Epigenetic regulation of dioxin response elements during mammalian development

哺乳动物发育过程中二恶英反应元件的表观遗传调控

• 批准号: RGPIN-2022-03647
• 负责人: Jones, Meaghan
• 金额: $ 2.26万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=744195
• 关键词: Epigenetic; regulation; dioxin; response; elements

The long-term objective of my research program is to understand the role of epigenetics in biological embedding of environmental exposures. I am interested in how DNA methylation (DNAm) helps cells remember past exposures and influences their future responses. The Aryl Hydrocarbon Receptor (AhR) pathway is one of the major metabolic routes of detoxification in the mammalian cell. Its primary ligands are dioxins, which are produced by combustion processes and are common products of forest fire smoke, combustion engines, and metal or petroleum refining. Upon stimulation, AhR activates genes involved in detoxification by binding to Dioxin Response Elements (DREs). There is some evidence that the cellular response to dioxins undergoes priming - that is, a second exposure to dioxins induces a greater response than the first, which implies that the cells retain a memory of the initial exposure. Importantly, the consensus binding site for DREs contains a CpG dinucleotide, which is the primary location of DNAm in the mammalian genome. However, we do not understand how epigenetic marks regulate DREs, how this priming effect is encoded in the epigenome, nor the specific role of DNAm compared to other epigenetic marks. My overarching hypothesis is that DNAm alterations are not required for induction of DREs, but are required for long-term priming of response. To test this hypothesis, in the next five years I propose to investigate three specific aspects of epigenetic regulation of DREs. 1. Establish the kinetics of DNAm, open chromatin, and AhR occupancy during dioxin response. First, we will investigate the timing of DNAm, AhR occupancy, and histone modifications at DREs during timecourses of stimulation with TCDD, a canonical dioxin. 2. Determine whether previous exposure to dioxins influences priming at DREs though epigenetic marks. Here, we will develop an exposure/re-exposure paradigm to detect the presence of primed responses, and compare priming in cells from early embryos as well as from adults, to see whether priming differs by developmental stage. 3. Delineate the specific role of DNAm in induction and priming of the dioxin response. Finally, we will compare induction and priming data from parts 1 and 2 in DREs which naturally contain different numbers of CpGs. We will also repeat the priming experiments in a system which is incapable of removing DNAm from DREs, to determine whether induction and/or priming are still occurring when DNAm levels do not change. The work proposed here will build a foundation for future work on cell-to-cell heterogeneity in cellular responses, as well as investigation of factors which can modify induction of the AhR pathway. HQP working in this research program will have the opportunity to gain skills and experience in a broad range of areas, setting them up for success in their future careers.

我的研究计划的长期目标是了解表观遗传学在环境暴露的生物嵌入中的作用。我对DNA甲基化（DNAm）如何帮助细胞记住过去的暴露并影响它们未来的反应感兴趣。芳烃受体（Aryl Hydrocarbon Receptor, AhR）途径是哺乳动物细胞解毒的主要代谢途径之一。它的主要配体是二恶英，由燃烧过程产生，是森林火灾烟雾、内燃机和金属或石油精炼的常见产物。在刺激下，AhR通过结合二恶英反应元件（DREs）激活参与解毒的基因。有一些证据表明，细胞对二恶英的反应经历了启动——即，第二次接触二恶英会引起比第一次更大的反应，这意味着细胞保留了对初次接触二恶英的记忆。重要的是，DREs的一致结合位点包含CpG二核苷酸，这是哺乳动物基因组中dna的主要位置。然而，我们不了解表观遗传标记如何调节DREs，这种启动效应如何在表观基因组中编码，也不了解与其他表观遗传标记相比，DNAm的具体作用。我的首要假设是，DNAm的改变不是诱发DREs所必需的，但却是长期启动反应所必需的。为了验证这一假设，我建议在未来五年内研究DREs表观遗传调控的三个具体方面。1. 建立二恶英反应过程中dna、开放染色质和AhR占用的动力学。首先，我们将研究TCDD（一种典型的二恶英）刺激期间DREs中DNAm、AhR占用和组蛋白修饰的时间。2. 通过表观遗传标记确定以前接触二恶英是否会影响DREs的启动。在这里，我们将开发一个暴露/再暴露范式来检测启动反应的存在，并比较早期胚胎细胞和成人细胞的启动，看看启动是否因发育阶段而异。3. 描述DNAm在诱导和启动二恶英反应中的具体作用。最后，我们将比较DREs中自然含有不同数量CpGs的第1部分和第2部分的诱导和启动数据。我们还将在一个无法从DREs中去除DNAm的系统中重复启动实验，以确定当DNAm水平没有变化时，诱导和/或启动是否仍在发生。本文提出的工作将为进一步研究细胞反应的细胞间异质性以及研究可以改变AhR通路诱导的因素奠定基础。在这个研究项目中工作的HQP将有机会获得广泛领域的技能和经验，为他们未来的职业生涯奠定基础。