Constitutive expression of the Wilms tumor suppressor gene WT1 in F9 embryonal carcinoma cells induces apoptotic cell death in response to retinoic acid

The product of the Wilms tumor suppressor gene, WT1, is thought to be a tissue specific transcription factor regulating cell growth and differentiation. To elucidate the function of WT1 in cellular differentiation, we examined the changes in the level of WT1 expression during retinoic acid induced-differentiation of embryonal carcinoma F9 cells into parietal endoderm cells. We found that, in response to retinoic acid addition, the expression of WT1 increased significantly after 12-24 h of incubation, then decreased and finally disappeared after 4 days, by which time most of the cells had differentiated into primitive endoderm cells. To examine the significance of these changes in WT1 expression, we established cell lines constitutively expressing one of the WT1 splicing variants. These cell lines showed a phenotype very similar to parental F9 cells in the absence of retinoic acid. However, in the presence of retinoic acid, they failed to differentiate into primitive endoderm cells and underwent apoptotic death 36 h after the addition of retinoic acid. These results suggest that downregulation of WT1 expression is necessary for normal differentiation of F9 cells into parietal endoderm cells.