Role of O-GlcNAc in human embryonic stem cell pluripotency and differentiation

O-GlcNAc 在人胚胎干细胞多能性和分化中的作用

• 批准号: 8214102
• 负责人: Lissette Andres
• 金额: $ 3.38万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-02-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8214102
• 关键词: Acetylglucosamine; Adult; Alzheimer' s Disease; Area; Binding; Biological Assay; Cell Survival; Cell physiology; Chemicals; Chromatin; Cytoplasmic Protein; DNA; Development; Disease; Drosophila genus; Engineering; Foundations; Gene Proteins; Gene Silencing; Genomics; Glycoproteins; Goals; Human; Lead; Link; Maintenance; Mediating; Metabolic; Modification; Molecular; Monosaccharides; Mutation; Neuronal Differentiation; Neurons; Nuclear Proteins; Oligosaccharides; Parkinson Disease; Play; Polycomb; Post-Translational Protein Processing; Process; Protein Array; Protein Binding; Proteins; Proteomics; Regenerative Medicine; Reporter; Repression; Response Elements; Role; Spielmeyer-Vogt Disease; Staging; Stem cells; Therapeutic Human Experimentation; Transcript; Transcription Repressor/Corepressor; Transcriptional Regulation; Work; adult stem cell; base; cell type; chromatin immunoprecipitation; human disease; human embryonic stem cell; neuron development; pluripotency; public health relevance; self-renewal; stem cell biology; stem cell fate; tool; transcription factor

DESCRIPTION (provided by applicant): The objective of this project is to characterize the functional role of protein O- GlcNAcylation in crucial areas of stem cell biology. O-GlcNAcylation is a post- translational modification in which a single monosaccharide (O-GlcNAc) is installed on Ser or Thr residues of cytosolic and nuclear proteins. This modification has been found on a wide array of proteins, including transcription factors. Although much is known about the crucial role of O-GlcNAcylation in cellular processes, this modification has yet to be studied in the context of regenerative medicine. I propose herein to characterize the functional role of O- GlcNAcylation in two critical areas of stem cell biology: neuronal differentiation (Aim 1), and the repression of genomic transcripts by polycomb group proteins (Aim 2). In Aim 1, I propose to use Metabolic Oligosaccharide Engineering (MOE) and glycoproteomic analysis to identify the protein substrates of O-GlcNAcylation present during various stages of neuronal differentiation. These studies will aid in the elucidation of the molecular determinants of stem cell fate decisions during neuronal differentiation. In Aim 2 I will study the putative role of O-GlcNAcylation in the maintenance of hESC and adult stem cells by polycomb group proteins (PcGs). Recent work in Drosophila has shown that O-GlcNAcylation governs PcG-mediated gene silencing during development. Herein we will identify both O-GlcNAcylated proteins and the genes that are being modulated by them by performing a chromatin immunoprecipitation (ChIP)-type assay facilitated by MOE. This work may expand the current understanding of the molecular mechanisms underlying self-renewal and pluripotency of hESCs. In summary, the successful completion of the studies proposed herein will constitute the first-ever comprehensive study of protein O-GlcNAcylation in hESCs, and will greatly expand the current understanding of molecular mechanisms governing neuronal differentiation, pluripotency and transcriptional control in hESC biology. PUBLIC HEALTH RELEVANCE: Human embryonic stem cells can be changed into virtually any cell type in the adult body, and thus, have the potential to cure a vast majority of existing human disorders. The results of the studies proposed in this project may lead to a greatly increased understanding of how stem cells retain their ability to be changed into other cell types, and also how the fate of stem cells is decided upon differentiation. Both are critical areas that need to be explored to enable modern regenerative medicine to realize its full potential as tool for the treatment of human diseases, such as Alzheimer's, Parkinson, and Batten disease.

描述（由申请人提供）：该项目的目的是表征蛋白质 O-GlcNAc 酰化在干细胞生物学关键领域的功能作用。 O-GlcNAc 酰化是一种翻译后修饰，其中将单个单糖 (O-GlcNAc) 安装在胞浆蛋白和核蛋白的 Ser 或 Thr 残基上。这种修饰已在多种蛋白质中发现，包括转录因子。尽管人们对 O-GlcNAc 酰化在细胞过程中的关键作用了解很多，但这种修饰尚未在再生医学的背景下进行研究。我在此建议描述 O-GlcNAc 酰化在干细胞生物学的两个关键领域中的功能作用：神经元分化（目标 1）和多梳蛋白对基因组转录物的抑制（目标 2）。在目标 1 中，我建议使用代谢寡糖工程 (MOE) 和糖蛋白组学分析来鉴定神经元分化各个阶段中存在的 O-GlcNAcNA 酰化的蛋白质底物。这些研究将有助于阐明神经元分化过程中干细胞命运决定的分子决定因素。在目标 2 中，我将研究 O-GlcNAcylation 在多梳组蛋白 (PcG) 维持 hESC 和成体干细胞中的假定作用。最近在果蝇中的研究表明，O-GlcNAc 酰化控制着发育过程中 PcG 介导的基因沉默。在此，我们将通过执行由 MOE 促进的染色质免疫沉淀 (ChIP) 型测定来鉴定 O-GlcNAcNA 酰化蛋白和受其调节的基因。这项工作可能会扩大目前对 hESC 自我更新和多能性分子机制的理解。总之，本文提出的研究的成功完成将构成对 hESC 中蛋白质 O-GlcNAc 酰化的首次全面研究，并将极大地扩展目前对 hESC 生物学中控制神经元分化、多能性和转录控制的分子机制的理解。 公共健康相关性：人类胚胎干细胞几乎可以转变为成人体内的任何细胞类型，因此有可能治愈绝大多数现有的人类疾病。该项目提出的研究结果可能会大大加深人们对干细胞如何保持其转变为其他细胞类型的能力，以及干细胞的命运如何在分化时决定的理解。这两个领域都是需要探索的关键领域，以使现代再生医学能够充分发挥其作为治疗阿尔茨海默病、帕金森病和巴顿病等人类疾病工具的潜力。