FIGURE 9

Influence of C80 and S212 residues on Wnt signaling activity (A) Morphological phenotypes at 24 hpf of embryos injected/treated with capped sense RNAs of control (EGFP 100 pg, 37/37 embryos), control +IWR (42/42 embryos, arrowhead: reduction in tail elongation), wt Wnt3 (100 pg, 47/53 embryos, arrowhead: loss of eye), Wnt3-EGFP + IWR (36/43 embryos, arrowhead: restoration of eye), Wnt3C80A-EGFP (100 pg, 41/43 embryos), Wnt3C80A-EGFP + IWR (41/44 embryos, arrowhead: reduction in tail elongation), Wnt3S212A-EGFP (100 pg, 52/54 embryos) or Wnt3S121A-EGFP + IWR (43/46 embryos, arrowhead: reduction in tail elongation). IWR was used at a concentration of 10 μM and DMSO was used in groups that were not treated with IWR. (B) Whole mount in situ hybridization (WMISH) in the transgenic Tg(7xTcf-Xla.Siam:nlsm-Cherry^ia) canonical Wnt/ß-catenin reporter embryos showing the effects of control (EGFP 100 pg, 33/33 embryos, arrowheads: anterior Wnt expression domains), control + IWR (45/45 embryos, arrowheads: reduction in anterior Wnt expression domains), Wnt3-EGFP (100 pg, 41/49 embryos, arrowheads: increase in posterior Wnt expression domain), Wnt3-EGFP + IWR (40/45 embryos, arrowheads: restoration of anterior Wnt expression domain), Wnt3C80A-EGFP (100 pg, 45/48 embryos, arrowheads: no effect on anterior Wnt expression domains), Wnt3C80A-EGFP + IWR (38/39 embryos, arrowheads: reduction in anterior Wnt expression domains), Wnt3S212A-EGFP (100 pg, 44/46 embryos, arrowheads: no effect on anterior Wnt expression domains) or Wnt3S121A-EGFP + IWR (50/53 embryos, arrowheads: reduction in anterior Wnt expression domains) on canonical Wnt signaling. IWR was used at a concentration of 10 μM and DMSO was used in groups that were not treated with IWR1. mCherry WMISH shows upregulation of signaling in Wnt/ß-catenin reporter embryos by Wnt3-EGFP but not by Wnt3C80A-EGFP or Wnt3S212A-EGFP.