Figure 3.

Ryk is essential for CE movements. (A–H) Overexpression or depletion of XRyk caused the disruption of CE movements. Two blastomeres of four-cell stage embryos were injected at the dorsal equatorial region with 500 pg XRyk mRNA, 40 ng XRyk MO, or 40 ng Co MO. (A–C) Compared with the Co MO–injected embryos, embryos expressing XRyk mRNA or MO at stage 33 showed dorsal flexure and a failure to straighten the anteroposterior axis. (E–G) Keller sandwich explants from Co MO–injected embryos elongated, but the explants expressing XRyk mRNA or MO were significantly inhibited. (D and H) Rescue of XRyk knockdown by coinjection of ORF XRyk mRNA. (I–O) XRyk did not affect mesodermal cell fate specification. (I–N) Whole mount in situ hybridization with Chordin, Xbra, and MyoD as probes. (O) RT-PCR analysis using primers specific for Siamois, Xnr3, Chordin, Goosecoid, and Xbra. ODC, ornithine decarboxylase, (a loading control); −RT, minus reverse transcription control sample. (P–R) XRyk or Wnt11 depletion inhibited the Dsh puncta colocalization with Rab5 in DMZ cells. 60 ng Wnt11 MO, 40 ng XRyk MO, or 40 ng Co MO was coinjected with GFP Dsh and myc Rab5 into the two dorsal blastomeres at the four-cell stage. DMZ explants were dissected at stage 11–11.5 and then subjected to fluorescence analysis. Bars, 20 μm.