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Fig. 3. Ednra is required for early neural crest specification. (A–I) Effect of EdnraMO on neural crest specification. Arrows indicate the injected side. (A'–D') Transverse sections of embryos at the level of cephalic neural crest. (A, B) Ednra-depleted embryos fail to express neural crest markers Snail2 (A) and FoxD3 (B). (A, B, insets) The injected side is recognized by the fluorescence of the lineage tracer FLDx. (C, D) Expression of neural plate marker Sox2 and epidermal marker XK81a is expanded on the EdnraMO-injected side. (E) Embryo processed by double in situ hybridization for Sox2 and XK81a genes showing the reduction of prospective neural crest domain in the injected side. (F) Injection of control morpholino (CoMO) showed no effect. (G) Coinjection of EdnraMO and Ednra′ mRNA rescues Snail2 expression. (H) Single injection of Ednra′ mRNA expanded the Snail2 expression domain. (I) Quantification of rescue experiments. Results are expressed as percentage of embryos normally expressing Snail2 for each treatment (yellow bars), and as percentage of embryos showing reduced Snail2 expression (purple bars). Two different concentrations of Ednra′ mRNA were coinjected for rescue experiments (0.5 ng/embryo and 0.9 ng/embryo). (J–N) Ednra participates in the early neural crest specification. Ednra-injected embryos show increased expression of Snail2 (J) and FoxD3 (K). Expression of the neural plate marker Sox2 (L) and epidermal marker XK81a (M) are reduced on the injected side. (N) Double in situ hybridization for Sox2 and XK81a shows the enlargement of prospective neural crest territory in the injected side. Broken line, dorsal midline; brackets indicate the width of the neural plate (C, L), the width of neural crest domain (E, N) or the width of the neural plate plus the neural crest domain (D, M). |
