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：
9901557
负责人：
MICHAEL S BOYCE
金额：
$ 30.08万
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 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9901557
关键词：
14-3-3 Family Ablation Affinity Arrhythmia Binding Binding Proteins Biochemical Biological Biological Models Biological Process Biophysics Cell Cycle Progression Cell Death Cell Extracts Cell Proliferation Cell Survival Cell physiology Cells Cellular Metabolic Process Collaborations Complex Crystallization Cytoplasmic Protein Deubiquitination Diabetes Mellitus Disease Enzymes Epitopes Family member Future Genetic Glycopeptides Glycoproteins Goals Human In Vitro Ligands Light Link Malignant Neoplasms Mammalian Cell Mammals Mediating Methylation Mitochondrial Proteins Modeling Multiprotein Complexes Mus Mutation Nuclear Envelope Nuclear Proteins Peptides Phosphoproteins Phosphorylation Physiological Physiology Play Post-Translational Protein Processing Process Property Protein Family Proteins Reader Role Signal Transduction Structure Tertiary Protein Structure Work cell growth chromatin remodeling design experimental study glycosylation human disease inorganic phosphate insight novel novel therapeutics paralogous gene protein complex protein function protein protein interaction public health relevance sugar

项目摘要

DESCRIPTION (provided by applicant): O-linked β-N-acetylglucosamine (O-GlcNAc) is a ubiquitous post-translational modification in mammals, decorating thousands of nuclear, cytoplasmic and mitochondrial proteins. O-GlcNAc cycling is an essential regulator of cell metabolism, cell cycle progression and cell death, and is dysregulated in numerous human diseases, such as cancer, diabetes and cardiac arrhythmia. Despite its broad pathophysiological significance, major aspects of O-GlcNAc signaling remain obscure, including how O-GlcNAc transduces biological information inside the cell. Recently, several studies showed that O-GlcNAcylation induces protein-protein interactions in processes as diverse as chromatin remodeling, deubiquitination and nuclear envelope assembly, suggesting that O-GlcNAc may signal through conserved modes of protein-protein interaction. However, little is known about either the structure or function of these intracellular glycoprotein-protein complexes. We hypothesized that mammalian "reader" proteins might exist and recognize O-GlcNAc moieties for signaling purposes. By analogy to reader proteins for other post-translational modifications, we reasoned that dedicated O-GlcNAc readers - or protein domains common to many readers - might relay glycosylation-encoded signals. In preliminary studies, we used a new biochemical approach to identify several proteins that bind specifically and directly to O-GlcNAcylated (but not unglycosylated) peptides and proteins in vitro and in cells. Interestingly, one group of putative O-GlcNAc reader proteins also binds to phosphoproteins, suggesting that they may be signal integrators, mediating the previously described crosstalk between O-GlcNAc and O- phosphate. This result may have wide-ranging implications for our understanding of cell signaling through post- translational modifications. The objective of this project is to characterize the biochemical and cell biological functions of the O- GlcNAc reader proteins that we identified. We will accomplish this goal through three specific aims. In Aim 1, we will define the biochemical scope of O-GlcNAc binding by the candidate readers. In Aim 2, we will identify the endogenous glycoprotein binding partners of O-GlcNAc reader proteins, and establish the role of these interactions in cell signaling. In Aim 3, we will determine the biophysical basis of an O-GlcNAc-mediated protein- protein interaction by solving the crystal structure of a reader protein bound to a glycosylated ligand. Our work will significantly advance the field of cell signaling by characterizing O-GlcNAc reader proteins at the cell biological, biochemical and atomic levels.

描述（由应用提供）：O连锁的β-N-乙酰葡萄糖胺（O-GLCNAC）是哺乳动物中普遍存在的翻译后修饰，装饰了数千种核，细胞质和线粒体蛋白。 O-GLCNAC自行车是细胞代谢，细胞周期进展和细胞死亡的重要调节剂，并且在许多人类疾病（例如癌症，糖尿病和心脏心律失常）中失调。尽管具有广泛的病理生理意义，但O-GLCNAC信号传导的主要方面仍然晦涩难懂，包括O-GLCNAC如何转化细胞内的生物学信息。最近，几项研究表明，O-Glcnacylation在潜水员（如染色质重塑，去泛素化和核包膜组装）的过程中诱导蛋白质 - 蛋白质相互作用，这表明O-GLCNAC可能通过蛋白质蛋白质相互作用的保守模式发出信号。然而，对于这些细胞内糖蛋白 - 蛋白质复合物的结构或功能知之甚少。我们假设哺乳动物的“读取器”蛋白可能存在，并识别O-GlCNAC部分以发出信号的目的。通过类似于其他翻译后修饰的读取器蛋白，我们认为专用的O-GLCNAC读取器 - 或许多读者常见的蛋白质结构域可能会中继糖基化编码的信号。在初步研究中，我们使用了一种新的生化方法来鉴定几种蛋白质，这些蛋白质在体外和细胞中直接与O-glcnacylated（但不是未糖基化的）辣椒和蛋白质结合。有趣的是，一组推定的O-GLCNAC读取子蛋白也与磷蛋白结合，表明它们可能是信号积分剂，介导了O-GLCNAC和O-磷酸盐之间先前描述的串扰。该结果可能对我们通过翻译后修改对细胞信号的理解具有广泛的影响。该项目的目的是表征我们确定的O-GLCNAC读取器蛋白的生化和细胞生物学功能。我们将通过三个特定目标来实现这一目标。在AIM 1中，我们将定义候选读者O-GLCNAC结合的生化范围。在AIM 2中，我们将确定O-GLCNAC读取器蛋白的内源性糖蛋白结合伴侣，并确定这些相互作用在细胞信号传导中的作用。在AIM 3中，我们将通过求解与糖基化配体结合的读取器蛋白的晶体结构来确定O-GLCNAC介导的蛋白质相互作用的生物物理基础。我们的工作将通过表征O-GLCNAC读取器蛋白在细胞生物学，生化和原子水平上来显着推动细胞信号传导领域。