Cell Type-Specific Targeted Mutations of Kras and Pten Document Proliferation Arrest in Granulosa Cells versus Oncogenic Insult to Ovarian Surface Epithelial Cells

The small G-protein KRAS is crucial for mediating gonadotropin-induced events associated with ovulation. However, constitutive expression of Kras(G12D) in granulosa cells disrupted normal follicle development leading to the persistence of abnormal follicle-like structures containing nonmitotic cells. To determine what factors mediate this potent effect of Kras(G12D) gene profiling analyses were done. We also analyzed Kras(G12D);Cyp19-Cre and Kras(G12);Pgr-Cre mutant mouse models that express Cre prior to or after the initiation of granulosa cell differentiation, respectively. Kras(G12D) induced cell cycle arrest in granulosa cells of the Kras(G12D);Cyp19-Cre mice but not in theKras(G12D);Pgr-Cre mice, documenting the cell context-specific effect of Kras(G12D). Expression of Kras(G12D) silenced the Kras gene, reduced cell cycle activator genes, and impaired the expression of granulosa cell and oocyte-specific genes. Conversely, levels of PTEN and phosphorylated p38 mitogen-activated protein kinase (MM)K) increased markedly in the mutant granulosa cells. Because disrupting Pten in granulosa cells leads to increased proliferation and survival, Pten was disrupted in the Kras(G12D) mutant mice. The Pten/Kras mutant mice were infertile but lacked granulosa cell tumors. By contrast, the Pten(fl/fl);Kras(G12D);Amhr2-Cre mice developed aggressive ovarian surface epithelial cell tumors that did not occur in the Pten(fl/fl); Kras(G12D);CYP19-Cre or Pten(fl/fl); Kras(G12D);Pgr-Cre mouse strains. These data document uneqivocally that Amhr2-Cre is expressed in and mediates allelic recombination of oncogenic genes in ovarian surface epithelial cells. That Kras(G12D)/Pten mutant granulosa cells do not transform but rather undergo cell cycle arrest indicates that they resist the oncogenic insults of Kras/Pten by robust self-protecting mechanisms that silence the Kras gene and elevate PTEN and phosphorylated p38 MAPK. [Cancer Res 2009;69(16):6463-72]