Fig. 3

Pioneer Proteins but not Intermediate Proteins Are Required to Establish the Mirror Symmetry of the Neural Tissue

(A) In 26-ss N-Cad^m117 mutants, apical marker ZO-1 and basal marker GFAP localized ectopically, and cells organized into cellular rosettes (circled by dashed lines), indicating the loss of the mirror symmetry. By contrast, the mirror symmetry remained in 26-ss nok^m520, pard6^fh266, and nok/pard6γb mutants, despite the presence of cellular bridges (arrows). Top panels, immunohistochemical images; bottom panels, drawings of the distributions of apical and basal markers.

(B) ZO-1 did not align at the midline in N-Cad^m117 mutants at 14-ss and 18-ss.

(C) Drawings contrast the distributions of apical marker ZO-1 in N-Cad^m117 mutants (blue) and wild-type embryos (red) at 14-ss and 18-ss.

(D) Apical marker ZO-1 localized more frequently in the midline region in N-Cad^m117 mutants that expressed N-Cad^wt-GFP (in the Tg(HSP70:N-Cad^wt-GFP)^pt137 transgenic background) than in N-Cad^m117 mutants that expressed N-Cad^m117-GFP (in the Tg(HSP70:N-Cad^m117-GFP)^pt136 transgenic background).

(E and F) To quantify the effects of rescuing expression of N-Cad^wt-GFP (D) on midline distribution of ZO-1, we devised a symmetry index (E), where the midline region is defined as the midline-striding vertical strip of two nuclear diameters wide. (F) Statistical significance was evaluated by one-way ANOVA and Tukey's post hoc analysis. The individual-value bar graphs represent the symmetry indexes of rescued embryos (with means ± SEM). ANOVA, analysis of variance; SEM, standard error of the mean.

(G–G″) In N-Cad^m117 mutants at 14-ss, the PAA-like structures (red arrows, G″) were detectable under TEM, but no apparent adhesion structures could be identified on the opposing apical surfaces (green arrowheads, G′). G′ and G″ are magnifications of the boxed regions in (G). Also see Figure S3.