Investigating Transcriptional Elongation and Nuclear RNA Surveillance in Melanoma

研究黑色素瘤的转录延伸和核 RNA 监视

• 批准号: 10215098
• 负责人: Megan Leigh Insco
• 金额: $ 5.95万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10215098
• 关键词: Affect; Aggressive behavior; Animal Model; Area; Award; Binding; Biological Assay; Boston; Cancer Biology; Cell Culture System; Cell Proliferation; Cells; Cellular Stress; Clinical; Collaborations; Complex; Cyclin-Dependent Kinases; Cyclins; Cytoplasm; Dana-Farber Cancer Institute; Data; Dissection; Dominant-Negative Mutation; Down-Regulation; Drug Targeting; Environment; Epigenetic Process; Failure; Family; Gene Expression; Genes; Genetic; Genetic Transcription; Grant; Growth; Heat shock proteins; Hematologist; Human; Image; Immune; Immunotherapy; In Vitro; Laboratories; Lead; Leadership; Learning; Lysosomes; Mainstreaming; Malignant Neoplasms; Measures; Melanoma Cell; Mentors; Metastatic Melanoma; Modeling; Molecular; Mutate; Mutation; Nuclear; Nuclear RNA; Oncogenic; Oncologist; Pathway interactions; Patients; Pediatric Hospitals; Peptides; Phenotype; Phosphorylation; Phosphotransferases; Phylogenetic Analysis; Pigments; Prognosis; Progressive Disease; Proteins; Proteomics; Publishing; RNA; Recurrence; Refractory; Reporter; Research; Research Personnel; Resistance; Scientist; Skin Cancer; Stress; System; Techniques; Testing; Therapeutic; Time; Training; Transcription Elongation; Translating; Translations; Work; Zebrafish; endoplasmic reticulum stress; exosome; in vivo; kinase inhibitor; medical schools; melanocyte; melanoma; member; mutant; premature; protein expression; skills; stress granule; transcriptome sequencing; transcriptomics; tumor; tumorigenesis

Project Summary Metastatic melanoma, a cancer derived from pigment-producing melanocytes, is the deadliest type of skin cancer. Metastatic melanoma that is refractory or resistant to current therapies has a poor prognosis. Melanoma growth and therapy resistance is dependent on dysregulated gene expression, and gene expression in melanoma remains poorly understood. The transcriptional cyclin dependent kinases (CDKs) are a phylogenetically distinct group of kinases that directly regulate transcriptional subprocesses. I found that CDK13 is mutated in melanoma; these mutations are kinase-dead and act by interfering with WT CDK13 function (dominant negative). Downregulation or mutation in CDK13 is associated with poor prognosis in metastatic melanoma patients, expression of mutant CDK13 in a zebrafish model expedites melanoma onset, and expression of mutant CDK13 in human melanoma cells causes the cells to be more proliferative. Mutant CDK13 fails to phosphorylate a protein that activates nuclear degradation of prematurely truncated ‘junk’ RNAs (ptRNAs). Subsequently, ptRNAs accumulate, are exported to the cytoplasm, and are translated into short proteins. This work is newly published on BioRxiv. I have also recently identified recurrent truncations in two nuclear RNA surveillance members: ZFC3H1 and ZC3H18. Here, I propose to further investigate the mechanism of mutant-CDK13 oncogenesis and nuclear RNA surveillance in melanoma. In Aim1, I plan to a) determine how protein stress pathways are affected in CDK13-mutant cells and b) to test whether truncated protein expression is sufficient to recapitulate the CDK13-mutant phenotype in zebrafish melanoma. In Aim 2, I will investigate whether recurrent truncating mutations in two nuclear surveillance complex members (ZFC3H1 and/or ZC3H18) cause more aggressive melanoma and if so, how nuclear RNA surveillance is impacted using proteomics and 3’ transcriptomics. This work has the potential to open a new cancer biology field and to lead to therapeutic strategies for cancers with deficient nuclear RNA surveillance. This research will be conducted in the laboratory of Dr. Leonard Zon, a renowned hematologist/oncologist and cancer biologist who has developed the zebrafish into a mainstream model organism for melanoma research. Under the guidance of Dr. Zon and an exceptional mentoring committee, that is personally invested in my growth into an independent investigator, this award will provide the necessary protected time to develop skills to study nuclear RNA surveillance in melanoma as an independent investigator. As recommended by this committee, I have developed a new collaboration with Dr. Steve Gygi in order to learn cutting-edge proteomic techniques. My rigorous training plan will help me build scientific and leadership skills necessary to succeed when I transition to independence. The research and clinical environment at Boston Children’s Hospital, Dana Farber Cancer Institute, and Harvard Medical School is the ideal environment to develop into a successful independent scientist who investigates nuclear RNA surveillance in melanoma.

项目概要 转移性黑色素瘤是一种源自产生色素的黑色素细胞的癌症，是最致命的皮肤类型 癌症。对当前疗法难治或耐药的转移性黑色素瘤预后较差。黑色素瘤 生长和治疗耐药性取决于基因表达失调，而基因表达在 黑色素瘤仍然知之甚少。转录细胞周期蛋白依赖性激酶 (CDK) 是 直接调节转录子过程的系统发育上不同的激酶组。我发现CDK13 在黑色素瘤中发生突变；这些突变是激酶死亡的，并通过干扰 WT CDK13 功能发挥作用 （显性阴性）。 CDK13 的下调或突变与转移性乳腺癌的不良预后相关 黑色素瘤患者，斑马鱼模型中突变 CDK13 的表达会加速黑色素瘤的发病，以及 人类黑色素瘤细胞中突变型 CDK13 的表达导致细胞更加增殖。突变CDK13 未能磷酸化激活过早截短的“垃圾”RNA 核降解的蛋白质 （ptRNA）。随后，ptRNA 积累，被输出到细胞质，并被翻译成短链。 蛋白质。该工作最新发表在 BioRxiv 上。我最近还发现了两个重复的截断 核RNA监视成员：ZFC3H1和ZC3H18。在此，我建议进一步研究其机制 黑色素瘤中突变型 CDK13 肿瘤发生和核 RNA 监测的研究。在目标 1 中，我计划 a) 确定如何 CDK13 突变细胞中的蛋白质应激途径受到影响，b) 测试蛋白质表达是否被截短 足以重现斑马鱼黑色素瘤中的 CDK13 突变表型。在目标 2 中，我将调查 两个核监视复合体成员（ZFC3H1 和/或 ZC3H18）是否反复出现截短突变 导致更具侵袭性的黑色素瘤，如果是这样，如何使用蛋白质组学和 3' 来影响核 RNA 监测 转录组学。这项工作有可能开辟一个新的癌症生物学领域并带来治疗方法 针对核 RNA 监测不足的癌症的策略。 这项研究将在著名血液学家/肿瘤学家 Leonard Zon 博士的实验室中进行 癌症生物学家，将斑马鱼发展成为黑色素瘤的主流模式生物 研究。在 Zon 博士和杰出的指导委员会的指导下， 我成长为一名独立调查员，该奖项将为我提供必要的受保护时间来发展技能 作为独立研究者研究黑色素瘤的核 RNA 监测。据此推荐 委员会，我与 Steve Gygi 博士建立了新的合作关系，以学习前沿的蛋白质组学 技术。我严格的培训计划将帮助我培养成功所需的科学和领导技能 当我过渡到独立时。达纳波士顿儿童医院的研究和临床环境 法伯癌症研究所和哈佛医学院是发展成为成功人士的理想环境 研究黑色素瘤核 RNA 监测的独立科学家。