Stat3 enhances Progesterone Receptor (PR)-mediated transcriptional activity through Sp-1 in the p21 promoter in breast cancer cells.

PROIETTI, CECILIA; ROSEMBLIT, CINTHIA; BEGUELIN W; SUNDBLAD, VICTORIA; DÍAZ FLAQUÉ, CELESTE; RIVAS, MARTÍN; CARNEVALE, ROMINA; CHARREAU, EDUARDO; SCHILLACI, ROXANA; ELIZALDE, PATRICIA V

San Diego,CA

Congreso; Annual Meeting American Association for Cancer Research; 2008

American Association for Cancer Research

We have previously demonstrated that the synthetic progestin medroxyprogesterone acetate (MPA) is able to induce Stat3 transcriptional activation, which is in turn an absolute requirement for progestin stimulation of in vitro and in vivo breast cancer growth, both in C4HD cells from an experimental model of hormonal carcinogenesis in which MPA induced mammary adenocarcinomas in Balb/c mice, and in the human breast cancer cell line T47D (Mol Cell Biol, 25: 4826, 2005). We have also shown that Stat3, acting as a coactivator, modulates PR function when binding to specific progesterone response elements (PRE) in the promoter regions of target genes, such as bcl-X (Proc Amer Assoc Cancer Res 2007;48:1029). It is known that progesterone induces the cyclin-dependent kinase inhibitor p21 expression through a transcriptional mechanism which involves interaction between progesterone-bound PR and the transcription factor Sp1 at proximal Sp1-binding sites. Here, we studied whether Stat3 could modulate PR activation when regulating the transcription of the p21 gene, which lacks canonical PREs in its promoter region. Assessment of the expression of the progestin-regulated p21 gene showed that MPA treatment of C4HD or T47D cells induced an increase in the levels of these proteins, which was further enhanced in a dose-dependent manner in the presence of a plasmid expressing a constitutively activated Stat3 mutant, Stat3-C. This effect was inhibited when the cells were transfected with Stat3 siRNA. To study the effect of Stat3 on PR-mediated transcriptional activity through this indirect mechanism, C4HD or T47D cells were transiently transfected with a p21 promoter reporter construct, together with increasing amounts of Stat3-C. We found that Stat3-C enhanced progestin-induced PR transcriptional activity in a dose-dependent manner. This effect was completely abolished when Stat3 expression was silenced using an siRNA strategy. We assessed the specific association of Stat3 and PR to the Sp-1 binding site in the p21 promoter in the context of living cells by performing Chromatin Immunoprecipitation Assays. We found that MPA treatment of T47D cells induced Sp-1, PR and Stat3 recruitment to the p21 promoter. These data identify, for the first time, Stat3 as a coactivator of progestin-activated PR when regulating p21 transcription by tethering to DNA-bound trans-acting factor Sp1.