Understanding epigenetic remodeling in primordial germ cells

了解原始生殖细胞的表观遗传重塑

基本信息

批准号：
8898861
负责人：
Amander Clark
金额：
$ 31.16万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-02-05 至 2019-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8898861
关键词：
Address Alleles Binding Birth Cell Differentiation process Child Cytosine DNA DNA Methylation Data Development Dioxygenases Disease Environment Enzymes Epigenetic Process Excision Fertilization Fingers First Pregnancy Trimester Funding Future Future Generations Gametogenesis Generations Genes Genome Genome Mappings Germ Cells Germ Lines Goals Gonadal structure Grant Health Homologous Gene Human In Vitro Individual Inherited Knock-out Laboratories Lead Left Mammals Maps Methylation Mus Parents Phase Pregnancy Production Protein-Arginine N-Methyltransferase Proteins Regulation Repetitive Sequence Repression Research Resolution Retrotransposon Risk Role Science Second Pregnancy Trimester Site Sorting - Cell Movement Staging Structure of primordial sex cell Technology Tetanus Helper Peptide Time Transgenic Mice Ubiquitin Work base bisulfite sequencing cofactor demethylation disease transmission embryonic stem cell epigenome gene repression genome-wide imprint in utero in vitro Model in vivo insight next generation prevent progenitor

项目摘要

DESCRIPTION (provided by applicant): During pregnancy three generations of DNA co-exist. Mum's, baby's and the germ line of baby. During the first and second trimester the majority of methylated cytosines from the DNA of baby's progenitor germ line cells, called primordial germ cells (PGCs) are removed. This act is essential to remove errors in methylation acquired during gametogenesis in the parents, and/or during early development of baby after fertilization. If errors in cytosine methylation are not removed, the abnormally methylated alleles have a risk of being inherited as disease epialleles in the following generation. Given that the environment can stably influence the genome, including the genome of baby's PGCs in utero, there is a need to understand the mechanisms that regulate DNA demethylation in PGCs in order to develop strategies to guard against the transmission of disease epialleles in future generations. Recently my group discovered that DNA demethylation in human PGCs is regulated in two phases however the mechanisms underlying demethylation globally (phase 1) and locally (phase 2) are unclear. In this project we aim to uncover new details on the dynamics of DNA demethylation in human PGCs and use conditional deletions of mouse PGCs in vivo as well as differentiation of mouse PGCs from embryonic stem cells in vitro to address hypotheses regarding the specific mechanisms responsible for demethylation in the mammalian germ line. In aim 1 we will identify the dynamic removal of cytosine methylation at base resolution for the very first time in human PGCs and address the hypothesis that Ubiquitin- like, containing PHD and RING finger domains, 1 (Uhrf1) repression by protein arginine methyltransferase 5 (Prmt5) is responsible for the phase 1 DNA demethylation in mammals. In aim 2 using a conditional deletion in mouse PGCs we will address the hypothesis that Dnmt1 maintains cytosine methylation at discreet loci in PGCs in the absence of its major cofactor Uhrf1. In aim 3, we turn to phase 2 demethylation to directly address the hypothesis that conversion of 5-methylcytosine to 5-hydroxymethylcytosine by Tet methylcytosine dioxygenases has a functional role in the demethylation of imprinting control centers in PGCs. Taken together, results from this grant will lead to new insights into the mechanisms that regulate germ line epigenetic inheritance, and in future work our goal will be to prevent epialleles from being acquired and transmitted.

描述（由申请人提供）：在怀孕期间，三代 DNA 共存。妈妈的、婴儿的和婴儿的生殖系。在妊娠早期和中期，婴儿祖生殖系细胞（称为原始生殖细胞 (PGC)）DNA 中的大部分甲基化胞嘧啶被去除。这一行为对于消除父母配子发生期间和/或受精后婴儿早期发育期间获得的甲基化错误至关重要。如果胞嘧啶甲基化错误未被消除，异常甲基化的等位基因就有可能作为疾病表观等位基因遗传到下一代。鉴于环境可以稳定地影响基因组，包括子宫内婴儿 PGC 的基因组，因此有必要了解调节 PGC 中 DNA 去甲基化的机制，以便制定预防疾病表观等位基因在后代中传播的策略。最近，我的团队发现人类 PGC 中的 DNA 去甲基化分两个阶段进行调节，但全局（第 1 阶段）和局部（第 2 阶段）去甲基化的机制尚不清楚。在这个项目中，我们的目标是揭示人类 PGC 中 DNA 去甲基化动态的新细节，并利用体内小鼠 PGC 的条件删除以及体外小鼠 PGC 与胚胎干细胞的分化来解决有关去甲基化具体机制的假设在哺乳动物种系中。在目标 1 中，我们将首次在人类 PGC 中以碱基分辨率动态去除胞嘧啶甲基化，并提出这样的假设：含有 PHD 和环指结构域的泛素样蛋白 1 (Uhrf1) 被蛋白精氨酸甲基转移酶 5 抑制（ Prmt5) 负责哺乳动物的第一阶段 DNA 去甲基化。在目标 2 中，我们将在小鼠 PGC 中使用条件删除来解决这样的假设：在缺乏主要辅因子 Uhrf1 的情况下，Dnmt1 在 PGC 中的离散位点维持胞嘧啶甲基化。在目标 3 中，我们转向第 2 阶段去甲基化，以直接解决这样的假设：Tet 甲基胞嘧啶双加氧酶将 5-甲基胞嘧啶转化为 5-羟甲基胞嘧啶，在 PGC 印记控制中心的去甲基化中具有功能性作用。总而言之，这项资助的结果将带来对调节种系表观遗传机制的新见解，在未来的工作中，我们的目标将是防止表观等位基因的获得和传播。