Coilin is a Mediator of m6A RNA Methylation

Coilin 是 m6A RNA 甲基化的介质

• 批准号: 10678140
• 负责人: Douglas McLaurin
• 金额: $ 4.77万
• 依托单位: UNIVERSITY OF MISSISSIPPI MED CTR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-17 至 2025-03-16
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10678140
• 关键词: 20 year old; Acute Lymphocytic Leukemia; Apoptotic; Binding; Biogenesis; CRISPR/Cas technology; Cause of Death; Cell Proliferation; Cell Survival; Cells; Child; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Data; Development; Elements; Embryonic Induction; Engineering; Future; Gene Expression; Hyperactivity; Incidence; Inhibition of Apoptosis; Investigation; Knock-out; Laboratory Finding; Link; Malignant Neoplasms; Mediating; Mediator; Methyltransferase; MicroRNAs; Microprocessor; Modeling; Modification; Molecular; Mus; NF-kappa B; Nuclear; Nucleoplasm; Pathway interactions; Patients; Phenotype; Plants; Post-Translational Protein Processing; Process; Prognostic Factor; Proliferating; Proteins; Publishing; RNA; RNA methylation; Reader; Regulation; Reporting; Ribonucleoproteins; Role; Signal Pathway; Signal Transduction; Small Nuclear Ribonucleoproteins; Telomerase; Testing; Therapeutic; Transformed Cell Line; Untranslated RNA; Up-Regulation; Work; Zebrafish; biomarker development; cancer cell; cell type; fly; knock-down; novel; prognostic; protein biomarkers; therapeutic development; therapy resistant; tumor progression; tumorigenesis; 20 year old; Acute Lymphocytic Leukemia; Apoptotic; Binding; Biogenesis; CRISPR/Cas technology; Cause of Death; Cell Proliferation; Cell Survival; Cells; Child; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Data; Development; Elements; Embryonic Induction; Engineering; Future; Gene Expression; Hyperactivity; Incidence; Inhibition of Apoptosis; Investigation; Knock-out; Laboratory Finding; Link; Malignant Neoplasms; Mediating; Mediator; Methyltransferase; MicroRNAs; Microprocessor; Modeling; Modification; Molecular; Mus; NF-kappa B; Nuclear; Nucleoplasm; Pathway interactions; Patients; Phenotype; Plants; Post-Translational Protein Processing; Process; Prognostic Factor; Proliferating; Proteins; Publishing; RNA; RNA methylation; Reader; Regulation; Reporting; Ribonucleoproteins; Role; Signal Pathway; Signal Transduction; Small Nuclear Ribonucleoproteins; Telomerase; Testing; Therapeutic; Transformed Cell Line; Untranslated RNA; Up-Regulation; Work; Zebrafish; biomarker development; cancer cell; cell type; fly; knock-down; novel; prognostic; protein biomarkers; therapeutic development; therapy resistant; tumor progression; tumorigenesis

PROJECT SUMMARY Survival and proliferation are fundamental cell activities that are exploited during oncogenesis. A key cellular pathway that supports survival and proliferation is the nuclear factor-Kappa B (NF-κB) signaling pathway, which is also hyperactive in cancer cells and is a noted element that contributes to therapeutic resistance, a phenomenon observed in various cancers, including acute lymphoblastic leukemia (ALL). With an incidence of about 30 cases per million, ALL is the most common cancer among children and the most frequent cause of death from cancer before 20 years of age. In a recent study, it was shown that the elevated expression of coilin, the Cajal body (CB) marker protein, is a negative prognostic factor in ALL patients. Recent work by our lab has discovered that coilin is a positive regulator in the biogenesis of microRNAs (miRNAs), a class of noncoding RNAs that negatively regulate gene expression. However, the mechanism for this function is unclear. Another positive regulator of miRNA biogenesis is N6-methyladenosine (m6A) RNA methylation, a reversible modification that is installed by writers, removed by erasers, and acted on by readers. Our lab hypothesized that coilin was influencing miRNA biogenesis through the regulation of m6A RNA methylation and we have generated exciting preliminary data showing that coilin suppression results in a decrease in the expression of m6A writer METTL3, reader hnRNPA2B1, and eraser ALKBH5 proteins, as well as a their mislocalization within the cell and a reduction in global m6A RNA levels. Additionally, work published by our lab found that coilin positively regulates the expression of miR-517, a NF-κB activator. Since the NF-κB pathway is both regulated by and a regulator of the biogenesis of several miRNAs, and in consideration of previously published work on coilin, we have generated the central hypothesis that coilin supports m6A RNA methylation through which it regulates miRNA biogenesis and facilitates the survival and proliferation of the cell. This hypothesis will be tested with the use of various primary and transformed cell lines, and our newly generated, CRISPR/Cas9-engineered coilin zebrafish models. To test our central hypothesis, two specific aims have been generated for which we will test the hypotheses that: 1) Coilin directs the optimal synthesis, posttranslational modification, and localization of m6A RNA Writers, Readers, and Erasers; and 2) Coilin mediates m6A RNA methylation by which miRNA biogenesis and survival & proliferative pathways are regulated. This work will characterize three novel CRISPR-engineered zebrafish coilin lines while also investigating a novel pathway by which coilin mediates m6A RNA methylation and regulates miRNA biogenesis. Findings in this proposal will highlight a novel role for coilin in facilitating survival and proliferation, as well as a potential prognostic or therapeutic role for coilin in cancers, such as ALL.

项目摘要 生存和增殖是在肿瘤发生期间被利用的基本细胞活性。钥匙细胞 支持生存和增殖的途径是核因子-kappa B（NF-κB）信号通路，该途径 在癌细胞中也多动，是有助于治疗性耐药性的著名元素 在各种癌症中观察到的现象，包括急性淋巴细胞白血病（ALL）。发生 大约每百万案例约30例，这是儿童中最常见的癌症，也是最常见的原因 20岁之前的癌症死亡。在最近的一项研究中，结果表明，卷轴的表达升高， Cajal体（CB）标记蛋白是所有患者的负预后因素。我们实验室的最近工作 发现螺旋蛋白是microRNA（miRNA）的生物发生的阳性调节剂，这是一类非编码 负调控基因表达的RNA。但是，此功能的机制尚不清楚。其他 miRNA生物发生的阳性调节剂是N6-甲基腺苷（M6A）RNA甲基化，可逆修饰 这是由作家安装的，由橡皮擦删除，并由读者采取行动。我们的实验室假设卷轴是 通过调节M6A RNA甲基化影响miRNA生物发生，我们产生了令人兴奋的 初步数据表明，螺丝蛋白抑制会导致M6A作者Mettl3的表达降低， 读取器HnRNPA2B1和橡皮擦ALKBH5蛋白，以及在细胞中的错误定位和A 全球M6A RNA水平的降低。此外，我们的实验室发布的工作发现，卷蛋白会积极调节 miR-517，NF-κB活化剂的表达。由于NF-κB途径既由 几种miRNA的生物发生，并考虑到先前发表的有关Coilin的工作，我们有 产生了中心假设，即卷蛋白支持M6a RNA甲基化，它通过该假设调节miRNA 生物发生并促进细胞的存活和增殖。该假设将通过使用 各种主要和转化的细胞系，以及我们新生成的CRISPR/CAS9工程螺旋蛋白斑马鱼 型号。为了检验我们的中心假设，已经产生了两个具体目标，我们将测试 假设：1）卷轴指导M6A的最佳合成，翻译后修饰和定位 RNA作家，读者和橡皮擦；和2）卷蛋白介导m6a RNA甲基化，miRNA生物发生 调节生存和增殖途径。这项工作将以三种小说CRIS工程为特征 斑马鱼线圈线还研究了一条新型途径，该途径通过该途径介导M6a RNA甲基化 并调节miRNA生物发生。该提案中的发现将突出卷轴在促进中的新作用 生存和增殖，以及在癌症中的卷蛋白的潜在预后或治疗作用，例如全部。