Novel Roles of Pluripotency Regulators in the Early Mouse Embryo

描述（由申请人提供）：控制胚胎干细胞（ESCs）多能性和分化的基因调控网络被广泛研究并得到很好的理解。然而，成功的体细胞核移植的遗传参数和建立ESC基因网络的遗传程序尚不清楚。ESC研究中这些关键问题的答案在于早期胚胎的基因调控，就我们的目的而言，早期胚胎包括受精后和囊胚形成之前的发育阶段。就像囊胚的内部细胞群必须保持多能性和自我更新的能力才能产生形成胎儿的细胞系一样，早期胚胎的卵裂球必须建立全能性和自我更新的能力来形成囊胚。然而，我们不知道卵裂球的全能性是如何实现的，以及它与胚胎基因组激活波的关系。因此，我们的总体目标是解决这个基因网络的一般结构，并确定在1到2细胞阶段建立正确的遗传回路至关重要和足够的特定调节因子，并确保随后的发育能力。此外，这些调节因子的功能将被确定，以了解在早期胚胎发育中必不可少的机制。我们已经建立了实验策略来询问转录调节因子在早期胚胎母胚转化过程中的确切作用，当母体和早期胚胎转录本可能同时存在时。我们发现Oct4是POU家族的同源结构域转录因子，在内细胞群、ESCs和生殖细胞的多能性中具有关键功能，在囊胚期之前的早期胚胎发育中具有新的作用，并且在多细胞和桑葚胚期之后的发育中是必需的。此外，我们的数据表明，多能性调节因子Sox2和Sall4也可能在囊胚形成之前具有关键功能。我们建议充分研究这些和其他多能性调控因子的新作用，并将它们作为“门户”，在核重编程和胚胎基因组激活的背景下解剖早期胚胎中的基因调控。了解早期胚胎的遗传需求将对再生和干细胞医学、不孕症治疗和癌症研究产生直接而重要的影响，因为与癌症有关的基因在这一遗传程序中高度丰富。

DESCRIPTION (provided by applicant): The gene regulatory network that controls pluripotency and differentiation in embryonic stem cells (ESCs) is widely studied and becoming well understood. However, the genetic parameters of successful somatic cell nuclear transfer and the genetic program that sets up the ESC gene network are not known. The answers to these key questions in ESC research lie in the gene regulation of the early embryo, which for our purpose here, encompasses developmental stages that occur after fertilization and prior to blastocyst formation. Just as the inner cell mass of the blastocyst must maintain pluripotency and the ability to self-renew in order to give rise to cell lineages that form the fetus, the blastomeres of the early embryo must establish totipotency and self-renewal abilities to form the blastocyst. However, we do not know how totipotency of blastomeres is achieved and how it relates to the waves of embryonic genome activation. Therefore, our overall goal is to address both the general architecture of this gene network and identify specific regulators that are critical and sufficient for establishing the correct genetic circuitry at the 1- to 2-cell stages, and that would ensure subsequent developmental competence. Further, the function of these regulators will be determined in order to understand mechanisms that are essential in early embryo development. We have established experimental strategies to interrogate the precise roles of transcriptional regulators during the maternal-embryonic transition of the early embryo, when both maternal and early embryonic transcripts may be present simultaneously. We discovered that Oct4, a homeodomain transcription factor of the POU family that is known for its critical functions in pluripotency in the inner cell mass, ESCs, and germ cells, has a novel role in early embryo development prior to the blastocyst stage and is required for progression beyond the multi-cell and morula stages. In addition, our data suggest that the pluripotency regulators, Sox2 and Sall4, may also have critical functions prior to blastocyst formation. We propose to fully investigate the novel roles of these and other pluripotency regulators and use them as "portals" to dissect gene regulation in the early embryo in the context of nuclear reprogramming and embryonic genome activation. Understanding the genetic requirements of the early embryo will have a direct and significant impact on regenerative and stem cell medicine, the treatment of infertility, and cancer research, as genes implicated in cancer are highly enriched in this genetic program. Public Health Relevance: The overall goal of this project is to investigate the novel roles of Oct4, Sall4, Sox2, and other pluripotency regulators in the early mouse embryo, with a focus on the 1- to 2-cell stages during the maternal-embryonic transition. Using these transcriptional regulators as "portals", we will dissect the gene regulatory network in the early embryo and identify specific regulators that are critical and sufficient for ensuring subsequent developmental competence. Understanding the genetic requirements of the early embryo will have a direct and significant impact on regenerative and stem cell medicine, the treatment of infertility, and cancer research.