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Fig. 6. Inhibition of Snail activity blocks the expression of neural crest markers. One blastomere of a 2-cell stage embryo was injected with 700 pg of the different dominant negative constructs, treated with dexamethasone at stage 12.5, fixed at stage 19, and the expression of the neural crest markers analysed. The injected side is indicated by an arrowhead. (A-D) XsnailZnFGR: dominant-negative of the Snail zinc fingers. Note that the dominant negative construct inhibited the expression of all the neural crest markers analysed. (E,F) Rescue of XsnailZnFGR by XsnailGR: both mRNAs were injected in equivalent amounts and analysed as previously described. Note the normal expression of the neural crest markers in the injected side. (G,H) SnailN-GR: dominant-negative using the Snail N-terminal domain. Note that the dominant-negative constructs inhibited the expression of all the neural crest markers analysed. (I,J) Rescue of XsnailNGR by XsnailGR: both mRNAs were injected in equivalent amounts and analysed as previously described. Note the normal expression of the neural crest markers in the injected side. |
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Fig. 8. Snail functions as a transcriptional repressor. One blastomere of a two-cell stage embryo was injected with 700 pg of a Snail repressor construct (A-D) or the Snail activator construct (E-H), treated with dexamethasone at stage 12.5, fixed at stage 19, and the expression of neural crest markers analysed. Arrowhead, injected side. Note that the Snail repressor construct (XsnailZnF-GR-EnR) produced an expansion of the neural crest markers on the injected side (A-D), while the Snail activator lead to an inhibition in the expression of the markers (E-H). |
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Fig. 4. The expansion of the neural crest territory induced by Snail or Slug does not require cell proliferation. One blastomere of a two-cell stage embryo was injected with 700 pg of Xsnail-GR mRNA (A,B) or Xslug-GR (C,D), treated with dexamethasone and HUA at stage 12.5, fixed at stage 19 and the expression of the neural crest markers Snail (A,C) and Slug (B,D) analysed. The injected side, which can be recognised by the blue FLDx staining, is indicated by the arrowhead. Note the expansion in the expression of the neural crest markers on the injected side. (E) HUA treated and (F) control embryos stained for histone H3 to verify the blockade in cell proliferation induced by the treatment. The inset shows a higher magnification of the embryos. Note the staining in absence of HUA treatment, but the lack of staining after HUA treatment. |
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Fig. 9. Snail controls neural crest migration. One blastomere of a two-cell stage embryo was injected with 700 pg of SnailGR (A,D), its dominant negative (B,E) or SlugGR (C,F) treated with dexamethasone at stage 16, fixed between stages 22 and 23, and the expression of the neural crest markers Slug analysed. (A-C) Injected side (arrowhead). (D-F) Uninjected side of the embryos shown in A-C. The leading edge of migration is indicated with a broken line. Note that SnailGR (A,D) and SlugGR (C,F) produces a stronger migration in the injected side; while the injection of ZnFXsnailGR (B,E) leads to an inhibition in the migration of the crest cells. |
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Fig. 10. Functional equivalence of the Snail genes assayed in Xenopus embryos. One blastomere of a two-cell stage embryo was injected with 500 pg of mRNA encoding for different members of the Snail gene family, treated with dexamethasone at stage 12.5, fixed at stage 25, and the expression of the neural crest marker Slug analysed. Arrowhead indicates injected side. (A,B) Xenopus genes: Xsnail-GR (A) or Xslug-GR (B). (C,D) Chick genes: Snail-GR (C) or Slug-GR (D). Note that in all the injected sides of the embryos a more vigorous and larger population of migratory crest cells (asterisks). |