Revealing how the mitotic spindle controls asymmetric cell division in vivo

摘要 这项提议的目标是发现新形式的微管组织如何驱动第一个不对称细胞 活体内哺乳动物有机体的分裂。多细胞生物最显著的特征之一是它们的 大量的细胞类型。其中许多是由不对称细胞分裂产生的，在不对称分裂中，亲本细胞 不对称地分配细胞命运决定因素，或产生不同形状、体积或细胞的子代细胞 极性特征。 在大多数细胞类型中，有丝分裂的纺锤体在细胞不对称分裂中起着基础性的作用。它包括 有两个关键部分，即与染色体相互作用并促进细胞动态沟形成的中心纺锤体， 以及一排星体微管。这些星形微管起源于纺锤体两极的中心体。 并允许极化的皮质元素并迫使调节器以不同的方式控制主轴以驱动不对称 细胞分裂。在培养细胞和非培养细胞中，细胞不对称分裂的机制已经得到了详细的研究。 哺乳动物的有机体。然而，小鼠胚胎并不继承其亲代细胞的中心粒，而是 建议缺乏功能中心体和星体微管阵列。因此，目前尚不清楚有丝分裂如何 磁盘轴是在发育的最早阶段组织起来的，以及它们可能如何导致不对称 细胞分裂是驱动哺乳动物细胞分化的第一类细胞。 在这里，我们结合了活体成像方法与分子和生物物理方法来测试一种新的 有丝分裂纺锤体组织的形式，我们在体内发现了驱动细胞不对称分裂的形式。与之前的不同 假设缺乏中心体和星体微管阵列，我们发现一些细胞建立了高度的 不对称纺锤体，仅有一个星形微管阵列。对这种不对称心轴的操纵破坏了 第一类细胞类型的分离。因此，这一提议的中心假设是，建立 这种不对称纺锤体组织的新形式推动了体内第一次不对称细胞分裂。为了测试这一点， 我们的目标将研究非对称主轴的装配和驱动机制(目标1) 不对称细胞分裂(目标2)。 目标1将专门侧重于剖析所需的分子调节剂的来源和功能 组装不对称纺锤体，细胞极性组件的作用，以及带来 星体阵列和中心纺锤体一起组装成一个完整的有丝分裂纺锤体。 目的2将研究不对称纺锤体驱动不对称细胞分裂的机制。我们会 具体测试基于细胞分裂定向的差异调节的三个基本机制， 子代细胞体积和细胞皮质张力。

Summary The goal of this proposal is to discover how new forms of microtubule organization drive the first asymmetric cell divisions of a mammalian organism in vivo. One of the most salient features of multicellular organisms is their vast number of cell types. Many of these are produced by asymmetric cell division, in which a parental cell asymmetrically distributes cell fate determinants, or generates daughter cells with different shape, volume or cell polarity features. In most cell types, the mitotic spindle apparatus plays a fundamental role in asymmetric cell division. It comprises two key parts, the central spindle that interacts with chromosomes and promotes cytokinetic furrow formation, and an array of astral microtubules. These astral microtubules originate from centrosomes at the spindle poles and allow polarized cortical elements and force regulators to differentially control the spindle to drive asymmetric cell division. The mechanisms of asymmetric cell division have been studied in detail in cultured cells and non- mammalian organisms. Yet, the mouse embryo does not inherit centrioles form its parental cells and has been proposed to lack functional centrosomes and astral microtubule arrays. Therefore, it remains unclear how mitotic spindles are organized during the earliest stages of development, and how they may contribute to asymmetric cell division to drive the differentiation of the first mammalian cell types. Here, we combine live-imaging approaches with molecular and biophysical methods to test the function of a new form of mitotic spindle organization, which we found to drive asymmetric cell division in vivo. Unlike previous assumptions of lack of centrosomes and astral microtubule arrays, we found that some cells establish a highly asymmetric spindle with only one astral-like microtubule array. Manipulations of this asymmetric spindle disrupt the segregation of the first cell types. Thus, the central hypothesis of this proposal is that the establishment of this new form of asymmetric spindle organization drives the first asymmetric cell divisions in vivo. To test this, our aims will investigate the mechanisms by which the asymmetric spindle is assembled (Aim 1) and can drive asymmetric cell division (Aim 2). Aim 1 will specifically focus on dissecting the source and function of the molecular regulators required to assemble the asymmetric spindle, the role of cell polarity components, and the physical forces required to bring together the astral array and the central spindle to assemble a complete mitotic spindle apparatus. Aim 2 will investigate the mechanisms by which the asymmetric spindle drives asymmetric cell division. We will specifically test three fundamental mechanisms based on the differential regulation of cell cleavage orientation, daughter cell volume and cell cortex tension.