MITOTIC SPINDLES IN NEMATODES: COMPARATIVE BIOPHYSICS TO EVOLUTIONARY BIOLOGY

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 有丝分裂纺锤体在组织和生物体之间可以显示出显著的变异性，但在细胞分裂中， 目前对这种多样性的进化基础知之甚少。因此，目前还不清楚为什么在不同的真核生物中期纺锤体表现出一系列的形态和这些纺锤体的体积变化超过一千倍。在这里，我们提出了一个比较研究，以调查的进化力量，形状的有丝分裂纺锤体。我们将在有丝分裂中使用单细胞线虫胚胎来解决三个广泛的问题： 1)锭子动力学和结构的变化程度如何？我们将通过研究来自线虫门的~60个物种来调查小的和大的同源距离-包括Caushabditis属的所有已知物种。这项工作将使用一些光学显微镜技术来确定线虫纺锤体变异的范围。 2)这种变异的生物物理基础是什么？我们将在10个选择物种的微管动力学和纺锤体结构进行深入分析。荧光技术（在非模型物种中可能时），薄片和3-D电子显微镜，以及物理和遗传（可行时）的扰动实验，将用于测量微管的动态，数量和行为。我们将发展生物物理模型来解释微管行为的差异如何导致纺锤体结构的差异。 3)这种变异的进化基础是什么？我们将使用表型进化的模型来深入了解：（i）系统发育效应的程度;（ii）纺锤体特征对细胞大小，核型，生活史和其他属性的依赖;以及（iii）选择和漂移在塑造纺锤体各种成分中的重要性。一个物种的平行研究，秀丽隐杆线虫，将提供一个更详细的分析进化的力量：我们将使用突变积累线来衡量自发突变如何修改有丝分裂纺锤体，我们将确定这种变异性是如何通过比较这些线与~40种不同的野生菌株的演变。这种独特的比较方法结合了分子方法，定量显微镜和数学建模，开发了一个全面的模型，通过突变，选择，漂移和细胞生物物理学的进化，有丝分裂纺锤体是如何形成的。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Mitotic spindles can show remarkable variability between tissues and organisms, but there is currently little understanding of the evolutionary basis of this diversity. Thus it is unclear why metaphase spindles in different Eukaryotes exhibit a range of morphologies and the volumes of these spindles vary over one thousand fold. Here, we propose a comparative study to investigate the evolutionary forces that shape the mitotic spindle. We will use one-cell nematode embryos in mitosis to address three broad questions: 1) What is the extent of variation in spindle dynamics and structure? We will investigate both small and large phylogenic distances by studying ~60 species from across the nematode phylum - including all known species of the Caenorhabditis genus. This work will use a number of light microscopy techniques to determine the range of spindle variation in nematodes. 2) What is the biophysical basis of this variation? We will perform an in-depth analysis of microtubule dynamics and spindle architecture in ~10 select species. Fluorescence techniques (when possible in nonmodel species), thin-section and 3-D electron microscopy, and perturbations experiments, both physical and genetic (when feasible), will be used to measure the dynamics, number and behavior of microtubules. We will develop biophysical models to explain how differences in microtubule behavior lead to differences in spindle architecture. 3) What is the evolutionary basis of this variation? We will use models of phenotypic evolution to gain insight into: (i) the extent of phylogenetic effects; (ii) the dependence of spindle features on cell size, karyotype, life history, and other attributes; and (iii) the importance of selection and drift in shaping various components of the spindle. A parallel study on one species, Caenorhabditis elegans, will provide a more detailed analysis of evolutionary forces: we will use mutation accumulation lines to measure how spontaneous mutations modify the mitotic spindle, and we will determine how this variability is shaped over evolution by comparing these lines with ~40 different wild isolates. This unique comparative approach combines molecular methods, quantitative microscopy, and mathematical modeling, to develop a comprehensive model of how the mitotic spindle is shaped through evolution by mutation, selection, drift, and cellular biophysics.