Structural and Functional Analysis of Nucleocytoplasmic Protein O-Glycosyltransferases in Plants

植物核胞质蛋白 O-糖基转移酶的结构和功能分析

• 批准号: 10648930
• 负责人: Tai-Ping Sun
• 金额: $ 41.92万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10648930
• 关键词: Affect; Agriculture; Allosteric Regulation; Anabolism; Animals; Arabidopsis; Bacteria; Binding; Biochemical; Biological Assay; Biology; Cell physiology; Cells; Complex; Cryoelectron Microscopy; Defect; Diabetes Mellitus; Disease; Environment; Enzymes; Event; Family; Fucosyltransferase; Genes; Genetic; Genetic Transcription; Gibberellins; Growth; Guanosine Diphosphate Fucose; Health; Human; Individual; Insecta; Knowledge; Length; Life; Link; Malignant Neoplasms; Mediating; Metabolic; Modification; Molecular; Molecular Conformation; Mutation; N-terminal; Neurodegenerative Disorders; O-GlcNAc transferase; Organism; Phenotype; Phytochrome; Plant Growth Regulators; Plants; Play; Protein Glycosylation; Proteins; Proteome; Proteomics; Public Health; Regulation; Resistance; Role; Sequence Homology; Signal Pathway; Signal Transduction; Site-Directed Mutagenesis; Structure; Substrate Specificity; Transcription Factor 3; Transcription Repressor; Variant; Virulence; environmental change; gene conservation; genetic analysis; glycosylation; glycosyltransferase; human disease; insight; link protein; mutant; nanobodies; novel; paralogous gene; plant growth/development; public health relevance; response; sugar; tool; transcription factor; transcriptional reprogramming

Project Summary/Abstract O-linked protein glycosylation orchestrates diverse signaling events in plant development in response to the changing environment. We have discovered the first nucleocytoplasmic O-fucosyltransferase, SPINDLY (SPY) in Arabidopsis, which together with its paralog SECRET AGENT (SEC), an O-GlcNAc transferase (OGT), post- translationally modify the conserved DELLA family of transcription regulators, such as RGA, to control their ability to modulate multiple signaling pathways. SPY-catalyzed O-fucosylation of RGA enhances the RGA interaction with key transcription factors to suppress plant growth, whereas SEC-mediated O-GlcNAcylation weakens RGA activity and promotes plant development. Despite their functional significance, how these two homologous glycosyltransferases, SPY and SEC, have evolved distinct substrate specificities and how these two types of glycosylation, O-fucosylation by SPY and O-GlcNAcylation by SEC, regulate the RGA conformation to influence plant development at the molecular level have remained unknown. We propose to elucidate the molecular details of SPY, SEC, and their substrate and product complexes, as well as the distinct structural and functional consequences of RGA O-fucosylation and O-GlcNAcylation. Recent proteomic analysis has unveiled hundreds of SPY and SEC substrates beyond RGA, highlighting the broad implication of protein glycosylation in regulating cellular functions. In order to overcome the potential pleotropic effects brought by spy and sec mutations in planta and decipher the functional consequences of O-fucosylation and O-GlcNAcylation for specific target proteins, we will develop nanobody-fused SPY and SEC variants for precise target modification. Collectively, our study will fill the critical knowledge gap about the structural and functional consequences of protein glycosylation in plants. As SPY-like genes are conserved in diverse organisms, including plants, bacteria, and parasitic protists, and as SEC-like (OGT) genes are broadly distributed in plants and animals, a molecular understanding of these enzymes and their distinct structural and functional consequences will have a profound impact in plant biology, agriculture, and human health and diseases.

项目概要/摘要 O-连接蛋白糖基化协调植物发育中的多种信号事件，以响应 不断变化的环境。我们发现了第一个核质 O-岩藻糖基转移酶 SPINDLY (SPY) 在拟南芥中，它与它的旁系同源物 SECRET AGENT (SEC) 一起，是一种 O-GlcNAc 转移酶 (OGT)， 翻译修饰保守的 DELLA 转录调节因子家族，例如 RGA，以控制其能力 调节多种信号通路。 SPY 催化的 RGA O-岩藻糖基化增强了 RGA 相互作用 具有抑制植物生长的关键转录因子，而 SEC 介导的 O-GlcNAc 酰化会削弱 RGA 活性并促进植物发育。尽管它们具有重要的功能，但这两个同源物如何 糖基转移酶 SPY 和 SEC 已进化出不同的底物特异性，以及这两种类型如何 糖基化、SPY 的 O-岩藻糖基化和 SEC 的 O-GlcNA 酰化，调节 RGA 构象以影响 分子水平上的植物发育仍然未知。我们建议阐明分子细节 SPY、SEC 及其底物和产物复合物，以及独特的结构和功能 RGA O-岩藻糖基化和 O-GlcNAcylation 的后果。最近的蛋白质组学分析揭示了数百个 RGA 之外的 SPY 和 SEC 底物，强调了蛋白质糖基化在调节中的广泛含义 细胞功能。为了克服植物中spy和sec突变带来的潜在多效性效应 并破译 O-岩藻糖基化和 O-GlcNAcNA 酰化对特定靶蛋白的功能影响， 我们将开发纳米抗体融合的 SPY 和 SEC 变体，以实现精确的目标修饰。总的来说，我们的研究 将填补关于蛋白质糖基化的结构和功能后果的关键知识空白 植物。由于 SPY 样基因在多种生物体中保守，包括植物、细菌和寄生原生生物， 由于 SEC 样（OGT）基因广泛分布于植物和动物中，因此对这些基因的分子理解 酶及其独特的结构和功能后果将对植物生物学产生深远的影响， 农业、人类健康和疾病。