Figure 7

Heart tube formation and positioning is sensitive to loss of vangl2 function. Representative heart tube phenotypes of homozygous vangl2^m209 mutant at 26 hpf, highlighted by mRNA in situ hybridization of the pan cardiac marker myosin, light chain 7, regulatory (myl7). (A) Normal, left jogging (A′) right jogging (A″) no jogging and (A‴) cardia bifida, the presence of two shorter heart tubes at the midline. (B) Quantification of abnormal heart jogging scored live between 28–30 hpf to examine efficacy of WT VANGL2 to rescue homozygous vangl2^m209 mutant phenotype. WT VANGL2 mRNA injection completely rescues heart jogging. Colour-coded regions defined by homozygous 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 16–29 embryos. (C) Quantification of abnormal heart jogging scored live between 28–30 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 6 for statistical tests). Each dot denotes a clutch, consisting of 21–42 embryos, colour coded regions and dots denote comparison against the range of uninjected mutant data from panel B. (D) Quantification of cardia bifida scored live between 28–30hpf to examine efficacy of WT VANGL2 to rescue homozygous vangl2^m209 phenotype. WT VANGL2 mRNA injection completely rescues cardia bifida. Colour-coded regions defined by homozygous 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 16–29 embryos. (E) Quantification of cardia bifida scored live between 28–30 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 7 for statistical tests). Each dot denotes a clutch, consisting of 21–42 embryos, colour coded regions and dots denote comparison against uninjected homozygous vangl2^m209 mutant data from panel D. Heart jogging and cardia bifida were analysed in the same clutch. A–A‴: dorsal views, anterior up. B-E: One-way ANOVA with multiple comparisons, Mean ± SEM, ^****: p < 0.0001.