Fig. 7.

Ectopic expression of GATA6 or GATA4 enables Grb2 (−/−) ES cells to form endoderm layer in embryoid bodies. Embryoid bodies were produced from aggregation of RW4 or Grb2 (−/−) ES cells transfected with expression vectors for GATA4, GATA6, and histone H2B-GFP (control). The 3-day embryoid bodies were processed for analysis by Western blot and immunofluorescence microscopy. A: Western blot was performed to compare markers of the cell aggregates. B: Representative embryoid bodies show the presence of partial endoderm layer from Grb2-null ES cells transfected with GATA4 or GATA6. The endoderm cells were stained with Dab2 (green), and GATA4 (red) in GATA4-transfected cells, and GATA6 (red) in GATA6-transfected cells. DAPI marks the nuclei. C: Representative embryoid bodies show the presence of partial endoderm layer from Grb2-null ES cells transfected with GATA4 or GATA6, compared to embryoid bodies from RW4 ES cells. The endoderm cells are stained with Dab2 (green), and laminin (red) for basement membrane. DAPI marks the nuclei. D: The stainings for Dab2 (green) and laminin (red) are shown in a higher magnification for the presence of partial endoderm layer from Grb2-null ES cells transfected with GATA4 or GATA6, comparing to embryoid bodies from RW4 ES cells.