MICRO RNA MEDIATED REGULATION OF MIEN1 IN PROSTATE CANCER

Purpose: MIEN1 is a membrane bound signaling molecule that triggers downstream signaling through the AKT/NF-𝛋B pathway by up-regulating key proteases and has a role in migration and invasion of cancer cells. The overall objective of this study is to identify the mechanisms that lead to the differential regulation of MIEN1 in normal and cancer cells. Though there are multiple mechanisms that are commonly studied, here, we propose to focus on the post-transcriptional regulation of MIEN1. Our data leads to the hypothesis that MIEN1 is post-transcriptionally regulated by a specific microRNA (miR) that is down-regulated in cancer, thus explaining aberrant increased expression of MIEN1 in cancer. Methods: We have performed miR in silico analysis, expression profiling, northern blotting, qPCRs, western blotting, luciferase reporter assays, migration and invasion assays, colony formation assays and stability assays to determine the effects of the miR on different aspects of MIEN1 regulation. Results: Our data indicate that MIEN1 is post-transcriptionally regulated by a specific miR which is lost in cancer cells. This miR is highly expressed in normal cells compared to a decrease in various cancer cells and this expression is inversely correlated to the expression of MIEN1. Introduction of the mimic (precursor) or inhibitor (antagomiR) led to a decrease or increase in MIEN1 expression respectively at both the RNA and protein levels. The downstream effectors were also similarly affected. The luciferase activity significantly reduced when the 3'UTR of MIEN1 was transfected with the target miR compared to the control miR, validating the direct binding of the miR. The miR decreases the migration and invasion of cells as well as the stability of MIEN1 RNA. There was a significant reduction in the colony formation capabilities and the morphology of the colonies differed in cells that were transfected with the miR along with a decrease in E-cadherin, suggesting potential involvement of the miRNA in reducing epithelial to mesenchymal transition and hence inhibiting metastasis. Conclusions: Our results demonstrate that aberrant expression of MIEN1 in cancer is attributed to a specific miR. We are currently exploring the potential of using this miR as a biomarker in prostate cancer. Since the importance of MIEN1 as a key signaling molecule in cancer is well established, understanding the mechanisms involved in its regulation will aid in designing novel and effective therapeutic strategies to treat cancer patients.