Figure 7.

Maintained nuclear expression of Vax2 blocks retinal development. Chick optic vesicles (E1.5; HH stage 11–12) were unilaterally electroporated (arrow; right side of the figure) with RCAS–IRES-EGFP (A), RCAS–Vax2–IRES-EGFP (D), RCAS– Vax2(S170A)–IRES-EGFP (G), or RCAS–Vax2(S170D)–IRES-EGFP (J). We injected a total of 20 ng of DNA per embryo (A–G)—except for the phospho-mimicking S170D construct (H–J), where we introduced 10-fold higher levels of DNA per embryo. Images are frontal (A,C,F,I) and lateral (B,D,E,G,H,J) views of electroporated embryos at E7.5. D, G, and J are the electroporated sides of each embryo, and B, E, and H are the control sides. (The expression of the electroporated Vax2 genes is confirmed in Supplementary Fig. S5). (K,L) E7.5 control (K) and Vax2 S170A electroporated (L) eyes, corresponding to those in E and G, respectively, were cryosectioned at 16 μm as indicated by the superimposed frames, to generate the sections schematized in M and N. (O,P) These sections were stained with an anti-Pax6 antibody (red) to visualize retinal tissue and RPE (O, control; P, Vax2S170A). (Q,R) Adjacent sections were stained with a Pax2 antibody (green) to visualize optic stalk/nerve tissue. A sharp border between Pax2-positive optic nerve astrocytes and Pax6-positive retinal cells is observed in the control (O,Q), whereas there is only a Pax2-positive optic stalk, and no Pax6-positive retina, which develops from the Vax2S170A-electroporated neuroepithelium (P,R). Bars, 100 μm.