METTL3-NUP93 interaction facilitates the nuclear export of m6A-modified mRNAs

METTL3-NUP93 相互作用促进 m6A 修饰 mRNA 的核输出

基本信息

批准号：
10299046
负责人：
Kexin Xu
金额：
$ 33.31万
依托单位：
UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-10 至 2026-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10299046
关键词：
3&apos Untranslated Regions Adaptor Signaling Protein Adenosine Affect Apoptosis Articulation Binding Biological Biological Assay Biology CRISPR/Cas technology Cell Cycle Progression Cell Nucleus Cell Proliferation Cell physiology Cells Chronic Kidney Failure Complex Cytoplasm DNA Sequence Alteration Data Defect Development Enzymes Epigenetic Process Fibrinogen Fractionation Gene Expression Genes Hand Health Human Immunoprecipitation Kidney Kinetics Knowledge Label Ligation Link Mediating Messenger RNA Methylation Methyltransferase Microscopy Modification Molecular Monitor Movement Mutation Nitrogen Nuclear Nuclear Envelope Nuclear Export Nuclear Pore Complex Nuclear Pore Complex Proteins Nuclear Structure Ontology Pathogenesis Pathologic Pathway interactions Patients Pattern Polyribosomes Positioning Attribute Process Production Proteins Proteomics RNA RNA Binding RNA Transport Regulation Reporting Research Resolution Role Sampling Signal Transduction Site Speed Steroid-resistant idiopathic nephrotic syndrome System Technology Testing Transcript Untranslated Regions base biological systems clinically relevant clinically significant congenital anomaly density epitranscriptome epitranscriptomics human disease insight knock-down mutant new therapeutic target novel receptor recruit scaffold spatiotemporal stem theories tool transcriptome transcriptome sequencing

项目摘要

PROJECT ABSTRACT The nucleus-cytoplasm shuttling of messenger RNAs (mRNAs) is a key determinant in the spatiotemporal articulation of gene expression. This process is finely tuned by multiple factors, one of which is the addition of a methyl moiety onto the adenosine base at the nitrogen-6 position (m6A). Emerging evidence suggests that m6A enhances the nuclear export of decorated mRNAs. Current theories explaining m6A-assisted mRNA export focus on the crosstalk between the protein enzymes involved in m6A modification and the typical RNA- binding adaptor proteins that hand the mRNA cargoes over to the classical export receptors. In this study, we have found a noncanonical transport axis for m6A-marked mRNAs. Our preliminary data have shown that METTL3, an integral subunit of the m6A methyltransferase complex, directly interacts with NUP93, a component of the nuclear pore complex (NPC). RNA-binding capacity of METTL3 helps link the m6A-modified mRNAs to NUP93 on the nuclear envelope, which accelerates the passage of mRNA cargoes through the NPCs with the assistance of the principal export factor NXF1-NXT1. Integrative analysis of the m6A profile and fractionation transcriptome has identified a large group of m6A- harboring mRNAs whose nuclear export is dependent on the METTL3-NUP93 complex. Interestingly, these mRNAs are distinguished from other transcripts, as they tend to have longer 3' untranslated regions (3'-UTR), which is associated with more m6A sites and stronger methylation intensities. In addition, Gene Ontology analysis has revealed important functions of these transcripts in cellular physiology. In this regard, we found that the genetic mutations on NUP93, which have been reported to cause steroid-resistant nephrotic syndrome (SRNS) in humans, disrupts its interaction with METTL3, implying a potential association between dysregulation of METTL3-NUP93 axis and development of certain pathological conditions. Being guided by these preliminary data, we have developed biological systems to monitor the m6A status of test mRNA species and will employ cutting-edge technologies, such as single-point edge-excitation subdiffraction microscopy and epitranscriptome-editing tools, to define the role of METTL3-NUP93 axis in the nuclear export of m6A- conjugated mRNAs. Our Specific Aims are: (1). To examine how the METTL3-NUP93 interaction facilitates the nuclear export of m6A-marked mRNAs; (2). To determine which transcripts are specifically regulated by the METTL3-NUP93 axis; (3). To elucidate why this new mRNA transport pathway is biologically important. Our proposed research will impact upon the fields of RNA biology and epigenetics. Findings from this study will help delineate a novel mechanism of m6A-assisted mRNA export and offer new therapeutic targets for certain human diseases, such as kidney abnormalities that are caused by NUP93 mutations.

项目摘要信使RNA（mRNA）的核-细胞质穿梭是时空的关键决定因素基因表达的阐明。这个过程是通过多种因素进行微调的，其中之一是添加了甲基部分连接到腺苷碱基的氮 6 位 (m6A)。新出现的证据表明 m6A 增强修饰 mRNA 的核输出。当前解释 m6A 辅助 mRNA 的理论导出重点关注参与 m6A 修饰的蛋白酶与典型的 RNA- 之间的串扰结合接头蛋白将 mRNA 货物传递给经典的输出受体。在这项研究中，我们发现了 m6A 标记 mRNA 的非规范运输轴。我们的初步数据表明，METTL3（m6A 甲基转移酶复合物的一个完整亚基）直接与核孔复合物 (NPC) 的组成部分 NUP93 相互作用。 RNA 结合能力 METTL3 有助于将 m6A 修饰的 mRNA 与核膜上的 NUP93 连接起来，从而加速在主要输出因子 NXF1-NXT1 的帮助下，mRNA 货物通过 NPC。 m6A 谱和分级转录组的综合分析已鉴定出一大群 m6A- 含有核输出依赖于 METTL3-NUP93 复合物的 mRNA。有趣的是，这些 mRNA 与其他转录物不同，因为它们往往具有更长的 3' 非翻译区 (3'-UTR)，这与更多的 m6A 位点和更强的甲基化强度相关。此外，基因本体论分析揭示了这些转录本在细胞生理学中的重要功能。对此，我们发现 NUP93 的基因突变，据报道会导致类固醇抵抗性肾病综合征（SRNS）在人类中，破坏了它与 METTL3 的相互作用，这意味着之间的潜在关联 METTL3-NUP93 轴失调和某些病理状况的发展。受到指导根据这些初步数据，我们开发了生物系统来监测测试 mRNA 物种的 m6A 状态并将采用尖端技术，例如单点边缘激发亚衍射显微镜和表观转录组编辑工具，用于定义 METTL3-NUP93 轴在 m6A- 核输出中的作用缀合的 mRNA。我们的具体目标是：（1）。检查 METTL3-NUP93 交互如何促进 m6A 标记 mRNA 的核输出；（2）。确定哪些转录本受到具体监管 METTL3-NUP93轴；（3）。阐明为什么这种新的 mRNA 转运途径具有重要的生物学意义。我们提出的研究将影响 RNA 生物学和表观遗传学领域。由此得出的结论研究将有助于描绘 m6A 辅助 mRNA 输出的新机制并提供新的治疗靶点用于某些人类疾病，例如由 NUP93 突变引起的肾脏异常。