To investigate the differentiation defects of Bptf knockout embryoid bodies at a molecular level we monitored the transcription of well documented markers of endoderm, mesoderm and ectoderm differentiation (Figure 3A–3C). We observed minimal Bptf dependence for the primitive ectoderm markers FGF5 and Otx2 (Figure 3A). However, we did observe significant defects in Nestin transcription, a marker of neural stem cell progenitors derived from the primitive ectoderm (Figure 3A). These results indicate that primitive ectoderm lineages are less dependent on Bptf than the more differentiated lineages. As anticipated we observed severe defects in the expression of mesoderm and endoderm markers during the differentiation time course. We observed little to no activation of mesoderm markers T, FGF8, Evx1, Wnt3, and Gsc which are significantly activated in wild type cultures by day 5. (Figure 3B). Similarly, endoderm markers Sox17, Cer1, Hnf4a, and Foxa2 show severe expression defects in Bptf mutant embryoid bodies compared to controls (Figure 3C). In contrast to the large differences in expression of mesoderm and endoderm markers we observed less than two fold changes in expression of cell cycle regulators
