Spindle orientation along the developmental axes in echinoderm embryos

描述（由申请人提供）：动物发育的一个共同特征是受精后，胚胎经历一种物种特异性的、典型的分裂模式，将发育决定因素隔离到特定的子细胞中。虽然谱系追踪和细胞消融研究已经确定了这些发育不对称细胞分裂的重要性，但对有丝分裂纺锤体相对于发育轴排列的机制的研究在很大程度上仅限于少数模式生物。海胆经历放射状的卵裂，每一个卵裂面都与前一个卵裂面平分。在第四次分裂结束时，与植物皮质相关的因子被不对称的细胞分裂成一个细胞群，而这个细胞群反过来又指导内膜的形成。尽管对微粒在海胆中的发育作用和其他系统中的不对称细胞分裂进行了深入的研究，但我们对棘皮动物发育过程中促进纺锤体相对于动物-植物轴定位的机制知之甚少。在迄今为止所研究的几乎所有细胞类型中，有丝分裂纺锤体都遵循Hertwig法则，并沿着细胞的长轴排列，这是由微管马达（动力蛋白）产生的皮质拉力促进的。本研究旨在了解在海胆胚胎初始分裂过程中，有丝分裂纺锤体相对于发育轴的定位。我们假设，在有丝分裂开始之前，早期卵裂球中决定细胞极性的因素使细胞核及其复制的中心体排列，从而影响对称和非对称细胞分裂过程中的纺锤体方向。为了验证这一假设，我们将对海胆和海星胚胎进行一系列活细胞分析，它们的互补特征将有助于定义我们认为是早期胚胎纺锤体排列高度保守的机制。形成本提案具体目标的实验线将：1)确定PAR复合体在卵裂早期确定卵裂球极性和纺锤体方向中的作用；2)评价极性因素和散乱在微粒形成过程中的作用。总之，预计这些研究将导致对极性因素如何影响高度刻板和精确的细胞分裂模式的机制理解，这将最终导致轴的确定和棘皮动物胚胎胚层的规范。

DESCRIPTION (provided by applicant): A common feature of animal development is that following fertilization, embryos undergo a species-specific, stereotypical pattern of cleavages that sequester developmental determinants into specific daughter cells. And while lineage tracing and cell ablation studies have firmly established the significance of these developmentally asymmetric cell divisions, study of the mechanisms that align the mitotic spindle relative to the developmental axes has been largely limited to a few model organisms. Sea urchins undergo radial cleavage, with each cleavage plane bisecting the plane of the previous division. By the end of the fourth division, factors associated with the vegetal cortex ar sequestered by asymmetric cell division into a population of cells that will in turn direct the formation of the endomesoderm. And despite the intense scrutiny placed on the developmental role of micromeres in sea urchins and asymmetric cell division in other systems, we understand little of the mechanisms that facilitate spindle positioning relative to the animal-vegetal axis during echinoderm development. In almost all cell types studied to date, the mitotic spindle obeys Hertwig's rule and aligns itself along the long axis of the cell, facilitated by cortical puling forces generated by the microtubule motor, dynein. This proposal seeks to understand how the mitotic spindle is positioned relative to developmental axes during the initial cleavages of the sea urchin embryo. We hypothesize that factors that define cell polarity in early blastomeres align the nucleus and its duplicated centrosomes prior to the onset of mitosis to influence spindle orientation during both symmetric and asymmetric cell division. To test this hypothesis, we will perform a series of live cell analyses in sea urchin and starfish embryos, whose complementary features will aid in defining what we believe is a highly conserved mechanism for spindle alignment in early embryos. The lines of experimentation that form the Specific Aims of this proposal will: 1) Define the role of the PAR complex in defining blastomere polarity and spindle orientation during early cleavage; and 2) Assess the role of polarity factors and Disheveled during micromere formation. Together, is anticipated that the these studies will lead to a mechanistic understanding of how polarity factors act to effect a highly stereotypical and precise pattern of cell divisions that will ultimately lead to axis determination and the specification of the germ layers on echinoderm embryos.