Fig. 8

Def6 RNA rescues the CE movement defects observed in wnt5b morphants.

Embryos injected with def6 RNA (150 pg), wnt5b MO (5 ng) alone and together at the 1–2 cell stage were fixed at tail-bud stage (10 hpf), and stained for dlx3 (marks the anterior borders of the neural plate) and ntl (marks the presumptive notochord) expression. (A–C) Expression of dlx3 shows restoration of the wider neural plate in the embryos co-injected with def6 RNA and wnt5b MO. Def6 RNA was also sufficient to rescue the reduced anterior extension of the presumptive notochord as revealed by ntl expression. (D) Representative embryo at tail-bud stage viewed from the dorsal side with anterior to top. The measures taken are shown. The black line indicates the width of the embryo whereas the red double-headed arrow is the width of the neural plate. (E) The ratio of the width of the neural plate divided by the width of the embryo was quantified for each category of embryos. ANOVA single factor indicated a significant (p<0.001) difference between the three groups of embryos. Two-tailed Student′s t-tests showed a significant (p<0.001, three asterisks) increase in this ratio in wnt5b morphants versus def6 RNA-injected embryos and statistical significance (p<0.001; three asterisks) in wnt5b MO-injected embryos versus embryos co-injected with def6 RNA and wnt5b MO. There was no statistical difference (N.S) between def6 RNA only versus rescued embryos. (F) Embryos at 24 hpf were morphologically analysed and separated into four categories according to their phenotype: normal (blue box), mild (purple box), moderate (green box), and severe (red box). (G) The phenotypes of the embryos from three independent experiments were scored and the percentages of normal (blue bar), mild (green bar), moderate (yellow bar) and severe (red bar) phenotypes were plotted on the graph.