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Fig. 1. In Xenopus, Krm2 is regulated by Wnt signaling and expressed in the NC region. (A-D) Whole-mount in situ hybridizations. (A) Comparison of Xkrm2, XWnt8 and XWnt3a expression patterns in gastrula and early neurula stage embryos. Top: gastrula stage. Vegetal view, dorsal is up. Bottom: neurula stage. Dorsal view, anterior is up. (B) Comparison of Xkrm2 and slug expression patterns at the indicated stages. Dorsal view, anterior is up. Lowermost panel: view of frontally cut stage 16 embryos, dorsal is up. Brackets indicate overlapping expression domains of krm2 and slug. (C-E) Effect of Wnt pathway perturbations on krm2 expression. (C) Embryos at the 32-cell stage were injected equatorially in two opposite blastomeres with 1 ng PPL or dnWnt8 and 250 pg lacZ mRNA and analyzed at gastrula stage. Arrowheads indicate β-gal lineage tracer staining (blue). Vegetal view, dorsal is up. (D) Embryos at the four-cell stage were injected animally with 200 pg pCS-PPL or pCSKA-Wnt8 DNA or 100 pg pCS-Wnt3a DNA in one blastomere and analyzed at gastrula stage. Lateral view, dorsal to the right. (E) Statistical overview of experiments shown in C and D. (F) Embryos at the four- to eight-cell stage were injected animally with 100 pg XWnt8 or Wnt3a mRNA, or 1 or 2 ng BMP4 mRNA. Animal caps were cut at stage 8-9, cultured until stage 20 equivalent and were analyzed by RT-PCR for expression of the indicated genes. (G) Embryos at the 32-cell stage were treated with 120 mM LiCl for 50 minutes, cultured until stage 11.5 and analyzed by RT-PCR. Histone H4 was used for normalization. -RT, minus reverse transcriptase control. |
