Contribution of Estrogen Receptor-α and Progesterone Receptor-B to Oncogenic K-Ras-Mediated NIH3T3 Cell Transformation

Abstract

We investigated the biological significance of estrogen receptors (ER) in NIH3T3 cell transformation by the [12Val] K-Ras mutant. This mutant enhanced the steady state level of ER. Cells expressing mutant K-Ras (K12V cells) were tumorigenic. To determine the role of ER accumulation in Ras-transformed cells, we developed cells (KwtER cells) that overexpressed both wild-type (wt) K-Ras and ER, and found that these cells were also tumorigenic. In the presence of 10% serum in the medium, the activation of ER appeared only in transformed KwtER and K12V cells. There was a significant reduction in the expression of progesterone receptor (PR)-B in Rasmediated NIH3T3 cell transformation. Coexpression of the PR with mutant K-Ras led to suppression of tumorigenicity and inhibition of the activation of ER. Functional inactivation of ERα by a dominant negative mutant of ERα (DNER) in the presence of the activated K-Ras 4B mutant arrested the cell cycle at G0/G1, subsequently provoking replicative cell senescence, and finally abrogating tumorigenic potential. p53-dependent upregulation of p21 was implicated in the induction of this cell senescence. The oncogenic K-Ras 4B mutant significantly increased MDM2 (mouse double minute 2) proteins coprecipitated with p53 and suppressed p53 transcriptional activity. In turn, DNER blocked these effects of the oncogenic K-Ras 4B mutant. Finally, we demonstrated that c-Jun expression overcame the suppression and resultant enhancement of p21 protein levels in response to DNER. The data imply that the ERα/AP-1 (activator protein-1) pathway activated by the oncogenic K-Ras 4B mutant contributes to NIH3T3 cell transformation by modulating p53 transcriptional activity through MDM2.