Progesterone regulates the proliferation and differentiation of normal mammary epithelium. In
breast cancer cells,
progesterone and its synthetic analogs,
progestins, induce long-term growth inhibition and differentiation. However, the mechanisms responsible are not fully understood. When T-47D
breast cancer cells were treated with the
synthetic progestin ORG 2058 (16alpha-ethoxy-21-hydroxy-19-norpregn-4-en-3,20-dione), all
isoforms of
Wilms' tumor protein 1 (Wt1)
mRNA and
protein were rapidly downregulated. We reasoned that the decrease in Wt1 levels may contribute to the long-term antiproliferative and differentiative effects of
progestins as proliferation and differentiation are known end points of Wt1 action. Consistent with this idea, Wt1
small interfering RNA led to a decrease in S phase and
cyclin D1 levels, and increased
Oil-Red-O staining, indicating increased lipogenesis. Conversely, overexpression of Wt1 attenuated the decrease in S phase induced by
ORG 2058 at 48-96 h. This was accompanied by the sustained expression of
cyclin D1 despite
progestin treatment, and increased levels of
retinoblastoma (Rb) phosphorylation at sites targeted by
cyclin D1-Cdk4 (Ser249/Thr252). Wt1 overexpression also attenuated the ORG 2058-mediated increase in
fatty acid synthase levels and reduced lipogenesis. Thus, Wt1 downregulation was sufficient to mimic the effects of
progestin and was necessary for complete
progestin-mediated proliferative arrest and subsequent differentiation. Furthermore, Wt1 overexpression modulated the effects of
progestins but not anti-
estrogens or
androgens. These results indicate that Wt1 is an important early target of
progestins that regulates both proliferation and differentiation in
breast cancer cells.