Biology of RNA G-quadruplexes

RNA G 四链体的生物学

基本信息

批准号：
10557098
负责人：
Pavel Ivanov
金额：
$ 37.59万
依托单位：
BRIGHAM AND WOMEN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-02-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10557098
关键词：
3-Dimensional 7-deazaguanine Affect Base Pairing Binding Binding Proteins Biochemical Biological Biological Assay Biological Process Biological Testing Biology Biophysics Cancer Biology Cell physiology Cells Cellular Stress Collaborations DNA DNA Maintenance Data Development Disease Elements Etiology Eukaryotic Cell Event Functional disorder G-Quartets Gene Expression Genes Genetic Transcription Goals Growth Guanine Guanosine Homeostasis Human In Vitro Intervention Investigation Ions Laboratories Malignant Neoplasms Mediating Messenger RNA Metabolism Methods Molecular Molecular Target Mutation Nerve Degeneration Neurodegenerative Disorders Nucleic Acid Regulatory Sequences Nucleic Acids Pathologic Pathology Pathway interactions Physiological Process Proteins Proteomics Publishing RNA RNA Binding RNA Sequences RNA metabolism Regulation Regulatory Element Reporting Research Ribonucleic Acid Regulatory Sequences Role Stress Structure System Techniques Testing Therapeutic Transcript Translational Regulation Translations Work biological adaptation to stress biophysical techniques candidate selection genetic manipulation genome-wide analysis human disease in vivo interest mRNA Stability mRNA Translation novel novel strategies novel therapeutics permissiveness pharmacologic physiologic stressor posttranscriptional prevent telomere three dimensional structure transcriptome tumorigenic

项目摘要

Summary G-quadruplexes (G4s) are non-canonical secondary structures in nucleic acids that are formed by guanine-rich sequences. G4 structural and functional studies have largely focused on DNA G4s, and the number of biological functions assigned to these motifs has grown rapidly since the discovery of their involvement in telomere biology. RNA G4s (RG4s) are less studied, but interest is increasing due to their association with multiple processes. A comprehensive understanding of how RNA G4s contribute to cell physiology and pathophysiology is the long-term research goal of the applicant. Multiple reports clearly demonstrate that G4s are enriched in mRNA 5’- and 3’-untranslated regulatory regions. There is an increasing evidence that RG4s control gene expression at transcriptional and post- transcriptional levels, although such data is largely based on in vitro studies. Testing the biological significance of RG4s requires proving that RG4s exist in vivo. Intriguingly, RG4s appear predominantly unfolded in eukaryotic cells, whereas they are readily folded in vitro, suggesting that in cells RG4s are constitutively recognized and actively unfolded. We hypothesize that RG4 folding-unfolding regulates mRNA homeostasis. This hypothesis is based on our analysis of human transcriptome that identifies RG4s as stress-responsive RNA elements. We will test this hypothesis with three specific aims. In AIM1, we will determine and characterize the fraction of the human transcriptome that contains putative stress-responsive RG4s in living cells. In AIM2, we will identify bona fide RG4-binding proteins using a novel approach based on proteomic/biochemical analysis of interactions between the 7-deazaguanine RNA derivatives and proposed binding factors. In AIM 3, we will use functional assays to determine the biological significance of RG4s in mRNA stability, localization and translation. We will use biophysical and biochemical methods to validate selected RG4 candidates. This work will elucidate how RG4-mediated functions contribute to cellular mRNA homeostasis, and will identify physiologically significant RG4-binding partners, which in turn may reveal molecular targets and pathways with therapeutic potential. The understanding of cellular functions of RG4 motifs is particularly relevant to the biology of cancer and neurodegeneration. RNA regions containing RG4s significantly overlap with regions containing disease- associated mutations. The proposed studies may elucidate molecular events underlying normal and pathological aspects of cell physiology, and identify events contributing to the tumorigenic or neurodegenerative changes.

概括 G-四链体 (G4s) 是核酸中的非规范二级结构，由 G4 结构和功能研究主要集中于 DNA G4 和富含鸟嘌呤的序列。自从发现这些基序以来，分配给这些基序的生物学功能数量迅速增加 RNA G4 (RG4) 参与端粒生物学的研究较少，但由于它们的作用而引起的兴趣正在增加。全面了解 RNA G4 如何对细胞做出贡献。生理学和病理生理学是申请人的长期研究目标。多个报告清楚地表明 G4 富含 mRNA 5'-和 3'-非翻译调节因子越来越多的证据表明 RG4 在转录和转录后控制基因表达。转录水平，尽管这些数据主要基于体外研究测试生物学意义。 RG4 的研究需要证明 RG4 存在于体内，有趣的是，RG4 在体内主要呈未折叠状态。真核细胞，而它们在体外很容易折叠，这表明在细胞中 RG4 是组成型的我们认识到 RG4 折叠-解折叠调节 mRNA 稳态。这一假设基于我们对人类转录组的分析，该分析将 RG4 识别为应激反应性 RNA 元素。我们将在 AIM1 中通过三个具体目标来检验这一假设。描述活体中包含推定应激反应 RG4 的人类转录组部分在 AIM2 中，我们将使用基于的新方法来鉴定真正的 RG4 结合蛋白。 7-脱氮鸟嘌呤 RNA 衍生物之间相互作用的蛋白质组学/生化分析，并提出在 AIM 3 中，我们将使用功能测定来确定 RG4 的生物学意义。我们将使用生物物理和生化方法来验证mRNA的稳定性、定位和翻译。选定的 RG4 候选者这项工作将阐明 RG4 介导的功能如何影响细胞 mRNA。稳态，并将识别生理上重要的 RG4 结合伙伴，这反过来可能揭示具有治疗潜力的分子靶点和途径。对 RG4 基序细胞功能的理解与癌症生物学特别相关含有 RG4 的 RNA 区域与含有疾病的区域显着重叠。相关的突变研究可能会阐明正常和潜在的分子事件。细胞生理学的病理学方面，并确定导致致瘤或致瘤的事件神经退行性改变。