To assess the specification of the VE and DVE, we monitored the expression of Cer1, Hex1, Gata6, and Nodal in E5.5 Bptf mutant embryos. We observed that the markers Cer1, Hex1 are significantly reduced in Bptf mutants relative to the wild type controls (Figure 2B). The absence of expression of these markers indicates that the DVE does not form in the absence of Bptf. Interestingly Gata6 expression, normally expressed only in the VE at E5.5 and E6.5, is absent in the VE but present in the epiblast in mutants at E5.5 (Figure 2B). This suggests that Bptf could also have roles in specifying the VE as well as the DVE. A key regulator of DVE specification is Nodal. Nodal expression is found as early as E4.5 but is not significantly expressed in the epiblast and VE until implantation is well underway at E5.0 [36],[38]. In E5.5 Bptf mutant embryos we observe normal expression levels of Nodal in the epiblast and VE as in wild type embryos (Figure S10A). This E5.5 pattern of Nodal expression continues into E6.5 in Bptf mutant embryos (Figure S10B).
