Reduced eye size following GDF6 depletion. (A) In vitro translation of GDF6 mRNA is blocked by GDF6 MOrpholino (GDF6 MO) but is not affected by the standard control MOrpholino (STD MO). (B) Dorsal view of stage 41 tadpole injected with 10 ng GDF6 MO on the left side. (C) Lateral view of tadpoles showing (top to bottom) an unaffected tadpole; GDF6 MO injected sides with an eye 84% of normal contra lateral size and 66% of normal contra lateral size. (D) Graphical representation of the percent of embryos injected with GDF6 MO with the small eye phenotype at stage 41 compared with those injected with STD MO. (E) A severely ventralized embryo injected with 250 pg GDF6 mRNA at stage 41. The percent of ventralized embryos is shown. (F) An example of a partially rescued embryo injected with 250 pg GDF6 mRNA + 20 ng GDF6 MO. The percent of similarly ventralized embryos is shown. (G) A completely rescued embryo injected with 250 pg GDF6 mRNA + 20 ng GDF6 MO. (H) The untreated side of a stage 24 embryo immunostained with anti-phosphorylated-Smad1/5/8 showing staining within the retina and along the neural tube (arrow). Full embryo is shown in the inset. (I) The GDF6 MO (20 ng) injected side of the same embryo showing a loss of phosphorylated-Smad1/5/8 in the retina and neural tube. (J) Frontal view of the same stage 24 embryo showing the reduction in the intensity of the phosphorylated-Smad1/5/8 stain on the injected side (i). (K) Endogenous Smad1/5/8 phosphorylation in St.11.5–12.5 embryonic extracts, lane 2. No Smad phosphorylation is detected in 64 cell embryos lane 1.