The role of MIRO2 in tumor cell invasion and metastasis

MIRO2在肿瘤细胞侵袭和转移中的作用

• 批准号: 10704512
• 负责人: Dillon Boulton
• 金额: $ 3.72万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10704512
• 关键词: 3-Dimensional; 4T1; Ablation; Address; Binding; Binding Proteins; Biological Assay; Breast; Breast Cancer Model; Breast Cancer Patient; Breast cancer metastasis; Calcium; Calcium Binding; Cause of Death; Cell Death; Cell Survival; Cell physiology; Cells; Cessation of life; Co-Immunoprecipitations; Data; Detection; Development; Disease; Distal; Dominant-Negative Mutation; EF-Hand Domain; GTPase-Activating Proteins; Goals; Growth; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Immunohistochemistry; Immunoprecipitation; In Vitro; Injections; Invaded; Knockout Mice; Ligation; MDA MB 231; Malignant Neoplasms; Malignant neoplasm of prostate; Mass Spectrum Analysis; Matrix Metalloproteinases; Measurement; Mediating; Messenger RNA; Mitochondria; Mitochondrial Membrane Protein; Modeling; Molecular; Myosin ATPase; Myosin S-2; Neoplasm Metastasis; Neuroglia; Neurons; Normal tissue morphology; Organ; Organelles; Outer Mitochondrial Membrane; Pancreas; Patients; Phenocopy; Phenotype; Primary Neoplasm; Process; Production; Proliferation Marker; Prostate; Recombinant Proteins; Role; Sampling; Secondary to; Signal Pathway; Signal Transduction; Site; Small Interfering RNA; System; Tertiary Protein Structure; Testing; Tissues; Transfection; Tumor Cell Invasion; Tumor Promotion; United States; Work; cell motility; cell type; hazard; in vitro Assay; in vivo; in vivo Model; in vivo imaging system; interest; knock-down; live cell imaging; mRNA Expression; melanoma; neoplastic cell; new therapeutic target; novel; novel therapeutics; pancreatic cancer cells; pharmacologic; prostate cancer cell line; protein function; rho GTP-Binding Proteins; rho GTPase-activating protein; small hairpin RNA; targeted treatment; therapeutic target; trafficking; tumor; tumor growth; tumorigenesis; tumorigenic

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.

项目摘要/总结 癌症是美国第二大死因。肿瘤患者的总生存期 仍局限于起源组织的癌症仍然相对较高，而据估计 90% 的癌症 相关死亡是由于转移到二级部位所致。这强调了重要性 识别促进转移的信号通路。线粒体是多功能细胞器 在调节 ATP 产生之外的许多细胞过程中发挥着重要作用，并因其 在肿瘤发生和转移中的作用。该项目旨在定义一种促进肿瘤细胞生长的新机制 通过线粒体 Rho GTPase 之间先前未识别的信号通路进行侵袭和转移 2 (MIRO2) 和非典型肌球蛋白 IXB (MYO9B)。 MIRO2 是一种线粒体外膜蛋白，属于 Ras GTPases MIRO 亚家族的一部分。 MIRO 首先在神经元中被表征，它们参与顺行和逆行运输 线粒体。多个研究小组已证明 MIRO2 对于线粒体运输而言是可有可无的 许多非神经元细胞类型。我们的实验室之前已经证明 MIRO2 mRNA 在致瘤性中富集 与正常组织相比，许多肿瘤类型中的组织。此外，我们还发现前列腺癌 MIRO2 在肿瘤细胞活力、贴壁依赖性和非依赖性生长以及肿瘤生长中很重要 体内。尽管有这些初步证据，但仍不清楚 MIRO2 是否只影响原发肿瘤，或者是否会影响原发肿瘤。 在转移中也很重要。我的初步结果表明 MIRO2 的缺失降低了肿瘤的侵袭能力 来自许多肿瘤类型的细胞，包括乳腺癌、黑色素瘤、胰腺和前列腺。此外，我还展示了 在新发现的最重要的 MIRO2 结合伴侣中，MYO9B 的丢失将最大程度地降低侵袭能力 程度。已知 MYO9B 通过定位到迁移细胞的前缘并失活来控制细胞运动 RhoA 通过 MYO9B 的 Rho GTP 酶激活蛋白 (GAP) 结构域。令人兴奋的是，我展示了 MIRO2 的丢失 和 MYO9B 导致活性 RhoA 增加。鉴于这一证据，我假设 MIRO2 促进肿瘤细胞 通过正向调节MYO9B GAP活性来抑制侵袭和转移。 在这个提案中，我将 1) 确定 MIRO2 在肿瘤细胞侵袭和转移中的作用，2) 确定 MIRO2 调节肿瘤细胞侵袭的机制。该提案将使用许多模型 包括但不限于：2D/3D体外侵袭系统、迁移细胞的活细胞成像、体内乳腺 癌症转移模型、无细胞 GTP 酶测定、免疫共沉淀和邻近连接测定。 总体而言，该提案的目标是作为开发靶向新疗法的基础 转移性疾病。