DNA Methylation at N6-Adenine in Placental Trophoblast Development

胎盘滋养层发育中 N6-腺嘌呤 DNA 甲基化

基本信息

批准号：
10033546
负责人：
Zongliang Jiang
金额：
$ 64.53万
依托单位：
LOUISIANA STATE UNIV AGRICULTURAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10033546
关键词：
Abnormal placentation Address Adenine Alleles Biology Cattle Cell Culture Techniques Cell Lineage Cells Clinical Communication Comparative Study Complex DNA Methylation Defect Development Disease Elements Embryo Embryo Loss Embryonic Development Enhancers Ensure Epigenetic Process Experimental Models Failure Fetal Growth Retardation Fetus Gene Expression Gene Targeting Genes Genetic Genetic Transcription Genomics Human Human Genome Hypoxia Impairment In Vitro Knock-out Knockout Mice Laboratories Lead Light Locus Control Region Mammals Maps Modeling Modification Molecular Mothers Mus Nature Nucleic Acid Regulatory Sequences Pathology Physiological Placenta Placentation Play Pluripotent Stem Cells Positioning Attribute Pre-Eclampsia Pre-implantation Embryo Development Regulatory Element Research Role Signal Pathway Single Nucleotide Polymorphism Spontaneous abortion Study models Testing Uterus Work blastocyst cancer cell comparative conditional knockout demethylation embryonic cell culture epigenetic regulation failure Implantation human pluripotent stem cell human stem cells imprint improved in utero in vivo innovation insight interest mammalian genome mouse model natural Blastocyst Implantation novel overexpression reproductive response self-renewal somatic cell nuclear transfer stem cell differentiation stem cell model stem cells trophoblast

项目摘要

Trophoblasts play an essential role in communication between the fetus and the mother. Failure to accurately establish the trophoblast lineage results in developmental arrest prior to the blastocyst stage, implantation failure, early miscarriage, and placental abnormalities. Recently, we made the first discovery of a novel DNA methylation on N6-adenine (N6-mA) in mammalian genomes (Wu et al., Nature, 2016; Xie et al., Cell, 2018). Our most recent preliminary studies have shown that N6-mA is predominantly present in the trophectoderm/trophoblast lineage of early embryos, and is conserved in the mouse and bovine and probably across all mammals. Alkbh1 (N6-mA demethylase)-deficient in mice die in utero due to loss of imprinted control and severe placental development defects in post-implantation embryos. Interestingly, ALKBH1 deficient bovine embryos have impaired trophoblast stem cell emergence and blastocyst formation. In addition, N6-mA abundance is remarkably high in trophoblast cultures differentiated from human pluripotent stem cells, providing an excellent model for studying N6-mA in human placentation. These results indicated that this novel epigenetic mark has both conserved and unique roles in trophoblast development in different eutherians. However, the molecular mechanism underlying N6-mA function, and its role in trophectoderm and trophoblast development remain elusive. The overall hypothesis is that N6-mA represents a novel epigenetic mark that regulates gene expression network directing determination of trophoblast cell fate. The overall objective of the proposed studies is to conduct a systematic, comparative study on trophoblast from mice and bovine embryos, as well as human trophoblast cell culture models, focusing on the role of N6-mA in trophoblast formation, gene expression and epigenetic reprogramming. Three independent and yet intellectually-related specific aims are proposed. Aim 1 will characterize N6-mA function in trophoblast lineage differentiation and imprinted gene control with a trophoblast-specific conditional knockout mouse model. Aim 2 will investigate the role of N6-mA in trophoblast stem cell emergence and blastocyst formation in a bovine embryo model. Aim 3 will utilize human stem cell-derived trophoblast differentiation models to establish N6-mA functions in human placental trophoblast formation. This project leverages the expertise of the Jiang laboratory in embryo/trophoblast development and the expertise of the Xiao laboratory in epigenetic regulation to address fundamental questions in trophoblast lineage development. This study is innovative because it a) provides insight into the role of a novel DNA methylation mark in trophoblast lineage formation, b) sheds new light on epigenetic regulation of imprinted genes, a longstanding issue in placental development, and c) establishes novel, comparative embryo and cell culture models to understand the conserved and unique epigenetic regulation of trophoblast cell fate in eutherians. The proposed project is significant because it will improve our understanding of the molecular basis of early embryonic loss and clinical reproductive disorders that are associated with abnormal placentation.

滋养层细胞在胎儿和母亲之间的沟通中起着至关重要的作用。未能准确建立滋养层谱系会导致囊胚期之前发育停滞、着床失败、早期流产和胎盘异常。最近，我们首次发现哺乳动物基因组中N6-腺嘌呤（N6-mA）上的一种新的DNA甲基化（Wu等，Nature，2016；Xie等，Cell，2018）。我们最近的初步研究表明，N6-mA 主要存在于早期胚胎的滋养外胚层/滋养层谱系中，并且在小鼠和牛中以及可能在所有哺乳动物中保守。缺乏 Alkbh1（N6-mA 去甲基酶）的小鼠会因印迹控制丧失和植入后胚胎严重胎盘发育缺陷而在子宫内死亡。有趣的是，ALKBH1 缺陷的牛胚胎会损害滋养层干细胞的出现和囊胚的形成。此外，在人多能干细胞分化的滋养层培养物中，N6-mA 丰度非常高，为研究人胎盘形成中的 N6-mA 提供了一个极好的模型。这些结果表明，这种新的表观遗传标记在不同真兽类滋养层发育中具有保守且独特的作用。然而，N6-mA 功能的分子机制及其在滋养外胚层和滋养层发育中的作用仍不清楚。总体假设是，N6-mA 代表一种新的表观遗传标记，可调节基因表达网络，指导滋养层细胞命运的决定。拟议研究的总体目标是对小鼠和牛胚胎的滋养层以及人类滋养层细胞培养模型进行系统的比较研究，重点关注 N6-mA 在滋养层形成、基因表达和表观遗传重编程中的作用。提出了三个独立但在智力上相关的具体目标。目标 1 将通过滋养层特异性条件敲除小鼠模型来表征 N6-mA 在滋养层谱系分化和印记基因控制中的功能。目标 2 将研究 N6-mA 在牛胚胎模型中滋养层干细胞出现和囊胚形成中的作用。目标 3 将利用人类干细胞衍生的滋养层分化模型来建立 N6-mA 在人类胎盘滋养层形成中的功能。该项目利用江实验室在胚胎/滋养层发育方面的专业知识和肖实验室在表观遗传调控方面的专业知识来解决滋养层谱系发育的基本问题。这项研究具有创新性，因为它a）深入了解了新型DNA甲基化标记在滋养层谱系形成中的作用，b）为印迹基因的表观遗传调控（胎盘发育中长期存在的问题）提供了新的线索，以及c）建立了新颖的、比较性的方法胚胎和细胞培养模型，以了解真兽类滋养层细胞命运的保守且独特的表观遗传调控。该项目意义重大，因为它将提高我们对早期胚胎丢失和与胎盘异常相关的临床生殖疾病的分子基础的理解。