|
Fig. 3
Disruption of the noncanonical Wnt pathway or the EphB signaling cascade induces the same CE phenotype. (A–D) Morphology [72 hours postfertilization (hpf)] of the wild-type embryo (A), embryo injected with zebrafish NDaam1a mRNA (B), mRNA encoding a soluble form of mouse EphB1 (C), and mRNA encoding a K44A mutant of zebrafish Dnm1 (D). These embryos share a short and bent body axis (red arrowheads), as well as cyclopia (blue arrowheads). (E) EGFP protein was expressed in the notochord cells to visualize their shape. In the wild-type embryo, the notochord cells elongate to show the intercalation movement. When zNDaam1 was expressed, the notochord cells became round and did not develop a polarized shape. Dorsal views at 11 hpf are shown. Rostral to the left, caudal to the right. (F) At 14 hpf, myoD is expressed in the somites and the adaxial cells in the wild type. (G) Injection of zNDaam1 mRNA resulted in a short body axis with compressed somites. (H and I) Injection of mRNA encoding the soluble form of mouse EphB1 (H) or the dominant-negative dynamin1 [Dnm1(K44A)] (I) induced a similar phenotype. (J) Injection of Dnm1-MO resulted in similar but more severe defects. (K) Coinjection of EphB4-MO and EphB2-MO induced severe defects. The notochord was short, broader, and bent. Pictures shown in F–K are shown at the same magnification. (L) Injection of mRNA encoding zNDaam1 or the soluble form of EphB1 induced the abnormalities of CE movement in 19.2% or 11.6% of injected embryos, respectively, as judged by expression of myoD at 14 hpf. Coinjection of these mRNAs induced the same abnormality in 49.4% of embryos. (M) Injection of a soluble mEphB1-FC mRNA induced the CE defect in 29.8% of injected embryos. Coinjection with zCDaam1a mRNA rescued this defect, and the CE abnormality was observed in only 11.4% of embryos. Injection of zCDaam1 mRNA alone induced the defect in 9.0% of embryos.