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: 10464387
• 负责人: Dillon Boulton
• 金额: $ 3.62万
• 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/10464387
• 关键词: 3-Dimensional; 4T1; Ablation; Address; Binding; Binding Proteins; Biological Assay; Breast Cancer Model; Breast Cancer Patient; Breast Melanoma; Breast cancer metastasis; Calcium; 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; Knockout Mice; Ligation; MDA MB 231; Malignant Neoplasms; Malignant neoplasm of pancreas; Malignant neoplasm of prostate; Mass Spectrum Analysis; Matrix Metalloproteinases; Measurement; Mediating; Messenger RNA; Metastatic to; 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; Pharmacology; 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; Tumor Cell Invasion; 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; neoplastic cell; new therapeutic target; novel; novel therapeutics; 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 仍然局限于起源组织的癌症仍然相对较高，而据估计，90%的癌症 相关的死亡是由于转移到第二部位。这突出了以下方面的重要性： 识别促进转移的信号通路。线粒体是多功能的细胞器， 在调节ATP产生之外的许多细胞过程中起重要作用，并因其 在肿瘤发生和转移中的作用。该项目旨在确定一种新的机制，促进肿瘤细胞 线粒体Rho GT3与肿瘤侵袭转移的信号通路 2（MIRO 2）和非典型肌球蛋白IXB（MYO 9 B）。 MIRO 2是一种线粒体外膜蛋白，是Ras GTP酶MIRO亚家族的一部分。 MIRO首先在参与顺行和逆行运输的神经元中进行表征 of mitochondria线粒体. MIRO 2已经被几个研究小组证明是线粒体运输的关键。 许多非神经细胞类型。我们的实验室先前已经表明，MIRO 2 mRNA在致瘤性细胞中富集， 与正常组织相比，许多肿瘤类型的组织。另外，我们在前列腺癌中发现， MIRO 2在肿瘤细胞活力、锚定依赖性和非依赖性生长以及肿瘤生长中是重要的 in vivo.尽管有这一初步证据，但仍不清楚MIRO 2是否专门影响原发性肿瘤，或者是否 在转移中也很重要。我的初步结果表明，MIRO 2的丢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、胰腺癌和前列腺癌。此外，我还展示了 在最新鉴定的MIRO 2结合配偶体中，MYO 9 B的缺失最大程度地降低了侵入能力 程度已知MYO 9 B通过定位于迁移细胞的前缘并使其失活来控制细胞运动性。 RhoA通过MYO 9 B的Rho GTP酶激活蛋白（GAP）结构域。令人兴奋的是，我表明，损失的两个MIRO 2 和MYO 9 B导致活性RhoA增加。鉴于这一证据，我假设MIRO 2促进肿瘤细胞 MYO 9 B GAP活性与肿瘤侵袭转移密切相关。 在这个提案中，我将1）确定MIRO 2在肿瘤细胞侵袭和转移中的作用，2） 确定MIRO 2调节肿瘤细胞侵袭的机制。该提案将利用许多模型 包括但不限于：2D/3D体外侵入系统、迁移细胞的活细胞成像、体内乳腺癌 癌症转移模型、无细胞GT3测定、免疫共沉淀和邻位连接测定。 总的来说，这项提案的目标是作为开发新疗法的基础， 转移性疾病