Downregulation of the tumor-suppressor miR-16 via progestin-mediated oncogenic signaling contributes to breast cancer development

RIVAS MA, VENTURUTTI L, HUANG YW, SCHILLACI R, HUANG TH, ELIZALDE PV.

BREAST CANCER RESEARCH

BIOMED CENTRAL LTD

Lugar: Londres; Año: 2012 vol. 14 p. 1 - 20

1465-5411

AbstractIntroduction: Experimental and clinical evidence points to a critical role of progesterone and the nuclearprogesterone receptor (PR) in controlling mammary gland tumorigenesis. However, the molecular mechanisms ofprogesterone action in breast cancer still remain elusive. On the other hand, micro RNAs (miRNAs) are shortribonucleic acids which have also been found to play a pivotal role in cancer pathogenesis. The role of miRNA inprogestin-induced breast cancer is poorly explored. In this study we explored progestin modulation of miRNAexpression in mammary tumorigenesis.Methods: We performed a genome-wide study to explore progestin-mediated regulation of miRNA expression inbreast cancer. miR-16 expression was studied by RT-qPCR in cancer cell lines with silenced PR, signal transducerand activator of transcription 3 (Stat3) or c-Myc, treated or not with progestins. Breast cancer cells were transfectedwith the precursor of miR-16 and proliferation assays, Western blots or in vivo experiments were performed. Targetgenes of miR-16 were searched through a bioinformatical approach, and the study was focused on cyclin E.Reporter gene assays were performed to confirm that cyclin E 3?UTR is a direct target of miR-16.Results: We found that nine miRNAs were upregulated and seven were downregulated by progestin in mammarytumor cells. miR-16, whose function as a tumor suppressor in leukemia has already been shown, was identified asone of the downregulated miRNAs in murine and human breast cancer cells. Progestin induced a decrease in miR-16 levels via the classical PR and through a hierarchical interplay between Stat3 and the oncogenic transcriptionfactor c-Myc. A search for miR-16 targets showed that the CCNE1 gene, encoding the cell cycle regulator cyclin E,contains conserved putative miR-16 target sites in its mRNA 3? UTR region. We found that, similar to the molecularmechanism underlying progestin-modulated miR-16 expression, Stat3 and c-Myc participated in the induction ofcyclin E expression by progestin. Moreover, overexpression of miR-16 abrogated the ability of progestin to inducecyclin E upregulation, revealing that cyclin E is a novel target of miR-16 in breast cancer. Overexpression of miR-16also inhibited progestin-induced breast tumor growth in vitro and in vivo, demonstrating for the first time, a rolefor miR-16 as a tumor suppressor in mammary tumorigenesis. We also found that the ErbB ligand heregulin (HRG)downregulated the expression of miR-16, which then participates in the proliferative activity of HRG in breasttumor cells.Conclusions: In this study, we reveal the first progestin-regulated miRNA expression profile and identify a novelrole for miR-16 as a tumor suppressor in progestin- and growth factor-induced growth in breast cancer.* Correspondence: