Wnt-11/Fz7 and PAPC cooperate in decreasing C-cadherin–mediated cell adhesion. (A) Scheme of cell dispersal assay. One animal blastomere of a 32-cell–stage embryo was injected with synthetic mRNAs together with the lineage tracer GAP43-GFP. At stage 12.5, intermingling of GFP-expressing cells was monitored. The patch morphology reflects changes in cell sorting activity as a result of changes in cell adhesion: tight patches with sharp borders and high fluorescence intensity indicate strong cell sorting properties. (B) Cell dispersal assay. Representative images of stage-12.5 embryos. Dashed lines reflect embryo size. Bar, 900 µm. Corresponding pseudocolor images show fluorescence intensity. Relative intensity of fluorescence is indicated by the color bar, where blue and violet represent the lowest and the highest level of detectable fluorescence, respectively. (C) Evaluation of the increasing changes in cell sorting activity. n, number of embryos; # exp., number of experiments. (D) Quantitative evaluation of cell dispersal assay. The diagram displays the cell sorting activity expressed as the relative ratio between intensity and area (for more details, see Fig. S3 and Materials and methods). Error bars indicate SEM. Student’s t test was performed (**, P < 0.005). (E) Scheme of blastomere adhesion assay. Synthetic mRNAs were injected in the animal hemisphere of 4-cell–stage embryos together with the lineage tracers GAP43-GFP and H2B-mCherry. At stage 8.5, animal caps were dissected. The blastomeres of the inner layer were dissociated, spotted on 1 µg/ml CEC1-5–coated substrates, and allowed to adhere. The adhesion strength of blastomeres was measured by the ratio of the number of blastomeres remaining attached after (N_t) versus the number before (N₀) flipover of the Petri dish. (F) Representative images of dissociated blastomeres expressing the indicated constructs attached to the CEC1-5 substrate before and after the flipover. Bars, 100 µm. (G) Quantification of the blastomere adhesion assay. The diagram displays the xC-cadherin adhesion index as the relative ratio between blastomeres attached to the CEC1-5 substrate after and before flipover. Injection amount was as follows: 500 pg xPAPC RNA, 500 pg xFz7 RNA, 20 pg xWnt-11 RNA, 100 pg GAP43-GFP RNA, and 250 pg H2B-mCherry RNA. Error bars indicate SEM. Student’s t test was performed (*, P < 0.05; **, P < 0.005). n, number of counted blastomeres.