Figure 5: CD146 is required for CE during zebrafish gastrulation.
From: Wnt5a uses CD146 as a receptor to regulate cell motility and convergent extension
(a) CD146 is essential for CE movement. Embryos at the one-cell stage were injected with 0.5 nM control MO, 4 ng CD146 MO, 50 pg N-terminal Myc-CD146 mRNA and 50 pg Wnt5a mRNA or with CD146 MO plus Myc-CD146 mRNA. These embryos were either viewed laterally, anterior to the top, at 10 hpf and 12 hpf or viewed from the dorsal side at 12 hpf. The angle between the anterior end and the posterior end was measured (scale bar, 250 μm). (b) Knockdown or overexpression of CD146 affects the expression of CE movement-related genes as determined by WISH. The expression pattern of the neuroectoderm marker gsc was determined by WISH at 6 hpf and that of the pre-chordal plate marker hgg1, neuroectoderm marker dlx3 and notochord marker ntl were determined at 10 hpf. The expression pattern of the somite marker myoD was determined at 12 hpf (scale bar, 250 μm). (c) Knockdown of CD146 inhibits JNK activity. Embryos were collected at 10 hpf after injection with control MO, CD146 MO, Myc-CD146 mRNA, Wnt5a mRNA or Myc-CD146 mRNA plus CD146 MO. The expression level of CD146, JNK2 and p-JNK2 was determined by western blotting. β-actin was used as a loading control. (d) CD146 regulates CE movement cell autonomously. Lateral (top two panels) or axial (bottom two panels) cells (red) derived from control, CD146 MO, CD146 mRNA or Wnt5a-mRNA-injected embryos at the shield stage (6 hpf) were transplanted into the corresponding regions of WT recipients at the shield stage and then collected and analysed at the bud stage (10 hpf; scale bar, 250 μm).
