Investigating the mechanistic origins, maintenance and functions of gene body DNA methylation in plants

在大多数真核生物的基因组中，DNA中胞嘧啶碱基的甲基化是一种常见的修饰，长期以来被认为可以下调或关闭基因表达。这一观点受到了最近研究的挑战，研究表明甲基化对基因表达的影响取决于甲基化发生的位置。在包括人类和一些植物在内的许多生物中，位于基因旁边的DNA甲基化会使该基因关闭，而位于基因体内的DNA甲基化则存在于活跃的基因中。这个项目试图理解为什么这种模式是正确的。利用植物作为研究生物，遗传学工具已被用于将基因体甲基化引入通常缺乏这种甲基化的植物基因组中。这个实验操作模拟了基因体甲基化的进化起源。因此，通过研究对基因表达和多代适应度性状的影响，该项目将为这种有趣的DNA甲基化模式的起源和功能提供新的见解。该项目还将支持几项教育活动。该研究将由一名博士后和一名研究生进行，他们将接受实验和计算生物学方面的培训。将开发开源软件和教学材料，通过计算机模拟解决问题来教授遗传学入门原理，作为传统讲座的补充。高中学生将在夏季实习期间得到佐治亚大学青年犬项目的指导。基因启动子中的胞嘧啶DNA甲基化通常与转录抑制有关。相比之下，基因体甲基化（gbM）通常发生在活跃转录的基因上。这种DNA甲基化与基因表达的关系表明gbM具有不同的功能。在哺乳动物中，这些功能中的一些是已知的，而在植物和昆虫中，这些功能似乎是不同的，尽管尚未明确描述。在任何情况下，在基因体中建立DNA甲基化的机制都没有被理解。为了深入了解gbM的起源和特异性，本项目将使用两个相关的植物物种作为模型：拟南芥（其中gbM由CMT3甲基转移酶催化）和Eutrema salsigineum（缺乏CMT3和gbM）。在初步研究中，拟南芥CMT3基因导入Eutrema会导致gbM。为了遵循这一观察结果，实验将测试几种假设，以了解gbM的分子基础，哪些基因获得gbM，这些基因如何靶向，以及新发现的染色体组蛋白修饰（H3K23甲基化）可能发挥的作用。这些结果有可能揭示DNA甲基化在高等真核生物中调节染色质介导的基因表达中的重要未被发现的功能。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

In the genomes of most eukaryotes, methylation of cytosine bases in DNA is a common modification that has long been believed to down-regulate, or turn off, gene expression. This view has been challenged by recent studies showing that the effect on gene expression depends on where the methylation occurs. In many organisms including humans and some plants, DNA methylation located next to a gene turns the gene off, whereas DNA methylation located within the gene-body is found in genes that are active. This project seeks to understand why this pattern is true. Using plants as study organisms, genetic tools have been used to introduce gene-body methylation into the genome of a plant that normally lacks this type of methylation. This experimental manipulation simulates the evolutionary origin of gene-body methylation. Therefore, by studying the resulting effects on gene expression and fitness traits over multiple generations, this project will provide new insights about the origins and function of this intriguing pattern of DNA methylation. The project will also support several educational activities. The research will be carried out by a postdoctoral fellow and a graduate student, who will be trained in experimental and computational biology. Open-source software and instructional materials will be developed for teaching introductory principles of genetics via computer-simulated problem-solving, as a complement to traditional lectures. High school students will be mentored during summer internships sponsored by the University of Georgia Young Dawgs Program. Cytosine DNA methylation in the promoters of genes is typically associated with transcriptional repression. By contrast, gene-body methylation (gbM) is often found on actively transcribed genes. This relationship of DNA methylation to gene expression suggests that gbM has distinct functions. In mammals some of these functions are known, whereas in plants and insects, these functions appear to be distinct, though as yet poorly characterized. In no case are the mechanisms that establish DNA methylation in gene bodies understood. To gain insight into the origin and specificity of gbM, this project will use two related plant species as models: Arabidopsis thaliana (in which gbM is catalyzed by the CMT3 methyltransferase) and Eutrema salsigineum (which lacks CMT3 and gbM). In preliminary studies, introduction of the Arabidopsis CMT3 gene into Eutrema leads to gbM. To follow this observation, experiments will test several hypotheses to understand the molecular basis for gbM, what genes acquire gbM, how those genes are targeted, and what role may be played by a newly identified chromosomal histone modification (H3K23 methylation). The outcomes have the potential to reveal important undiscovered functions of DNA methylation in regulating chromatin-mediated gene expression in higher eukaryotes.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.