozygous vangl2^m209 mutants and homozygous vangl2^m209 mutants injected with WT VANGL2 mRNA (see Supplementary Table 5 for statistical tests). Each dot denotes a clutch, consisting of 18–43 embryos, colour coded regions and dots denote comparison against the uninjected homozygous vangl2^m209 mutant data from panel B. A–A″: Dorsal views, anterior up. B-C: One-way ANOVA with multiple comparisons, Mean ± SEM, ^****: p < 0.0001.