Nutrient regulation of Alternative splicing and transcription by O-GlcNAcylation

O-GlcNAc 化对选择性剪接和转录的营养调节

• 批准号: 10725030
• 负责人: Shouling Xu
• 金额: $ 7.01万
• 依托单位: CARNEGIE INSTITUTION OF WASHINGTON, D.C.
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-01 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10725030
• 关键词: Acinus organ component; Affinity Chromatography; Agriculture; Alternative Splicing; Amino Acids; Animals; Apoptosis; Arabidopsis; Arabidopsis Proteins; Biochemical; Biological Models; Biology; Cell Differentiation process; Chromatin; Complement; Complex; Cues; Data; Development; Dissection; Ensure; Eukaryota; Eukaryotic Cell; Event; Flowers; Fractionation; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genomic approach; Genomics; Germination; Goals; Health; Homologous Gene; Human; Introns; Label; Link; Maps; Mass Spectrum Analysis; Modification; Molecular; Mutagenesis; Names; Nutrient; O-GlcNAc transferase; Pathway interactions; Plant Gene Expression Regulation; Plants; Play; Post-Translational Protein Processing; Productivity; Proteins; Proteomics; RNA; RNA Splicing; RNA-Protein Interaction; Regulation; Regulatory Pathway; Research; Role; Signal Transduction; Site; Specificity; Stress; System; Technology; Testing; Transcriptional Regulation; Transgenic Organisms; Transgenic Plants; biological adaptation to stress; cell growth regulation; crosslink; crosslinking and immunoprecipitation sequencing; experimental study; gene repression; genetic approach; hormonal signals; in vivo; mutant; novel strategies; plant growth/development; posttranscriptional; programs; protein complex; response; sugar; synthetic protein; transcriptome sequencing

The long-term goal of this project is to understand the molecular mechanisms that control gene expression and developmental transitions. While transcription has been extensively studied, the posttranscriptional mechanisms of RNA alternative splicing is much less understood despite of their importance in cellular regulation, human health, and plant growth and development. We have discovered that the Arabidopsis protein AtAcinus is evolutionarily related to but highly divergent from the human Acinus protein, which plays important roles in regulating transcription, RNA alternative splicing, and apoptosis. Our unpublished studies have shown that AtAcinus is modified by O- GlcNAcylation, plays essential role in alternative splicing of a number of genes, many of which encoding key components of signaling and developmental pathways. In particular, our data indicate that AtAcinus play important roles in regulating seed germination and flowering, two major developmental transition in plants. Using a combination of proteomics, genetics, genomic and biochemical approaches in the Arabidopsis model system, we have made tremendous progress in understanding the functions of AtAcinus. Our results support a hypothesis that AtAcinus is controlled by O-GlcNAcylation in response to endogenous and environmental cues, and in turn it regulates key cellular pathways through both transcriptional and posttranscriptional mechanisms. In this proposal, we plan to continue using the combination of proteomic, genomic and genetic approaches to further advance our understanding of Acinus regulatory pathway. We will 1) dissect the molecular functions of AtAcinus, particularly taking advantage of proximity labeling, cross-linking mass spectrometry and biochemical fractionation, CLIP-seq and CLIP-MS technologies to understand how AtAcinus carries out multiple functions (aim 1 and 3); 2) dissect how AtAcinus functions are regulated by post- translational modifications (aim 2). The experiments outlined in this proposal will greatly advance our understanding of the molecular mechanism of RNA alternatively splicing and O-GlcNAcylation and the mechanisms of signal integration at post-transcriptional level. Given the evolutionary conservation of Acinus, this study not only is important for plant biology and agriculture, but also can potentially help us understand fundamental mechanisms of signaling and cellular regulation that are relevant broadly.

该项目的长期目标是了解控制基因的分子机制 表达和发育转变。虽然转录已被广泛研究， 尽管 RNA 选择性剪接的转录后机制有其自身的特点，但人们对它的了解却少之又少。 在细胞调节、人类健康以及植物生长和发育中具有重要意义。我们有 发现拟南芥蛋白 AtAcinus 在进化上与它相关但高度不同 人类腺泡蛋白，在调节转录、RNA替代中发挥重要作用 剪接和细胞凋亡。我们未发表的研究表明 AtAcinus 被 O- 修饰 GlcNAc 酰化在许多基因的选择性剪接中起着重要作用，其中许多基因 编码信号传导和发育途径的关键组成部分。特别是，我们的数据表明 AtAcinus 在调节种子发芽和开花这两个主要方面发挥着重要作用 植物的发育转变。结合蛋白质组学、遗传学、基因组学和 我们在拟南芥模型系统的生化方法方面取得了巨大进展 了解 AtAcinus 的功能。我们的结果支持 AtAcinus 受到控制的假设 通过 O-GlcNAcylation 响应内源性和环境信号，进而调节关键 通过转录和转录后机制的细胞途径。在这个提案中， 我们计划继续结合蛋白质组学、基因组学和遗传学方法来进一步研究 增进我们对腺泡调节途径的理解。我们将1）剖析分子功能 AtAcinus，特别是利用邻近标记、交联质谱和 生化分级分离、CLIP-seq 和 CLIP-MS 技术可了解 AtAcinus 如何携带 多种功能（目标 1 和 3）； 2）剖析AtAcinus功能是如何被后调节的 翻译修饰（目标 2）。本提案中概述的实验将极大地推进 我们对RNA选择性剪接和O-GlcNAcylation分子机制的理解 以及转录后水平的信号整合机制。鉴于进化论 Acinus 的保护，这项研究不仅对植物生物学和农业具有重要意义，而且可以 可能有助于我们理解信号传导和细胞调节的基本机制 广泛相关。