Fig. 3. Disturbed Nodal expression patterns in Nodal^600/– embryos. Genotypes are indicated towards the left of each panel. Black boxes indicate functional Nodal alleles and null alleles are shown as white boxes. The blue box represents the lacZ insertion into exon 2 of Nodal. Enhancers are indicated as circles, the PEE in green and the ASE in red. The 600 deletion is indicated by the cross. In all panels, anterior is towards the left. (A) As assessed by ß-gal staining, punctate Nodal^600.LacZ expression observed at 5.5 dpc throughout the epiblast becomes restricted to the proximal epiblast by 6.5 dpc and gradually resolves to the posterior. Expression continues in the primitive streak after the onset of gastrulation, but is undetectable in the VE. (B) Nodal expression directly assessed by whole-mount in situ hybridization. In contrast to wild type (WT), expression in Nodal^600/– embryos is noticeably reduced, restricted to the proximal epiblast, and fails to show obvious AP asymmetry. Section analysis confirms the loss of expression within the VE (data not shown). (C) Nodal expression in Nodal^LacZ/600 embryos is proximally restricted. The embryo shows a characteristic constriction at the embryonic/extra-embryonic boundary (arrowhead). The position of the section shown is indicated by the box. ß-Gal staining is clearly detectable in the VE of these embryos (arrows). (D) ß-Gal staining and tissue morphology in Nodal^600.LacZ/600 embryos resemble that in C. Arrowhead indicates embryonic/extra-embryonic boundary. However no lacZ expression is detected in the VE. VE, visceral endoderm; m, mesoderm; epi, epiblast.

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