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Fig. 5. The central motifs are crucial for the repressive role of Tcf3 in mesoderm development. (A) Schematic representation of the wild-type and mutated constructs used in this study. The ß-catenin binding domain, the HMG box, LVPQ and SXXSS motifs are as indicated. (B-M) Vegetal view of brachyury (Xbra) expression in stage 10.5 embryos, dorsal towards the top, injections into the right side. Blocking of Xbra expression by injection of 20 ng Tcf1 MO (B) is hardly rescued by co-injection of 0.3 ng XTcf3C (C) or 0.3 ng Tcf4A mRNA (D), but significantly rescued by 0.3 ng Tcf4C mRNA (E), 0.15 ng XTcf3L-SA (F) or 0.3 ng Lef1 mRNA (G). By contrast, downregulation of Xbra expression by injection of 60 ng Tcf3 MO (H) is rescued by co-injection of 0.3 ng XTcf3C (I) or 0.3 ng Tcf4A mRNA (J), but is not rescued by 0.3 ng Tcf4C mRNA (K), 0.1 ng XTcf3L-SA (L) or 0.3 ng Lef1 mRNA (M). (N-S) Vegetal view of Xpo expression in stage 10.5 embryos, dorsal towards the top, injections into the right side. Downregulation of Xpo expression by injection of XlLef1 MO (N) is not rescued by co-injection of 0.3 ng XTcf3C (O) or 0.3 ng Tcf4A mRNA (P), but is rescued by 0.3 ng Tcf4C mRNA (Q), 0.15 ng XTcf3L-SA (R) or 0.3 ng HA-Lef1 mRNA (S). (T) Numerical summary illustrating the penetrance of rescue effects of mRNAs of Lef1, Tcf4 isoforms or Tcf3 mutated constructs in Tcf1 MO- or Tcf3 MO-injected embryos, indicating reduced or absent Xbra expression. (U) Numerical summary illustrating the penetrance of rescue effects of mRNAs of Lef1, Tcf4 isoforms or Tcf3 mutated constructs in Lef1 MO-injected embryos, indicating reduced or absent Xpo expression.

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