Fig. 2

Wnt regulator mutants have differential sensitivities to exogenous Wnt8. (A–C) Overexpression of 100 pg of wnt8 in wild type embryos results in the classic Wnt gain of function eyeless phenotype in ∼ 5% of embryos, but more than 30% in axin2gh1/gh1 and nkd1gh2/gh2 mutants. Small eyes were considered to have eyes. In contrast, the axin2gh1/gh1;nkd1gh2/gh2 resembles the wild-type phenotype (C). (D–F) Analysis of body axis development revealed that only the nkd1gh2/gh2 mutants develop a short-twisted axis with exogenous wnt8 in about 30% of the embryos. (G) The Wnt reporter line was used to assess Wnt activity at the midbrain-hindbrain boundary at 1dpf when either Axin2 and/or Nkd1 was knocked down using sgRNA/Cas9. In each of the sgRNA/Cas9 injections there was an expansion of Wnt activity in the forebrain when compared with wild type embryos. Single plane brightfield images were overlaid with composite GFP images and as such the eye is not visible in all images. Error bars represent SEM, different letters above the bar signify significance with a p value < 0.01 using a one-way ANOVA.