Fig. 8

Tjp3/Zo-3 is essential for EVL permeability barrier function. (A) Biotin permeability assay: negative control without biotin (a), control morpholino (b) and tjp3/zo-3 morpholino (c, d) injected embryos were dechorionized at 6 hpf and exposed to 1 mg/ml Biotin in egg water. Cryosections were probed with labeled streptavidin and images taken using identical parameters. In Cmo injected embryos, although some background staining for streptavidin was observed (a), only little biotin diffusion was detected (b). In 40% of the tjp3/zo-3 morpholino injected embryos, a significant accumulation of biotin was observed in the segmentation cavity between the blastoderm and the yolk syncytial layer (c, d, arrows), demonstrating a compromised epidermal barrier in embryos lacking Tjp3/Zo-3. (B) Morphant embryos are more sensitive to the toxic heavy metal cadmium as compared to control embryos. Embryos injected with either control morpholino (Cmo) or tjp3/zo-3 splice morpholino (MO) were dechorionized at 6 hpf and grown in 1x egg water as a negative control or in egg water containing 0.5 mM CdCl₂ or 1 mM CdCl₂. Significance of differences was established using 2-way analysis of variance. Cmo embryos had a mortality of ∼ 20% after 42 h exposure to low toxicity CdCl₂, whereas the mortality of morphants was significantly higher at ∼ 50% (p < 0.01). At the high cadmium toxicity, Cmo embryos had ∼ 50% mortality and mortality of MO injected embryos was again significantly higher at ∼ 80% (p < 0.05). Mean and standard error (S.E.M.); n ≥ 20 per experiment; 3–4 independent experiments. 2-way ANOVA; p < 0.01. Asterisks indicate statistical significance.