Cell signaling through O-GlcNAc reader proteins

通过 O-GlcNAc 读取蛋白的细胞信号传导

基本信息

批准号：
10656649
负责人：
MICHAEL S BOYCE
金额：
$ 32.04万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-04-01 至 2027-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10656649
关键词：
14-3-3 Family Ablation Binding Binding Proteins Biochemical Biological Biological Process Biophysics Cell model Cell physiology Cells Comparative Study Complex Cytoplasmic Protein Cytoskeletal Filaments Cytoskeletal Proteins Deubiquitination Diabetes Mellitus Diffusion Disease Enzymes Future Genetic Glycoproteins Goals Homo Human Importins In Vitro Individual Intermediate Filament Proteins Intermediate Filaments Link Malignant Neoplasms Mammalian Cell Mammals Mediating Methods Modeling Multiprotein Complexes Mus Nerve Degeneration Nuclear Nuclear Envelope Nuclear Import Nuclear Pore Nuclear Pore Complex Proteins Nuclear Proteins Organelles Peptides Phenotype Phosphoproteins Phosphorylation Physiological Polysaccharides Post-Translational Protein Processing Process Proteins Reader Regulation Reporting Resolution Role Signal Transduction Structure Testing Vimentin Work X-linked intellectual disability chromatin remodeling comparative glycosylation human disease inorganic phosphate insight neurofilament novel novel therapeutics nucleocytoplasmic transport paralogous gene protein complex protein function protein protein interaction receptor receptor binding sugar trafficking transmission process ubiquitin-protein ligase

项目摘要

Project Summary/Abstract O-linked b-N-acetylglucosamine (O-GlcNAc) is a ubiquitous post-translational modification (PTM) in mammals, decorating thousands of nuclear and cytoplasmic proteins. O-GlcNAc cycling is an essential regulator of myriad aspects of cell physiology and is dysregulated in numerous human diseases, such as cancer, diabetes and neurodegeneration. Despite this broad pathophysiological significance, major aspects of O-GlcNAc signaling remain incompletely understood, including the biochemical mechanisms through which O-GlcNAc transduces information. At the start of our project, several studies had shown that O-GlcNAcylation mediates protein-protein interactions (PPIs) in processes as diverse as chromatin remodeling, deubiquitination and nuclear envelope assembly. These results suggested that O-GlcNAc might signal through conserved modes of PPI. However, little was known about either the structure or function of these intracellular glycoprotein-protein complexes. We hypothesized that mammalian “reader” proteins might exist and transduce O-GlcNAc signals. In the first project period, we devised and deployed a method to identify proteins that bind specifically and directly to O- GlcNAcylated peptides and proteins in vitro and in human cells. These include the 14-3-3 family, which are well- known to bind to phosphoproteins. Our results indicate that 14-3-3 proteins may be signal integrators, mediating the extensively documented crosstalk between O-GlcNAc and O-phosphate, with broad implications for intracellular signaling. In addition, we identified and characterized O-GlcNAc-mediated PPIs among nuclear import receptors, intermediate filament (IF) cytoskeleton proteins and ubiquitin E3 ligase complex components. Building on these results, we will study a focused set of diverse model O-GlcNAc-binding proteins in the next project period. This strategy will generate new insight into the functions of individual O-GlcNAc-mediated PPIs in particular cell biological processes, while also providing a comparative perspective across reader proteins, allowing us to discern general themes of O-GlcNAc-mediated PPIs at the biophysical, biochemical and physiological levels. The long-term objective of our work is to understand the principles, mechanisms and functions of O- GlcNAc-mediated PPIs in mammals. In the next project period, we will advance this goal through three Specific Aims. In Aim 1, we will define the biochemical scope and phenotypic effects of O-GlcNAc binding by 14-3-3 proteins. In Aim 2, we will determine the role of O-GlcNAc/nuclear transport receptor interactions in inter-organelle trafficking. In Aim 3, we will dissect the functional importance of O-GlcNAc-mediated interactions in two model IF proteins. We believe our proposed work will significantly advance the field of intracellular signaling both by elucidating the mechanisms and functions of individual O-GlcNAc-mediated PPIs and by revealing conserved biochemical principles of O-GlcNAc recognition across reader proteins.

项目摘要/摘要 O连锁的B-N-乙酰葡萄糖（O-GLCNAC）是无处不在的翻译后修饰（PTM）哺乳动物，装饰数千种核和细胞质蛋白。 O-GLCNAC自行车是必不可少的调节器细胞生理学的各个方面，并且在许多人类疾病（例如癌症）中失调和神经变性。尽管具有广泛的病理生理意义，但O-GLCNAC信号的主要方面保持不完全理解，包括O-GLCNAC传递的生化机制信息。在我们项目开始时，几项研究表明O-Glcnacylation介导蛋白质蛋白质像染色质重塑，去泛素化和核包膜一样潜水的过程中的相互作用（PPI）集会。这些结果表明O-GLCNAC可能通过配置的PPI模式发出信号。但是，很少已经知道这些细胞内糖蛋白 - 蛋白质复合物的结构或功能。我们假设哺乳动物的“读取器”蛋白可能存在并传递O-GLCNAC信号。在第一个项目时期，我们设计并部署了一种方法，以识别特异性结合并直接与O-的蛋白质在体外和人类细胞中，Glcnacylated Pepperides和蛋白质。其中包括14-3-3家族已知与磷蛋白结合。我们的结果表明14-3-3蛋白可能是信号积分器，介导 O-GLCNAC和O-磷酸盐之间的广泛记录的串扰，对细胞内信号传导。此外，我们在核进口中确定并表征了O-GLCNAC介导的PPI 受体，中间细丝（IF）细胞骨架蛋白和泛素E3连接酶复合成分。建筑在这些结果上，我们将在下一个项目时期研究一组重点的潜水员模型O-GLCNAC结合蛋白。该策略将对单个O-GLCNAC介导的PPI的功能产生新的见解生物过程，同时还提供读者蛋白质的比较观点，使我们能够辨别 O-GlCNAC介导的PPI的一般主题在生物物理，生化和物理水平上。我们工作的长期目标是了解O-的原理，机制和功能 GlcNAC介导的哺乳动物中的PPI。在下一个项目时期，我们将通过三个特定的特定目标提高这一目标目标。在AIM 1中，我们将定义O-GLCNAC结合的生化范围和表型效应14-3-3 蛋白质。在AIM 2中，我们将确定O-GLCNAC/核运输接收器相互作用在轨道间的作用贩运。在AIM 3中，如果在两个模型中，我们将阐述O-GLCNAC介导的相互作用的功能重要性蛋白质。我们认为，我们提出的工作将大大推动细胞内信号传导的领域阐明单个O-GlCNAC介导的PPI的机制和功能，并通过揭示构成读取器蛋白质O-GLCNAC识别的生化原理。