Actin-based mechanisms of chromosome segregation in mammalian oocytes and embryos

项目总结 染色体分离是由一个纺锤体机械驱动的，它分发 子代细胞之间的基因组。在卵子及其祖先卵母细胞中，染色体被分离成 专门的减数分裂程序。卵母细胞和卵子非常容易受到染色体的影响。 导致胚胎非整倍体的分离错误，这是导致自发流产的主要原因 和发育障碍。尽管如此，胚胎非整倍体即使在无错误的完成之后也会出现 减数分裂染色体分离。然而，我们对胚胎特有潜在原因的理解 非整倍体受到一种倾向的限制，即只关注减数分裂衍生的非整倍体。 直到最近，人们还认为微管是细胞骨架的唯一组成部分。 染色体分离所必需的。这一观点被我们发现的纺锤体成功地挑战了 F-肌动蛋白在卵母细胞和卵子中促进染色体纺锤体附着并防止非整倍体。你好吗？ 纺锤体F-肌动蛋白的组装以及它如何在染色体-微管界面发挥作用？ 这些都是我们理解细胞分裂的范式转变所提出的突出问题。 我的实验室的一个主要目标是了解保障准确 哺乳动物卵母细胞和胚胎中的染色体分离。我们发现纺锤体F-肌动蛋白 构成了这样一种保护机制，我们将先进的显微镜分析与 快速蛋白质降解工具确定纺锤体F-肌动蛋白组装和功能所需的蛋白质 卵母细胞。这一方法揭示了支配卵母细胞的关键肌动蛋白和微管结合蛋白。 染色体分离，其中一些与零星流产和 不孕症患者近期遗传学研究中的发育障碍。我们建议在此基础上再接再厉 胚胎特异性非整倍体的研究进展1)扩大基于候选基因的快速 蛋白质降解筛选到更大的肌动蛋白-微管串扰蛋白子集，以及2)发展 一种新的生化和蛋白质组学耦合的无偏鉴定新基因的实验流水线 卵母细胞和胚胎中的纺锤体F-肌动蛋白组装蛋白。此外，我们还将直接进行实验 应用显微注射和激光显微手术工具增加或移除中心体的方法 解释为什么纺锤形的F-肌动蛋白结构是无着丝体纺锤体所特有的。小鼠早期胚胎 有丝分裂是在没有规范的中心体的情况下进行的，它将为我们提供一个有吸引力的 用来回答细胞生物学中这个长期存在的问题的实验模型。 当这项研究完成后，我们将发现并确定功能特征 卵母细胞和胚胎中的纺锤体F-肌动蛋白组装蛋白。总体而言，这项研究将揭示出 细胞骨架系统相互协作，在早期发育阶段驱动准确的染色体分离。

PROJECT SUMMARY Chromosome segregation is driven by a spindle machinery that distributes copies of the genome between daughter cells. In eggs and their progenitor oocytes, chromosomes are segregated in a specialized meiotic division program. Oocytes and eggs are remarkably vulnerable to chromosome segregation errors that give rise to aneuploidy in embryos, a leading cause of spontaneous miscarriages and developmental disorders. Embryo aneuploidy can nonetheless arise even after error-free completion of meiotic chromosome segregation. However, our understanding of the underlying causes of embryo-specific aneuploidies has been restricted by a tendency to focus only on meiosis-derived aneuploidies. Until recently, it was believed that microtubules are the only cytoskeletal components required for chromosome segregation. This view was successfully challenged by our discovery of spindle F-actin in oocytes and eggs that boost chromosome-spindle attachments and prevent aneuploidy. How is spindle F-actin assembled and how does it exert its function at the chromosome-microtubule interface? These are among outstanding questions raised by this paradigm shift in our understanding of cell division. A major goal of my lab is to understand the mechanisms that safeguard accurate chromosome segregation in mammalian oocytes and embryos. Driven by our discovery that spindle F-actin constitutes one such protection mechanism, we are combining advanced microscopy assays with rapid protein degradation tools to identify proteins required for spindle F-actin assembly and function in oocytes. This approach has revealed key actin- and microtubule-binding proteins that govern oocyte chromosome segregation, some of which were independently implicated in sporadic miscarriages and developmental disorders in recent genetic studies of infertility patients. We propose to build on this progress and study the origins of embryo-specific aneuploidy by 1) expanding our candidate-based rapid protein degradation screens to a larger subset of actin-microtubule crosstalk proteins, and 2) developing a new biochemical and proteomics-coupled experimental pipeline for unbiased identification of novel spindle F-actin assembly proteins in oocytes and embryos. Furthermore, we will take direct experimental approaches of adding or removing centrosomes using microinjection and laser microsurgery tools to address why spindle-shaped F-actin structures are unique to acentrosomal spindles. Early mouse embryo mitotic divisions, which are executed without canonical centrosomes, will provide us with an attractive experimental model in which to answer this long-standing question in cell biology. When this research is completed, we will have discovered and functionally characterized spindle F-actin assembly proteins in oocytes and embryos. Overall, this study will reveal how distinct cytoskeletal systems cooperate to drive accurate chromosome segregation in early development.