MicroRNA profiling of the pubertal mouse mammary gland identifies miR-184 as a candidate breast tumour suppressor gene
INTRODUCTION: The study of mammalian development has offered many insights into the molecular aetiology of cancer. We previously used analysis of mammary morphogenesis to discover a critical role for GATA-3 in mammary developmental and carcinogenesis. In recent years an important role for microRNAs (miRNAs) in a myriad of cellular processes in development and in oncogenesis has emerged. METHODS: microRNA profiling was conducted on stromal and epithelial cellular subsets microdissected from the pubertal mouse mammary gland. miR-184 was reactivated by transient or stable overexpression in breast cancer cell lines and examined using a series of in vitro (proliferation, tumour-sphere and protein synthesis) assays. Orthotopic xenografts of breast cancer cells were used to assess the effect of miR-184 on tumourigenesis as well as distant metastasis. Interactions between miR-184 and its putative targets were assessed by quantitative PCR, microarray, bioinformatics and 3' untranslated region Luciferase reporter assay. The methylation status of primary patient samples was determined by MBD-Cap sequencing. Lastly, the clinical prognostic significance of miR-184 putative targets was assessed using publicly available datasets. RESULTS: A large number of microRNA were restricted in their expression to specific tissue subsets. MicroRNA-184 (miR-184) was exclusively expressed in epithelial cells and markedly upregulated during differentiation of the proliferative, invasive cells of the pubertal terminal end bud (TEB) into ductal epithelial cells in vivo. miR-184 expression was silenced in mouse tumour models compared to non-transformed epithelium and in a majority of breast cancer cell line models. Ectopic reactivation of miR-184 inhibited the proliferation and self-renewal of triple negative breast cancer (TNBC) cell lines in vitro and delayed primary tumour formation and reduced metastatic burden in vivo. Gene expression studies uncovered multi-factorial regulation of genes in the AKT/mTORC1 pathway by miR-184. In clinical breast cancer tissues, expression of miR-184 is lost in primary TNBCs while the miR-184 promoter is methylated in a subset of lymph node metastases from TNBC patients. CONCLUSIONS: These studies elucidate a new layer of regulation in the PI3K/AKT/mTOR pathway with relevance to mammary development and tumour progression and identify miR-184 as a putative breast tumour suppressor.
|ISBN||1465-542X (Electronic) 1465-5411 (Linking)|
|Authors||Phua, Y. W.; Nguyen, A.; Roden, D. L.; Elsworth, B.; Deng, N.; Nikolic, I.; Yang, J.; McFarland, A.; Russell, R.; Kaplan, W.; Cowley, M. J.; Nair, R.; Zotenko, E.; O'Toole, S.; Tan, S. X.; James, D. E.; Clark, S. J.; Kouros-Mehr, H.; Swarbrick, A.;|
|Responsible Garvan Author|
|Publisher Name||BREAST CANCER RES|
|URL link to publisher's version||http://www.ncbi.nlm.nih.gov/pubmed/26070602|
|OpenAccess link to author's accepted manuscript version||https://publications.gimr.garvan.org.au/open-access/13059|