(A) List and Venn diagram of 23 Nodal target genes found to be down-regulated in a microarray of aplnra/b morphant embryos compared to WT at 50% epiboly (5.25 hpf). (B) Relative luciferase activity regulated by the Activin response element (ARE) in WT, aplnra/b morpholino (MO), oep MO and lefty1/2 MO injected embryos at 30% epiboly (4.7 hpf). Data are represented as means ± SEM. *p<0.05 unpaired two-tailed t-test. (C–E) Phenotypic characterization of WT (D) and aplnra/b morphant embryos (E) when treated with the indicated concentration of the Alk4/5/7 inhibitor SB431542 from the sphere stage (4 hpf) onwards. (F–S) Visualization of the expression of the canonical nodal target genes gsc, flh and sox32 in WT (F,J,N), aplnra/b MOs injected (G,K,O), aplnrains; aplnrbhu4145 double mutant (H,L,P) and aplnrb RNA injected treated with the Aplnr agonist ML233 (I,M,Q) embryos at 8 hpf. Embryos are viewed from the dorsal side. Quantification of the number and spread of sox32 expressing cells (R,S). Data are represented as means ± SEM. *p<0.05, **p<0.01 unpaired two-tailed t-test.