Chemical Reporters for the Analysis of Lysine Methylation in Epigenetics

用于表观遗传学中赖氨酸甲基化分析的化学报告基因

• 批准号: 7571455
• 负责人: Howard C Hang
• 金额: $ 33.71万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-15 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7571455
• 关键词: Address; Biological; Biological Process; Chemicals; Chromatin; Chromosomes; Communities; Complex Mixtures; DNA Sequence; Development; Disease; Environment; Enzymes; Epigenetic Process; Family; Gene Expression; Generations; Genetic; Genetic Transcription; Health; Heart; Heterochromatin; Histone-Lysine N-Methyltransferase; Histones; Human; In Vitro; Individual; Label; Lysine; Methods; Methylation; Methyltransferase; Nucleosomes; Pathway interactions; Phenotype; Physiology; Play; Post-Translational Protein Processing; Proteins; Public Health; Reporter; Role; Signal Transduction; Specificity; Structure; Tail; Transcriptional Regulation; analog; cationite KMT; enzyme substrate; enzyme substrate analog; mutant; novel; rapid detection; tool; transmission process

DESCRIPTION (provided by applicant): Chemical Reporters for the Analysis of Lysine Methylation in Epigenetics The transmission of biological information over generations is regulated beyond the level of the DNA sequence. This fundamental principle has been observed in many biological contexts and has been termed "epigenetics", which begins to describe the basic contribution of environment to phenotype under a given genetic background [1]. At the heart of epigenetic mechanisms is regulated gene expression, which manifests itself in the fine control of nucleosome structure and function, the core unit of chromosomes. One of the key factors that modulate transcription is the posttranslational modification (PTM) of chromatin-associated proteins (histones) [2]. In particular, dynamic lysine methylation of histone tails appears to play essential roles in regulating gene expression and epigenetic phenomena [3, 4]. The families of enzymes that regulate lysine methylation on histones, lysine methyltransferases (KMTs) and lysine demethylases (KDMs), have now been identified and are associated with transcriptional regulation, x-chromosomal inactivation and heterochromatin formation [3, 4]. Interestingly, lysine methylation on several non-histone proteins has also been recently described, which has raised many questions regarding the specificity of KMTs and KDMs as well as their discrete roles in epigenetics [2-5]. Unfortunately, the lack of general methods to characterize lysine methylation on proteins and their respective enzymes has hindered a more general understanding of how this PTM regulates signal transduction and epigenetic mechanisms. To fully appreciate the roles of lysine methylation in biological pathways, new methods with higher sensitivity and generality are required. We therefore propose to develop chemical reporters for lysine methylation that will enable rapid detection and identification of methylated proteins in complex mixtures using bioorthogonal labeling methods (Aim 1). Furthermore, we will develop orthogonal enzyme-substrate pairs to identify selective protein substrates of individual lysine methyltransferases (Aim 2). These studies should uncover novel lysine-methylated proteins and identify specific enzymes that regulate their function in fundamental cellular pathways and epigenetic mechanisms of phenotypic inheritance. Ultimately, these tools should provide a more general understanding of protein methylation in normal physiology and disease. PUBLIC HEALTH RELEVANCE: The modulation of phenotypes by epigenetic mechanisms is central to many biological processes and diseases. A detail understanding of the underlying mechanisms that control epigenetics is therefore essential to human health. Reversible lysine methylation on proteins has emerged as an important PTM that regulates the inheritance of phenotypes, however, the analysis of lysine methylation requires more general methods to characterize specific substrates for KMTs and KDMs. To address this problem, this proposal describes the development of chemical reporters to identify lysine-methylated proteins and specific substrates of KMTs. If successful, these chemical approaches would provide the scientific community with a new set of tools to analyze the role of protein methylation in fundamental cellular pathways and in epigenetic processes.

描述（由申请人提供）：表观遗传学中赖氨酸甲基化分析的化学报告基因生物信息在几代人中的传递受到DNA序列水平以外的调控。这一基本原则已在许多生物学背景中观察到，并被称为“表观遗传学”，它开始描述在给定遗传背景下环境对表型的基本贡献[1]。表观遗传机制的核心是调控基因表达，这表现在对核小体结构和功能的精细控制中，核小体是染色体的核心单位。调节转录的关键因素之一是染色质相关蛋白（组蛋白）的翻译后修饰（PTM）[2]。特别是，组蛋白尾部的动态赖氨酸甲基化似乎在调节基因表达和表观遗传现象中发挥重要作用[3，4]。调节组蛋白上赖氨酸甲基化的酶家族，赖氨酸甲基转移酶（KMT）和赖氨酸脱甲基酶（KDM），现已被鉴定并且与转录调节、x染色体失活和异染色质形成相关[3，4]。有趣的是，最近还描述了几种非组蛋白蛋白上的赖氨酸甲基化，这引起了许多关于KMT和KDM的特异性以及它们在表观遗传学中的离散作用的问题[2-5]。不幸的是，缺乏通用的方法来表征赖氨酸甲基化的蛋白质和它们各自的酶，阻碍了更全面的了解这种PTM如何调节信号转导和表观遗传机制。为了充分认识赖氨酸甲基化在生物学途径中的作用，需要具有更高灵敏度和通用性的新方法。因此，我们建议开发化学报告赖氨酸甲基化，这将使快速检测和识别的甲基化蛋白质在复杂的混合物中使用生物正交标记方法（目的1）。此外，我们将开发正交酶底物对，以确定个别赖氨酸甲基转移酶（目标2）的选择性蛋白质底物。这些研究应该发现新的赖氨酸甲基化蛋白，并确定特定的酶，调节其功能的基本细胞途径和表型遗传机制。最终，这些工具应该提供对正常生理和疾病中蛋白质甲基化的更全面的理解。 公共卫生相关性：表观遗传机制对表型的调节是许多生物过程和疾病的核心。因此，详细了解控制表观遗传学的潜在机制对人类健康至关重要。蛋白质上的可逆赖氨酸甲基化已成为调节表型遗传的重要PTM，然而，赖氨酸甲基化的分析需要更通用的方法来表征KMT和KDM的特异性底物。为了解决这个问题，该提案描述了化学报告的发展，以确定赖氨酸甲基化的蛋白质和特定底物的KMT。如果成功的话，这些化学方法将为科学界提供一套新的工具来分析蛋白质甲基化在基本细胞途径和表观遗传过程中的作用。