Fig. 5

Ldb2a and Smad7 function in the same pathway.

Knockdown of smad7 increased expression of Nodal target genes, cyc (A–B) and mixer (D–E), whereas overexpression of smad7 mRNA abolished cyc expression (C). In (A–E), one experiment was performed, with the total number of analysed embryos indicated in each panel. These are mainly confirming results published already [39]. (F) Smad7 mRNA level decreased in ldb2a morphants from the sphere stage, shown by RT-qPCR. Error bars are drawn from three independent experiments, each with three technical replicates. (G) Western blot showed decreased protein levels of Smad7 in shield stage ldb2a morphants. β-actin was the loading control. (H–O) Co-injection of ldb2a MO with low-level smad7 MO that did not cause any defect on its own resulted in ectopic expression of Nodal targets, cyc and lefty1, in the ectoderm, suggesting a synergistic effect between ldb2a and smad7 MOs. Experiments were repeated twice, with the total number of analysed embryos indicated in each panel. The wildtype control refers to uninjected embryos that are stage matched.