Mechanisms of genomic reprogramming and transcriptional activation in the embryo

摘要 我们的长期目标是阐明支配基因组重组的分子机制 允许从指定的细胞类型转换到全能状态。这种转变发生得很快， 在胚胎发育的最初阶段，受精后立即有效。在这个紧要关头 在发育阶段，合子基因组处于转录静止状态。只有当细胞变成 全能的，准备分化的，广泛的合子转录启动。尽管这是一个事实 合子基因组的延迟转录激活在后生动物中几乎是普遍的，其机制 管理这一进程以及它与协调建立一个全能国家的关系仍然存在 未知。我们最近证明了果蝇蛋白Zelda(ZLD)在全球范围内起作用，促进 合子基因组的激活，并提出它是通过建立一个允许的染色质来做到这一点的 环境。通过全基因组结合研究，我们已经表明ZLD在发育早期就被结合了 到数千个基因组区域，随后驱动第一波合子转录。这些基因座是 后来被定义为开放染色质的区域，并由大量转录因子结合，这些转录因子 管理胚胎模式。通过我们的初步研究，我们可以通过以下方式定义这些机制 合子基因组被重塑以创造允许转录激活和 细胞分化。我们将使用遗传、基因组和生化策略来确定这种机制 ZLD通过以下方式成为塑造早期胚胎染色质环境的先驱因素：1) 确定ZLD在组织早期胚胎染色质结构中的作用和2)鉴定 ZLD介导的基因组激活所需的辅因子。我们将获得对ZLD的机械性见解 功能将促进我们对全能基因组是如何普遍建立的理解。最终这就是 这项工作将对在培养和培养中产生全能哺乳动物干细胞具有重要意义 在活体内。

ABSTRACT Our long-term goal is to elucidate the molecular mechanisms governing the restructuring of the genome that allows for the transition from a specified cell type to a totipotent state. This transition occurs rapidly and efficiently immediately following fertilization, during the initial stages of embryonic development. At this critical developmental stage, the zygotic genome is transcriptionally quiescent. Only when the cells have become totipotent and poised to differentiate does widespread zygotic transcription initiate. Despite the fact that this delayed transcriptional activation of zygotic genome is nearly universal among metazoans, the mechanisms governing this process and how it relates to the coordinated establishment of a totipotent state remain unknown. We have recently demonstrated that the Drosophila protein Zelda (ZLD) acts globally to facilitate the activation of the zygotic genome and propose that it is doing so by establishing a permissive chromatin environment. Through genome-wide binding studies, we have shown that ZLD is bound early in development to thousands of genomic regions that subsequently drive the first wave of zygotic transcription. These loci are later defined as regions of open chromatin and are bound by a large number of transcription factors that govern embryonic patterning. Through our initial studies, we are well positioned to define the mechanisms by which the zygotic genome is remodeled to create an environment permissive to transcriptional activation and cellular differentiation. We will use genetic, genomic, and biochemical strategies to determine the mechanism by which ZLD is acting as a pioneer factor to shape the chromatin environment of the early embryo by: 1) determining the role of ZLD in organizing the chromatin structure of the early embryo and 2) identifying cofactors required for ZLD-mediated genome activation. The mechanistic insights we will gain into ZLD function will advance our understanding of how totipotent genomes are established generally. Ultimately this work will have important implications for the generation of totipotent mammalian stem cells both in culture and in vivo.