Role of Mitochondria as Intracellular Shuttles for Nuclear Gene-regulatory Transcription Factors During Pluripotent Cell Division and Fate Specification

在哺乳动物中，受精是通过卵子和精子结合产生单细胞受精卵来实现的。这个单个细胞分裂几次，形成一个由大约100个相同细胞组成的囊胚，但随后要做出一个所谓的细胞命运决定：一些囊胚细胞发育成胚胎，其他囊胚细胞发育成胎盘，胎盘滋养发育中的胚胎。发育生物学中一个长期存在的问题是“早期囊胚中的哪些因素触发了细胞命运的决定？”这个项目将探索这样一种假设，即细胞的能量生产工厂，被称为线粒体，通过充当运送驱动细胞交替命运的因子的班车，来指导细胞命运的决定。新技术将用于比较小鼠胚胎和胎盘细胞的线粒体特征，如大小、能量分布和蛋白质组成。这些结果有望揭示细胞命运决定的关键步骤，并揭示线粒体在这一过程中以前未知的作用。在这项研究中，来自不同背景的本科生和研究生将参与创新培训活动，使用最先进的科学工具，在生物学和技术方面建立新的发现，旨在更深入地了解细胞命运是如何调节的。该项目还将实施一项创新的团队指导战略，为有兴趣从事科学、技术、工程、数学（STEM）职业的本科生提供更多参与研究的机会，重点关注那些在STEM领域历史上代表性不足的学生群体。在器官发育、功能、衰老和疾病中，破解直接细胞命运的核心机制仍然是一个高度优先的研究领域。哺乳动物的第一个细胞命运决定发生在着床前胚胎发生的后期，它驱动了内细胞群（注定成为胚胎）和滋养外胚层（注定成为胎盘）的形成。尽管已经提出了几个模型来解释这一过程是如何协调的，但胚胎细胞谱系规范的实际启动驱动因素的身份仍然未知。迄今为止发现的最早的命运决定信号之一是TEA结构域转录因子4 (Tead4)，它激活TE身份所需基因的转录。支持Tead4在细胞命运规范中的关键作用，在小鼠中，Tead4基因的破坏会产生不能产生TE的毛囊，导致胚胎死亡。由于Tead4被认为是胚胎细胞命运决定中最早的信号之一（如果不是最早的信号），我们仍然不清楚Tead4的功能如何仅限于那些注定要形成TE的细胞。该项目将把线粒体亚群分析的技术进步与线粒体谱系追踪方法相结合，以检验线粒体亚型的差异分配假说，线粒体亚型在胚胎发育期间的生化特性和蛋白质组学景观（包括Tead4内化）不同，驱动细胞谱系命运决定。通过测试线粒体亚型异质性和细胞器间通讯对TE和ICM规范的重要性，该项目将评估线粒体异质性是否指导细胞命运的决定，可能是通过线粒体在不对称细胞分裂过程中作为转录因子“穿梭体”的尚未发现的作用。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

In mammals, fertilization occurs through the joining of an egg and a sperm to produce a single-cell zygote. This single cell divides a few times to form a blastocyst composed of about 100 identical cells, but then a so-called cell fate decision is made: some blastocyst cells will develop into the embryo, and the others will develop into the placenta, which nourishes the developing embryo. A long-standing question in developmental biology is “What factors in the early blastocyst trigger cell-fate decision-making?” This project will explore the hypothesis that the energy-producing factories of cells, called mitochondria, guide cell-fate decisions by acting as shuttles for the delivery of factors that drive alternate cell fates. New technology will be applied to compare mitochondrial features--such as size, energy profiles, and protein makeup--in mouse embryo versus placental cells. The results are expected to shed light on key steps of cell-fate decision-making and uncover a previously unknown role for mitochondria in this process. During this research, undergraduate and graduate students from diverse backgrounds will participate in innovative training activities using state-of-the-art scientific tools to forge new discoveries in biology and technology, geared towards development of a deeper understanding of how cell fate is regulated. The project also will implement an innovative team-mentorship strategy to expand opportunities for undergraduate students interested in pursuing Science, Technology, Engineering, Mathematics (STEM)-based careers to engage in research, with a focus on student populations who are historically underrepresented in STEM.Deciphering the core mechanisms that direct cell fate remains a high-priority research area in organ development, function, aging, and disease. The first cell fate decision in mammals, which drives formation of the inner cell mass (destined to become the embryo) and the trophectoderm (TE; destined to become the placenta), occurs during the later stages of preimplantation embryogenesis. Although several models have been offered to explain how this process is orchestrated, the identity of the actual initiating driver(s) of embryonic cell lineage specification remains unknown. One of the earliest fate-determination signals identified thus far is TEA Domain transcription factor 4 (Tead4), which activates transcription of genes required for TE identity. Supporting its critical role in cell fate specification, disruption of the Tead4 gene in mice generates morulae that fail to generate TE, resulting in embryonic lethality. With Tead4 being identified as one of the, if not the, earliest signal(s) in embryonic cell fate decision-making, it remains unclear how Tead4 function is restricted only to those cells destined to form the TE. This project will combine a technological advancement in mitochondrial subpopulation analysis with a mitochondrial lineage-tracing approach to test the hypothesis that differential allocation of mitochondrial subtypes, which differ in their biochemical properties as well as in their proteomic landscapes--including Tead4 internalization--during embryonic development drives cell lineage fate determination. By testing the significance of mitochondrial subtype heterogeneity and inter-organelle communication to TE versus ICM specification, this project will assess if mitochondrial heterogeneity guides cell fate determination, perhaps through an as-yet undiscovered role for mitochondria as transcription factor ‘shuttles’ during asymmetric cell divisions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.