Protecting and Sustaining Germ Cell Identity

保护和维持生殖细胞身份

基本信息

批准号：
10736134
负责人：
RUTH LEHMANN
金额：
$ 41.93万
依托单位：
WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-11 至 2028-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10736134
关键词：
ATAC-seq Address Binding Binding Sites Biological Assay Cell Reprogramming Cells Chromatin Chromatin Remodeling Factor DNA Transposable Elements DNA damage checkpoint Deposition Development Elements Embryo Embryonic Development Enzymes Face Gene Activation Gene Expression Gene Expression Regulation Genes Genetic Genetic Transcription Genome Germ Germ Cells Goals Human Immunity Inherited Longevity Mediating Methods Modeling Molecular Mus Mutation Oogenesis Pathway interactions Phenotype Process Proteins RNA RNA Editing RNA-Binding Proteins Regulation Regulatory Pathway Reporter Repression Role Site Small RNA Somatic Cell Specific qualifier value Structure of primordial sex cell System Testing Totipotency Trans-Activators Transact Transcriptional Activation Transcriptional Regulation Translational Repression Translations Validation candidate identification cell type egg expectation experimental study fly member mutant nano nanobodies neuronal cell body next generation novel permissiveness piRNA preempt premature prevent programs reproductive ribosome profiling single-cell RNA sequencing sperm cell transcription factor transmission process tumor

项目摘要

Project Summary/Abstract Germ cells have the extraordinary potential to generate every cell type in the body. Yet, the molecular program that protects and sustains this promise for totipotency in germ cells, which can last more than forty years in humans, is poorly understood. To face this challenge, germline regulators assume dual roles in maintaining germ cell identity: activating a germline transcriptional program while simultaneously protecting germ cells from reprogramming to a somatic cell fate. Indeed, genes involved in germline specification, such as the conserved RNA-binding protein Nanos and the transposable element (TE) regulator Piwi, are erroneously upregulated in certain human tumors. These tumors are thought to undergo soma-to-germline transformations to acquire a more immortal, germline-like state. However, a specific program for germ cell transcriptional activation has yet to be described, mainly because a ‘master-regulator transcription factor’ for germ cell fate is missing. This proposal uses a multipronged approach to systematically characterize the gene expression program in primordial germ cells (PGCs) and identify prime regulators of germ cell fate. The goal is to distinguish between a ‘default’ model, by which repression of the somatic program ‘allows’ the PGC program, and an ‘instructive’ model, by which PGC specification is actively controlled. In Aim1, we probe the instructive model to uncover the germline transcriptional program. By single-cell RNA sequencing, we have detected specific, temporally-regulated germline genes. To identify the transacting regulatory factors, we will use ATAC-sequencing, characterize known factors with matching binding activities, and identify potential new regulatory candidates for germline genes. In Aim2, we address the function of a critical regulator of germ cell fate, the conserved RNA regulator Nanos. Using the RNA target identification method hyper-TRIBE and ribosome profiling, we will identify Nanos targets and functionally distinguish between those targets that promote the germline program and/or repress the somatic program. Aim3 focuses on the piRNA pathway, which provides maternally inherited immunity against TE activity by controlling the transcription of TE elements, specifically in germ cells. Using a novel degradation strategy to specifically degrade components of the piRNA pathway in PGCs without impacting oogenesis, we will determine the role of this pathway for embryonic PGC gene expression beyond TE control. By integrating multiplex findings, this proposal will uncover the regulatory landscape of early germ cells, thereby providing a necessary understanding of the process of gonadogenesis and, ultimately, the survival of the species.

项目概要/摘要然而，生殖细胞具有产生体内每种细胞类型的非凡潜力。保护和维持生殖细胞全能性的承诺，这种全能性可以持续四十多年人类对这一挑战知之甚少，生殖系调节因子在维持细菌方面发挥着双重作用。细胞身份：激活生殖系转录程序，同时保护生殖细胞免受事实上，基因涉及种系特化，例如保守的基因。 RNA 结合蛋白 Nanos 和转座元件 (TE) 调节因子 Piwi 在某些人类肿瘤被认为会经历体细胞到种系的转变以获得然而，生殖细胞转录激活的特定程序尚未出现。待描述，主要是因为生殖细胞命运的“主调节转录因子”缺失。该提案使用多管齐下的方法来系统地描述原始基因表达程序生殖细胞（PGC）并识别生殖细胞命运的主要调节因子，目标是区分“默认”细胞。模型，通过抑制体细胞程序“允许”PGC程序，以及“指导性”模型，通过 PGC 规范是主动控制的。在 Aim1 中，我们探讨了揭示种系的指导模型。通过单细胞 RNA 测序，我们检测到了特定的、时间调控的转录程序。为了识别交易调节因子，我们将使用 ATAC 测序来表征已知的基因。具有匹配结合活性的因素，并确定种系基因的潜在新调控候选者。目标2，我们利用 Nanos 来研究生殖细胞命运的关键调节因子——保守的 RNA 调节因子的功能。 RNA靶标识别方法hyper-TRIBE和核糖体分析，我们将识别Nanos靶标并在功能上区分促进种系程序和/或抑制体细胞的目标 Aim3 计划重点关注 piRNA 途径，该途径提供针对 TE 活性的母系遗传免疫力。通过控制 TE 元件的转录，特别是在生殖细胞中，使用一种新的降解策略来控制 TE 元件的转录。特异性降解 PGC 中 piRNA 途径的成分而不影响卵子发生，我们将确定通过整合多重研究结果，该途径对胚胎 PGC 基因表达的作用超出了 TE 的控制。该提案将揭示早期生殖细胞的监管格局，从而提供必要的了解性腺发生的过程以及最终物种的生存。