Smad4 as a Transcription Corepressor for Estrogen Receptor α

Abstract

Antiestrogen compounds exhibit a variety of different effects in different tissues and are widely used for the treatment of osteoporosis, breast cancer, and other diseases. Upon examining the molecular mechanisms, we found that Smad4, a common signal transducer in the bone morphogenetic protein (BMP)/transforming growth factor-β (TGF-β) signaling pathway, functions as a transcription corepressor for human estrogen receptor α (ERα). Endogenous ERα was co-immunoprecipitated with Smad4, and the interaction was induced by antiestrogen ligands such as tamoxifen, raloxifene, and droloxifen, which was confirmed in chromatin immunoprecipitation assays. Smad4 and ERα form a complex when ERα binds to the estrogen-responsive element within the estrogen target gene promoter. Importantly, the expression of Smad4 inhibits both antiestrogen-induced luciferase activity and estrogen downstream target gene transcription in breast cancer cells. Mapping of the interaction domains indicates that the activation function 1 (AF1) domain of ERα is essential for its interaction with Smad4, while the MH1 domain and linker region of Smad4 are essential for the interaction. Our findings represent a novel mechanism that TGF-β may regulate cell fate through Smad4-mediated cross-talk with estrogen. Antiestrogen compounds exhibit a variety of different effects in different tissues and are widely used for the treatment of osteoporosis, breast cancer, and other diseases. Upon examining the molecular mechanisms, we found that Smad4, a common signal transducer in the bone morphogenetic protein (BMP)/transforming growth factor-β (TGF-β) signaling pathway, functions as a transcription corepressor for human estrogen receptor α (ERα). Endogenous ERα was co-immunoprecipitated with Smad4, and the interaction was induced by antiestrogen ligands such as tamoxifen, raloxifene, and droloxifen, which was confirmed in chromatin immunoprecipitation assays. Smad4 and ERα form a complex when ERα binds to the estrogen-responsive element within the estrogen target gene promoter. Importantly, the expression of Smad4 inhibits both antiestrogen-induced luciferase activity and estrogen downstream target gene transcription in breast cancer cells. Mapping of the interaction domains indicates that the activation function 1 (AF1) domain of ERα is essential for its interaction with Smad4, while the MH1 domain and linker region of Smad4 are essential for the interaction. 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The of was with of the ligands as in The cell with and the with an The ERα and Smad4 in as the interaction two the interaction was to different with the treatment of different antiestrogen ligands the in however, the interaction was not in with which be to the expression of Smad4 in cells. antiestrogen ligands induced the interaction that the interaction Smad4 and ERα are by the function of the interaction Smad4 and a luciferase two of was with The of with or that transcription activity as The expression of Smad4 a of luciferase activity in with The was in a different SERMs exhibit transcription the antiestrogen compounds tamoxifen, raloxifene, and droloxifen also binding of ligands to ERα not in luciferase the of Smad4, however, the transcription by antiestrogen ligands to be the of luciferase of the with that SERMs the interaction Smad4 and that Smad4 as an ERα transcription and its activity is by different in not that transcriptional activation be by T. T. A. F. J. Biol. 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