The role of MIRO2 in tumor cell invasion and metastasis

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Project Abstract/Summary Cancer is the second leading cause of death in the United States. Overall survival for patients with tumors that remain localized to the tissue of origin remain relatively high, while it is estimated that 90% of all cancer related deaths are due to metastatic dissemination to secondary sites. This underscores the importance of identifying signaling pathways that promote metastasis. Mitochondria are multifunctional organelles with important roles in regulating many cellular processes outside of ATP production and are recognized for their roles in tumorigenesis and metastasis. This project seeks to define a novel mechanism that promotes tumor cell invasion and metastasis through a previously unidentified signaling pathway between Mitochondrial Rho GTPase 2 (MIRO2) and atypical myosin IXB (MYO9B). MIRO2 is an outer-mitochondrial membrane protein that is a part of the MIRO subfamily of Ras GTPases. MIROs were first characterized in neurons where they are involved in the anterograde and retrograde trafficking of mitochondria. MIRO2 has been shown by several groups to be dispensable for mitochondrial trafficking in many non-neuronal cell types. Our lab has previously shown that MIRO2 mRNA is enriched in tumorigenic tissues in many tumor types when compared to normal tissue. Additionally, we have shown in prostate cancer that MIRO2 is important in tumor cell viability, anchorage-dependent and -independent growth, and tumor growth in vivo. Despite this initial evidence, it remains unknown if MIRO2 exclusively impacts the primary tumor or if this is also important in metastasis. My initial results show that loss of MIRO2 reduces the invasive capacity of tumor cells from many tumor types including breast, melanoma, pancreas, and prostate. Furthermore, I have shown out of the top newly identified MIRO2 binding partners, loss of MYO9B reduces invasive capacity to the greatest extent. MYO9B is known to control cell motility by localizing to the leading edge of migrating cells and inactivating RhoA through MYO9B’s Rho GTPase activating protein (GAP) domain. Excitingly, I show that loss of both MIRO2 and MYO9B result in increased active RhoA. Given this evidence, I hypothesize MIRO2 promotes tumor cell invasion and metastasis through positively regulating MYO9B GAP activity. In this proposal I will 1) determine the role of MIRO2 in tumor cell invasion and metastasis and 2) determine the mechanism in which MIRO2 regulates tumor cell invasion. This proposal will utilize many models including, but not limited to: 2D/3D in vitro invasion systems, live cell imaging of migrating cells, in vivo breast cancer metastasis models, cell-free GTPase assays, co-immunoprecipitation, and proximity ligation assays. Overall, the goal of this proposal is to serve as the basis for the development of novel therapeutics to target metastatic disease.