Regulation of the Selenocysteine Stress Response in Cancer Metastasis

癌症转移中硒代半胱氨酸应激反应的调节

• 批准号: 10223174
• 负责人: Leona Nease
• 金额: $ 4.6万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-14 至 2023-07-13
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10223174
• 关键词: Address; Amino Acids; Anabolism; Anticodon; Binding Proteins; Biochemical; Biological Assay; Biology; Biotin; Cancer Etiology; Cause of Death; Cell Line; Cell Survival; Cells; Cellular Stress; Cellular Stress Response; Cessation of life; Codon Nucleotides; Cysteine; Development; Diet; Disease; Endoplasmic Reticulum; Enzymes; Equilibrium; Event; Family; Genetic; Genetic Transcription; Goals; Human; Immunoprecipitation; In Vitro; Investigation; Laboratories; Link; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Medicine; Melanoma Cell; Metabolic; Metastatic Melanoma; Methods; Methylation; Methyltransferase; Modeling; Modification; Molecular; Molecular Chaperones; Molecular Conformation; Mus; Mutate; Neoplasm Metastasis; Nodule; Nucleosides; Oxidation-Reduction; Oxidative Stress; Patient-Focused Outcomes; Patients; Pharmacology; Primary Neoplasm; Protein Family; Proteins; Reactive Oxygen Species; Regulation; Reporter; Research Personnel; Ribose; Role; Science; Scientist; Selenium; Selenocysteine; Site-Directed Mutagenesis; Solid Neoplasm; Speed; Stress; Structure; Sulfur; Techniques; Terminator Codon; Testing; Therapeutic; Trans-Activators; Transfer RNA; Translational Regulation; Translations; Tumor-Derived; Uridine; Work; X-linked mental retardation 9; Yeasts; antioxidant enzyme; base; biological adaptation to stress; cancer cell; career; crosslink; effective therapy; glutathione peroxidase; in vivo; insight; liquid chromatography mass spectrometry; malignant breast neoplasm; melanoma; novel; patient derived xenograft model; protein complex; protein folding; rat Secp43 protein; selenocysteine-tRNA; selenoprotein; small hairpin RNA; tRNA Methyltransferases; thioredoxin reductase; tool

PROJECT SUMMARY The adaptations that metastasizing cancer cells undergo are not well understood, contributing to a lack of effective therapies, making metastasis the leading cause of cancer-related deaths. Genetic drivers of metastatic progression have not been identified, thus it is imperative to study the disease at the transcriptional and translational level where cells are able to make reversible adaptations that allow them to survive throughout the metastatic cascade. Our long-term goal is to find novel and targetable vulnerabilities in cancer metastasis by identifying these molecular and metabolic adaptations. Early work from our lab shows that metastasizing cells undergo high levels of oxidative stress. Selenocysteine, the 21st amino acid, is incorporated into a family of proteins that are involved in detoxifying reactive oxygen species and maintaining redox balance in the cell. Translation of selenoproteins under cellular stress is regulated by a single 2’-O-ribose methylation on the wobble uridine (Um34) of the selenocysteine tRNA (tRNASec), completed by an unknown methyltransferase. Using our lab’s patient-derived tumor model of melanoma metastasis, I will test the hypothesis that Um34 methylation is increased in metastasizing cells and that this modification increases cell survival under oxidative stress by regulating a subset of stress response selenoproteins. To address this question, I will utilize an established patient-derived melanoma model where metastasis is predictive of patient outcome in parallel with melanoma cell lines in vitro. In Aim 1, I will determine the functional role of Um34 in metastasis and the oxidative stress response. I will develop tools to measure Um34 levels and I will perturb the modification event by targeting related enzymes, tRNASec and selenium availability. In Aim 2, I will identify the unknown Um34 methyltransferase by testing a candidate protein and through targeted immunoprecipitation approaches. I will use mass spectrometry to quantify Um34 levels and identify related protein complexes. I will then characterize the selenocysteine stress response and through depletion of the identified methyltransferase, I will determine its functional role in metastasis and the oxidative stress response. This work will be completed at Weill Cornell Medicine where I will develop scientifically and professionally into an independent researcher. I will receive expert guidance throughout my project from my committee of world-renowned scientists whose expertise span the many fields covered in my proposal. Through this work I will master biochemical analytical techniques, modeling a complicated disease in vivo and clearly communicating my ideas and results – all of which will prepare me for a career as an independent investigator in academic science. Identifying novel targets in metastatic disease is an urgent therapeutic need and I believe that this work will not only find one, but expand the current understanding of how metastasizing cancer cells adapt and survive.

项目摘要 转移性癌细胞所经历的适应性尚不清楚，这导致了缺乏对癌细胞的免疫调节。 有效的治疗方法，使转移成为癌症相关死亡的主要原因。转移性肿瘤的遗传驱动因素 进展尚未确定，因此必须在转录和转录水平上研究该疾病， 在翻译水平上，细胞能够进行可逆的适应，使它们能够在整个过程中存活。 转移级联反应我们的长期目标是通过以下方法找到癌症转移中的新的和可靶向的弱点： 识别这些分子和代谢适应。我们实验室的早期工作表明转移细胞 经受高水平的氧化应激。硒代半胱氨酸，第21个氨基酸，被纳入一个家族， 参与解毒活性氧和维持细胞内氧化还原平衡的蛋白质。 在细胞应激下硒蛋白的翻译受摆动上的单个2 '-O-核糖甲基化调节 硒代半胱氨酸tRNA（tRNASec）的尿苷（Um 34），由未知的甲基转移酶完成。使用我们 实验室的黑色素瘤转移的患者源性肿瘤模型，我将测试Um 34甲基化 在转移细胞中增加，并且这种修饰增加了在氧化条件下的细胞存活。 通过调节应激反应硒蛋白的子集来应激。为了解决这个问题，我将使用一个 建立了患者来源的黑色素瘤模型，其中转移可预测患者结局， 黑色素瘤细胞系。在目的1中，我将确定Um 34在转移和氧化损伤中的功能作用。 应激反应我将开发工具来测量Um 34水平，我将通过瞄准来干扰修改事件， 相关酶、tRNA Sec和硒的可用性。在目标2中，我将鉴定未知的Um 34甲基转移酶 通过测试候选蛋白和通过靶向免疫沉淀方法。我会用质量 光谱法来定量Um 34水平并鉴定相关蛋白质复合物。然后我将描述 硒代半胱氨酸应激反应，并通过确定的甲基转移酶消耗，我将确定其 在转移和氧化应激反应中的功能作用。这项工作将在威尔康奈尔完成 医学，我将发展成为一个独立的研究人员科学和专业。我将收到 在我的整个项目中，我的世界知名科学家委员会提供了专家指导， 我的提案中涉及的许多领域。通过这项工作，我将掌握生化分析技术， 在体内模拟复杂的疾病，并清楚地传达我的想法和结果-所有这些都将 为我作为学术科学独立调查员的职业生涯做好准备。确定新的目标， 转移性疾病是一个迫切的治疗需求，我相信这项工作不仅会找到一个，但扩大 目前对转移癌细胞如何适应和生存的理解。