Knockdown of SerpinE2 mimics the phenotype of neural excision and inhibits migration of neural crest cells
(A) Scheme of extirpation. Dorsal view of Xenopus embryo at stage 17, from which NC tissue was removed on both sides.
(B,C) Tadpole embryo at stage 40 following NC excision (B) and sibling control (C). Note the small head, absence of dorsal fin tissue (open arrowheads) and the reduced number of melanocytes (arrow) resulting from NC extirpation in A.
(D) Unaffected tadpole after microinjection with control-MO into all animal blastomeres at the 8-cell stage. Highlighted are the pigmented melanocytes (arrow) and the intact dorsal fin (filled arrowheads).
(E) SerpinE2-MO causes a reduction of head tissue, dorsal fin structures (open arrowheads) and melanocytes (arrow).
(F) Co-injection of SerpinE2-MO and 2 ng non-targeted Flag-SerpinE2 mRNA restores a normal phenotype.
(G) Scheme for microinjections in K-W. MOs and mRNAs were injected together with 100 pg nlacZ mRNA as lineage tracer (red nuclei) into one dorsal animal blastomere of embryos at the 8-cell stage. The injected side is marked with a star.
(H-Q) Anterior view of neurula embryos. Neither control-MO nor SerpinE2-MO affect Twist and Sox9 expression in NC cells in the head and trunk (arrows) at stage 18 (H-K). SerpinE2-MO inhibits the EMT of Foxd3+ and Snail2+ NC cells (open arrowheads) at stage 20, whereas the control-MO and SerpinE2-5MM-MO have no effect (filled arrowheads) (L-Q).
(R-T) Lateral view of stage 26 embryos. A single injection of control-MO does not affect the migration of Snail1+ NC cells (filled arrowhead). SerpinE2-MO leads to a migration defect on the injected side (open arrowhead). 333 pg Flag-SerpinE2 mRNA rescues NC migration in the SerpinE2-morphant embryo.
br, branchial crest segment; ey, eye primordium; hy, hyoid crest segment; ma, mandibular crest segment; MO, morpholino oligonucleotide; NC, neural crest; ot, otic vesicle. Doses of injected MOs per embryo were 40 ng (D-I) and 10 ng (H-T). Indicated phenotypes were shown in B, 10/11; D, 89/90; E, 64/83; F, 122/132; H, 29/29; I, 24/26; J, 7/7; K, 10/11; L, 9/9; M, 7/7; N, 7/9; O, 7/8; P, 8/9; Q, 9/11; R, 10/10; S, 13/15; T, 9/9. Scale bar, 0.5 mm.