Figure 3

Both the PDZ-binding domain and unique sequences in Epb4.1l5^long required for rescue of moe^- mutant defects. (A-C) At 30 hpf in wild-type embryos, Moe localizes cortically in brain and retinal neuroepithelial cells and is concentrated at the apical surface (A) and ZO-1 (green) and panCrb (red) localize to the apical surface of the retina (B) and brain (C). (D-F) At 30 hpf in moe^- embryos, there is no Moe labeling (D) and ZO-1 (green) and panCrb (red) fail to localize to the apical surface of the retina (E) and brain (F). (G) At 30 hpf in moe^- embryos injected with epb4.1l5^longmRNA, Epb4.1l5^long immunoreactivity is cortically localized in most retinal and brain neuroepithelial cells and ZO-1 (green) and panCrb (red) localize to the apical surface in the retina (H) and brain (I). Upper inset (G), magnified section of anti-Epb4.1l5^long labeling and lower inset, uninjected moe^- embryos shows no labeling with anti-Epb4.1l5^long. (J) At 30 hpf in moe^- mutants injected with epb4.1l5^shortmRNA, anti-Epb4.1l5^short is cytoplasmically localized and ZO-1 (green) and panCrb (red) do not localize to the apical surface in moe^- retina (K) and brain (L). Upper inset (G), magnified section of anti-Epb4.1l5^short labeling and lower inset, uninjected moe^- embryos shows no labeling with anti-Epb4.1l5^short. (M, N) At 30 hpf in moe^- mutants injected with myc-epb4.1l5^FERMmRNA, ZO-1 (green) and panCrb (red) do not localize to the apical surface in moe^- retina (M) and brain (N). (O, P) At 30 hpf in moe^- mutants injected with myc-epb4.1l5^long_ΔPBDmRNA, ZO-1 (green) and panCrb (red) do not localize to the apical surface in moe^- retina (O) and brain (P). (Q, R) At 30 hpf in moe^- mutants injected with myc-epb4.1l5^{short+long_PBD} mRNA, ZO-1 (green) and panCrb (red) localize to the apical surface in moe^- retina (Q) and brain (R). (S, T) At 30 hpf in moe^- mutants injected with myc-epb4.1l5^{long+short_PBD}mRNA, ZO-1 (green) and panCrb (red) localize to the apical surface in moe^- retina (S) and brain (T). (U) Western analysis of zebrafish embryos injected with epb4.1l5^longand epb4.1l5^shortmRNA were tested for expression of protein product with isoform-specific antibodies at time points from 6 hpf to 96 hpf. (V) Western analysis of zebrafish embryos injected with epb4.1l5^FERM(anti-Myc), epb4.1l5^long_ΔPBD(anti-Epb4.1l5^long), epb4.1l5^{short+long_PBD}(anti-Myc), epb4.1l5^{long+short_PBD}(anti-Epb4.1l5^long). Blots stripped and reprobed with Anti-α-Tubulin as a loading control. Scale bars, 10 μm (A, D, G, J), 50 μm (lower insets in G, J), 10 μm (remaining panels). (A-T), single confocal z-sections.