Transcriptional and Epigenetic Control of Pluripotency and Self-Renewal by Honey Bee Royalactin and its human structural analog

蜜蜂 Royalactin 及其人类结构类似物对多能性和自我更新的转录和表观遗传控制

• 批准号: 10646468
• 负责人: Kevin Chun-Kai Wang
• 金额: $ 39.61万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-15 至 2024-01-18
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10646468
• 关键词: Affect; Automobile Driving; Back; Binding; Biological Assay; Biology; Cell Maintenance; Cell Surface Receptors; Cell surface; Cells; Cellular Assay; Chromatin; Complex; Congenital Abnormality; Data; Degenerative Disorder; Derivation procedure; Development; Developmental Process; Diabetes Mellitus; Disease; Disease model; Embryo; Embryology; Embryonic Development; Epigenetic Process; Event; Gene Expression; Gene Family; Genetic; Genetic Transcription; Goals; Growth Factor; Heterogeneity; Human; Human Development; Impairment; Ligands; Maintenance; Malignant Neoplasms; Mammalian Cell; Mammals; Mediating; Membrane; Methods; Mission; Modeling; Molecular; Mus; Nature; Phenotype; Pluripotent Stem Cells; Proteins; Proteomics; Public Health; Receptor Mediated Signal Transduction; Regenerative Medicine; Regulator Genes; Signal Pathway; Signal Transduction; Specific qualifier value; Stem Cell Factor; Stem cell pluripotency; Stimulus; Study models; Therapeutic; Transposase; United States National Institutes of Health; Work; analog; blastomere structure; candidate validation; cell behavior; developmental disease; embryo cell; embryonic stem cell; epigenetic regulation; epigenome; epigenomic profiling; flexibility; forward genetics; gene regulatory network; genome-wide analysis; human disease; human embryonic stem cell; implantation; improved; improved outcome; insight; loss of function; novel; novel strategies; pluripotency; pluripotency factor; preservation; receptor; regenerative; regenerative cell; regenerative therapy; response; self-renewal; single cell analysis; stem; stem cell biology; stem cell differentiation; stem cell fate; stem cell self renewal; stem cells; stemness; tissue repair; transcriptome; transcriptome sequencing; transcriptomics; whole genome

Limitations in current treatment options for many congenital and acquired diseases in humans, including birth defects, cancer, degenerative disorders and diabetes, highlight the need for the development of novel approaches for regenerative medicine to dramatically improve tissue repair. The pluripotent nature of mammalian embryonic stem cells (ESCs) makes them a convenient model for studying aspects of early development and an invaluable starting point for deriving numerous therapeutically relevant cells for regenerative medicine. Despite the remarkable progress made in deciphering mechanisms driving ESC pluripotency, fundamental gaps remain in understanding how human embryonic stem cells (hESCs) regulate the pluripotent state. If we are to use hESCs as a high-fidelity model for embryonic development, and if we wish to improve outcomes of hESC differentiation and the fidelity of cellular reprogramming to pluripotency, then it is imperative that we understand how hESCs fit into the paradigm of mammalian embryonic development. In this project we seek to answer some of these fundamental questions, by characterizing and validating a potential alternative pluripotency state. Our paradigm-shifting hypothesis stems from our unexpected discovery that the honey bee queen-maker protein, Royalactin, and its structural analog in mammals, Regina, have unexpected robust pluripotency maintenance effects in mammalian stem cells. We hypothesize that Regina/Royalactin stabilize and capture a pivotal pluripotent state distinct from the existing pre- (naïve) and post- (primed) implantation associated stem cell states. I outline here a plan to molecularly characterize this novel cellular metastable state with 3 specific aims. In Aim 1, we will Isolate and characterize the composition and activity of the receptor complex(es) in ESCs. We hypothesize that Regina/Royalactin, as secreted molecules, likely directly interact with a receptor partner on the membrane of responsive cells to affect gene expression and subsequent cellular behavior. We will identify the receptor(s) through multiple high throughput forward genetics and proteomic strategies. In Aim 2, we will derive and maintain murine and human ESCs to functionally demonstrate that the Regina/Royalactin-mediated state of pluripotency can be related back to the signaling pathways involved in lineage specification and maintenance in the embryo itself. Establishment of a new distinct stage of mammalian pluripotency will be an important advance in our understanding of early lineage commitment. Lastly, in Aim 3 we will elucidate and characterize the critical mechanisms that interface between Regina/Royalactin and downstream epigenetic and transcriptomic events. The genome-wide analyses will be compared to current established conditions to determine whether genetic and epigenetic instability of the ESCs, associated with impaired developmental potential, exists. Taken together, our data will directly establish how a novel endogenous mammalian pluripotency factor instigates fate decisions in ESCs, and provide a new platform to study the principles governing cell potency, epigenetic regulation, and the mechanisms that regulate developmental processes in naïve pluripotent stem cells.

许多先天性和人类疾病的当前治疗方案的限制，包括 生日缺陷，癌症，退化性疾病和糖尿病，强调了发展新颖的需求 再生医学的方法可以显着改善组织修复。哺乳动物的多能性质 胚胎干细胞（ESC）使其成为研究早期发展方面和 用于获取众多治疗性相关细胞的重生医学细胞的宝贵起点。 尽管在驱动ESC多能的解密机制方面取得了显着进展，但基本差距 继续了解人类胚胎干细胞（HESC）如何调节多能状态。如果我们要 将hESC用作胚胎开发的高保真模型，如果我们希望改善hESC的结果 分化和细胞重编程对多能性的忠诚度，那么我们必须理解 HESC如何适合哺乳动物胚胎发育的范式。在这个项目中，我们试图回答一些 在这些基本问题中，通过表征和验证潜在的替代多能状态。我们的 范式转移假设始于我们意外发现的蜜蜂女王制造蛋白， 王室成分及其在哺乳动物中的结构类似物，里贾纳（Regina）具有意想不到的多能维持 哺乳动物干细胞的作用。我们假设里贾纳/王室素可以稳定并捕获关键 多能状态与现有前（幼稚）和（启动）植入相关的干细胞状态不同。 我在这里概述了一个计划，以分子表征这种新型细胞亚稳态，具有3个特定目标。目标 1，我们将隔离并表征ESC中受体复合物（ES）的组成和活性。我们 假设Regina/Royalactin作为分泌分子，可能直接与接收者伴侣在 反应性细胞的膜影响基因表达和随后的细胞行为。我们将确定 接收器通过多个高吞吐量前向遗传学和蛋白质组学策略。在AIM 2中，我们将得出 并维持鼠和人类的ESC，以表明里贾纳/王室素介导的状态 多能性可能与谱系规范和维护中有关的信号通路有关 胚胎本身。建立一个新的哺乳动物多能阶段将是重要的进步 在我们对早期血统承诺的理解中。最后，在AIM 3中，我们将阐明和表征关键 里贾纳/王室素与下游表观遗传和转录组事件之间接口的机制。 将全基因组分析与当前既定条件进行比较，以确定是否遗传和 存在与发育潜力受损相关的ESC的表观遗传不稳定。总的来说，我们的 数据将直接确定新型内源性哺乳动物多能因素如何促进脂肪决策 ESC，并提供一个新的平台来研究控制细胞效力，表观遗传调节和 调节幼稚多能干细胞中发育过程的机制。