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/摘要 癌症是美国第二大死亡原因。肿瘤患者的总生存期 仍然局限于原始组织的局部性相对较高，而估计所有癌症的90％ 相关的死亡是由于转移到次要部位的转移。这强调了 识别促进转移的信号通路。线粒体是多功能细胞器 在控制ATP生产以外的许多细胞过程中的重要作用，并因其认可 在肿瘤发生和转移中的作用。该项目旨在定义促进肿瘤细胞的新型机制 通过先前未鉴定的线粒体Rho GTPase之间的侵袭和转移 2（miro2）和非典型肌球蛋白IXB（myo9b）。 MIRO2是一种外膜膜蛋白，是Ras GTPase的Miro subfigily的一部分。 米罗首先在神经元中进行了特征 线粒体。几个小组已显示MiRO2可用于线粒体贩运 许多非神经细胞类型。我们的实验室先前已经表明MiRO2 mRNA富含肿瘤性 与正常组织相比，许多肿瘤类型的组织。此外，我们已经在前列腺癌中显示 MiRO2在肿瘤细胞的生存力，锚定依赖性和非依赖性生长以及肿瘤生长中很重要 体内。尽管有最初的证据，但MiRO2是否仅影响原发性肿瘤，或者是否存在 在转移中也很重要。我的最初结果表明，Miro2的丧失会降低肿瘤的侵入性能力 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，我已经表明 在最高的新确定的MiRO2绑定伙伴中，损失Myo9b将侵入性能力降低到最大的能力 程度。众所周知，Myo9b通过将其定位到迁移细胞的前缘并灭活来控制细胞运动。 通过MyO9b的Rho GTPase激活蛋白（GAP）结构域的RhoA。令人兴奋的是，我表明了两个miro2的损失 Myo9b导致活跃的RhoA增加。鉴于此证据，我假设MiRO2促进了肿瘤细胞 通过积极调节myo9b间隙活性的侵袭和转移。 在此提案中，我将1）确定miro2在肿瘤细胞侵袭和转移中的作用以及2） 确定MIRO2调节肿瘤细胞浸润的机制。该建议将利用许多模型 包括但不限于：2D/3D体外入侵系统，迁移细胞的活细胞成像，体内乳房 癌症转移模型，无细胞GTPase分析，共免疫沉淀和接近结扎分析。 总体而言，该提案的目的是作为开发新疗法以靶向的基础 转移性疾病。