ITAM PHOSPHORYLATION OF THE ADAPTER PROTEIN MIEN1 CONTRIBUTES TO ITS MIGRATION AND INVASION POTENTIAL

Purpose: Metastasis is one of the major causes of treatment failure in cancer patients. This process involves cell migration, stromal invasion, intravasation into the circulatory system leading to cancer proliferation. Identifying metastasis promoting and suppressing genes and their mechanisms of action will provide new insights into the pathogenesis and management of cancer. Previously known as C17orf37 or C35, Migration and Invasion enhancer 1 (MIEN1) is a novel gene that is over-expressed in a wide range of tumors. MIEN1 has been reported to be a critical regulator of cell migration and invasion. MIEN1 has a prenylation motif and an immunoreceptor tyrosine-based activation motif (ITAM), commonly found in signaling chains and receptors. The presence of a prenylation and ITAM domains in MIEN1 suggests that prenylation and/or tyrosine phosphorylation of the ITAM motif are important for its functions. The objective of the present study is to elucidate the molecular mechanisms through which MIEN1 promotes migration in breast cancer and whether or not tyrosine phosphorylation is important. Methods: We knockdown the expression of MIEN1 in MDA-MB231 and MCF10CA1 by siRNA and compared the migration of these cells with their respective controls. To study the importance of tyrosine phosphorylation of MIEN1- ITAM in migration and invasion, NIH3T3 were stably transfected with wild type MIEN1 or its phospho-mutants and used for migration and invasion assays. Results: Silencing of MIEN1 expression in MDA-MB231 and MCF10CA1 led to a significant decrease in breast cancer cell migration, conforming to our previous results in prostate cancer cells. Analyses of in-vitro migration and invasion assays revealed that NIH3T3 cells over-expressing MIEN1 phospho mutants failed to induce significant migration and invasion when compared with NIH3T3 over-expressing MIEN1 wild type. Conclusions: Our results show that MIEN1 plays an important role in the migration and invasion of breast cancer cells and tyrosine phosphorylation of MIEN1-ITAM significantly contributes to its functions