Nutrient regulation of Alternative splicing and transcription by O-GlcNAcylation

O-GlcNAcylation 对选择性剪接和转录的营养调节

• 批准号: 10063997
• 负责人: Shouling Xu
• 金额: $ 33.92万
• 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/10063997
• 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; 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; Seeds; Signal Transduction; Site; Specificity; Stress; System; Technology; Testing; Transcriptional Regulation; Transgenic Organisms; Transgenic Plants; base; 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; 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 Our unpublished studies have shown that AtAcinus is modified by O- GlcNAcylation, plays essential role in alternative splicing of a number of genes, many of which encoding key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表达的分子机制。 表达和发育转变。虽然转录已被广泛研究， RNA选择性剪接的转录后机制是了解少得多，尽管他们 在细胞调节、人类健康和植物生长发育中的重要性。我们有 发现拟南芥AtAcinus蛋白质在进化上与拟南芥AtAcinus蛋白质相关，但与拟南芥AtAcinus蛋白质高度不同。 人腺泡蛋白在转录调控中起重要作用，RNA替代物 剪接和凋亡。我们未发表的研究表明，AtAcinus被O- GlcNAc酰化在许多基因的选择性剪接中起重要作用，其中许多基因 编码信号传导和发育途径的关键成分。特别是，我们的数据表明， AtAcinus在调节种子萌发和开花中起重要作用，这两个主要的 植物的发育转变利用蛋白质组学、遗传学、基因组学和 生化方法在拟南芥模型系统中，我们已经取得了巨大的进展， 了解AtAcinus的功能。我们的研究结果支持了一个假设，即AtAcinus是受控制的 通过O-GlcNAcylation响应内源性和环境线索，反过来它调节关键的 通过转录和转录后机制的细胞途径。在这一提议中， 我们计划继续使用蛋白质组学、基因组学和遗传学方法的组合， 促进我们对腺泡调节通路的理解。我们将1）剖析分子功能 的AtAcinus，特别是利用邻近标记，交联质谱和 生物化学分馏，CLIP-seq和CLIP-MS技术，以了解AtAcinus如何携带 多功能（目标1和3）; 2）解剖AtAcinus功能如何受后 翻译修饰（目的2）。该提案中概述的实验将大大推进 我们对RNA选择性剪接和O-GlcNAc化的分子机制的理解 以及转录后水平的信号整合机制。考虑到进化论 本研究不仅对植物生物学和农业有重要意义， 可能帮助我们理解信号和细胞调节的基本机制， 广泛相关。