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

• 批准号: 10705982
• 负责人: Zongliang Jiang
• 金额: $ 46.91万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 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/10705982
• 关键词: DNA; Methylation; N6; Adenine; Placental

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.

滋养细胞在胎儿与母亲之间的交流中起着至关重要的作用。无法准确建立滋养细胞谱系会导致胚泡阶段，植入失败，早产和胎盘异常之前的发育停滞。最近，我们在哺乳动物基因组中首次发现了在N6-腺嘌呤（N6-MA）上的新型DNA甲基化（Wu等人，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剂/滋养细胞谱系中，并且在小鼠和牛中保守，并且可能在所有哺乳动物中。 AlkBH1（N6-MA脱甲基酶） - 由于失去印迹对照和植入后胚胎的严重胎盘发育缺陷而使子宫内的小鼠缺陷型。有趣的是，ALKBH1缺乏的牛胚胎会损害滋养细胞的出现和胚泡形成。此外，在与人多能干细胞区分的滋养细胞培养物中，N6-MA丰度非常高，为研究人体胎盘中的N6-MA提供了极好的模型。这些结果表明，这种新颖的表观遗传标记在不同的欧地人的滋养细胞发展中具有保守和独特的作用。然而，N6-MA功能的分子机制及其在滋养剂和滋养细胞发育中的作用仍然难以捉摸。总体假设是N6-MA代表了一个新型的表观遗传标记，该标记调节基因表达网络指导滋养细胞命运的测定。拟议研究的总体目的是对小鼠和牛胚胎的滋养细胞进行系统的比较研究，以及人类滋养细胞细胞培养模型，重点是N6-MA在滋养细胞形成，基因表达和表观遗传重编程中的作用。提出了三个独立但与智力有关的特定目标。 AIM 1将以滋养细胞特异性的有条件敲除小鼠模型来表征N6-MA函数在滋养细胞谱系分化和印迹基因控制中。 AIM 2将研究N6-MA在牛胚胎模型中N6-MA在滋养细胞干细胞出现和胚泡形成中的作用。 AIM 3将利用人类干细胞衍生的滋养细胞分化模型在人胎盘滋养细胞形成中建立N6-MA功能。该项目利用江安实验室在胚胎/滋养细胞开发方面的专业知识以及表观遗传调节中的小小的实验室的专业知识来解决滋养细胞谱系发展中的基本问题。 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.拟议的项目很重要，因为它将提高我们对与异常胎盘相关的早期胚胎丧失和临床生殖障碍的分子基础的理解。