Mechanisms of asymmetric cell division during female meiosis

描述(申请人提供)：我们的目标是了解雌性减数分裂纺锤体在卵子皮质的位置和调节极体形成的机制。从长远来看，我们希望阐明为什么不对称减数分裂纺锤体位置在动物门中是如此保守的原因。这项建议的具体目标是阐明三种不同途径的详细机制，这些途径调节秀丽线虫减数分裂纺锤体定位和极体形成。1)在减数分裂中期-后期转换之前，需要动蛋白-1重链、动蛋白轻链和一种名为KCA-1的新蛋白将减数分裂纺锤体移动到卵皮层。相反，减数分裂纺锤体在中期-后期转换后向卵皮层的运动是不依赖动蛋白的。我们将使用荧光显微镜和体外生物化学来检验这一假说，即Kinesin/KCA-1复合体直接沿着细胞质微管运输纺锤体，并且在野生型减数分裂过程中，一个不同的机制导致了纺锤体的移位和纺锤体的旋转。2)微管切断蛋白katanin的完全丧失导致不能组装减数分裂纺锤体，而katanin功能的部分丧失则导致减数分裂纺锤体异常长和极体异常大。我们将使用荧光显微镜和遗传学来验证微管切断活动限制减数分裂纺锤体长度从而限制极体大小的假设。3)SPE-11是一种精子蛋白，在受精时进入卵子，是极体形成所必需的。我们将使用荧光显微镜和遗传学相结合的方法来验证SPE-11通过与卵子中保守的胞质分裂调节因子相互作用来调节极体形成的假设。减数分裂纺锤体与公共卫生有关，主要有两个原因。首先，减数分裂纺锤体功能异常导致非整倍体，导致唐氏综合征和流产。其次，目前利用患者匹配的干细胞治疗多种人类疾病的潜力受到动物克隆过程中异常的减数分裂纺锤体组装的限制。因此，对减数分裂纺锤体功能的详细分子理解可能会导致预防出生缺陷和允许干细胞治疗人类疾病的治疗方法。

DESCRIPTION (provided by applicant): Our goal is to understand the mechanisms that position female meiotic spindles at the egg cortex and that mediate polar body formation. In the long term, we hope to elucidate the reasons why asymmetric meiotic spindle positioning is so conserved across animal phyla. The specific goals of this proposal are to elucidate the detailed mechanisms of three distinct pathways that mediate meiotic spindle positioning and polar body formation in Caenorhabitis elegans. 1) Kinesin-1 heavy chain, kinesin light chains and a novel protein called KCA-1 are each required to move the meiotic spindle to the egg cortex before the metaphase-anaphase transition. In contrast, movement of the meiotic spindle to the egg cortex after the metaphase-anaphase transition is kinesin-independent. We will use fluorescence microscopy and in vitro biochemistry to test the hypothesis that a kinesin/KCA-1 complex directly transports the spindle along cytoplasmic microtubules and that a distinct mechanism is responsible for both spindle translocation in the absence of kinesin and spindle rotation during wild-type meiosis. 2) Complete loss of the microtubule-severing protein, katanin, results in failure to assemble a meiotic spindle whereas partial loss of katanin function results in abnormally long meiotic spindles and abnormally large polar bodies. We will use fluorescence microscopy and genetics to test the hypothesis that microtubule-severing activity restricts meiotic spindle length and thereby polar body size. 3) SPE-11 is a sperm protein introduced into the egg at fertilization and which is required for polar body formation. We will use a combination of fluorescence microscopy and genetics to test the hypothesis that SPE-11 mediates polar body formation by interacting with conserved cytokinesis regulators in the egg. Meiotic spindles are relevant to public health for two major reasons. First, abnormal meiotic spindle function leads to aneuploidy that causes Down syndrome and miscarriage. Second, the potential for treating numerous human diseases with patient-matched stem cells is currently limited by abnormal meiotic spindle assembly during animal cloning. A detailed molecular understanding of meiotic spindle function could thus lead to treatments that prevent birth defects and that allow stem cell therapy of human diseases.