Since Bptf mutant embryos are unable to form a functional DVE and AVE, we anticipated that they should be defective in specifying the primitive streak and differentiating mesoderm and definitive endoderm. Several critical transcription factors and signaling molecules such as T, Lhx1, Fgf8, Gsc, Foxa2, Nodal, and Cripto (Tdgf1) serve as effective markers for development of the primitive streak in the gastrulating embryo [34], [39]–[42]. Our analyses revealed that T, Lhx1, Fgf8, Gsc, and Foxa2 were undetectable at E6.5, and in the case of T, Fgf8 but not Lhx1, were delocalized in expression at E7.5 (Figure S11A, Figure S11B). The absence of expression of primitive streak markers at E6.5 confirms our histological analyses, and further supports the observation that gastrulation and mesoderm formation do not occur in Bptf mutants. Interestingly, the delocalized Nodal and Cripto expression patterns observed in the Bptf mutants at E6.5 are highly reminiscent of their expression patterns prior to the establishment of the DVE (Figure 2B) (Figure S11A) (Figure S10A, Figure S11B) [34],[39]. Taken together, the data suggests that Bptf mutant embryos arrest at a stage prior to DVE formation (<E5.5).
