Mechanisms of mitosis and size control in Xenopus

非洲爪蟾有丝分裂和大小控制的机制

• 批准号: 10378687
• 负责人: Rebecca W Heald
• 金额: $ 87.01万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-12 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10378687
• 关键词: Address; Affect; Architecture; Area; Automobile Driving; Biochemical; Biological; Biological Assay; Biology; Biophysical Process; Cell Nucleus; Cell Size; Cell division; Cell physiology; Cells; Centromere; Chromosome Condensation; Chromosome Segregation; Chromosomes; Ciona intestinalis; Collaborations; Computer Models; Conflict (Psychology); Cytoplasm; Cytoskeleton; DNA; Defect; Development; Embryo; Embryonic Development; Ensure; Event; Evolution; Gene Expression; Genome; Genomics; Hybrids; Interphase; Kinetochores; Laboratories; Lead; Life; Link; Malignant Neoplasms; Measures; Mediating; Meiosis; Microfluidics; Microscopy; Microtubules; Mitosis; Mitotic; Mitotic Chromosome; Mitotic spindle; Molecular; Morphology; Oocytes; Organelles; Organism; Phylogenetic Analysis; Physiology; Ploidies; Process; Proteomics; Rana; Research; Resolution; Role; Shapes; Structure; Surface; System; Techniques; Testing; Time; Urochordata; Xenopus; base; cell type; chromosome missegregation; daughter cell; egg; embryo cell; human disease; in vivo; innovation; insight; laser tweezer; novel; novel strategies; reproductive; segregation; sensor; single molecule; sperm cell; virtual; xenopus development

PROJECT SUMMARY Mechanisms of Mitosis and Size Control in Xenopus Research in my laboratory is focused on two major areas: Cell division is arguably the most dramatic event in the life of a cell. Chromosomes condense, organelles vesiculate, and the microtubule cytoskeleton rearranges into a bipolar spindle that attaches to chromosomes at their kinetochores and segregates a complete genome to each daughter cell. Although the morphological changes that occur during mitosis were first observed over a century ago, we still do not understand how these dynamic events are orchestrated. Many factors have been identified that contribute to spindle assembly and function, but the molecular and biophysical mechanisms and interactions that ensure mitotic fidelity remain unclear. Our current projects address outstanding questions including 1) What are the molecular underpinnings and functional consequences of different spindle architectures? Spindle size and organization vary dramatically across cell types and organisms, and factors known to affect these parameters are altered in many cancers, but how specific spindle features are established and their effects on chromosome segregation and cell division are poorly understood. We will leverage morphometric and phylogenetic comparisons together with biochemical and functional assays to investigate the dramatic changes in spindle architecture that occur between oocyte meiosis and the mitotic divisions of early development in Xenopus and the sea squirt Ciona intestinalis. We will elucidate the role of specific factors in this transition, and examine the consequences of altering spindle architecture on embryo cell division. 2) What defects in cell division mechanisms underlie speciation? We have observed chromosome mis-segregation in inviable hybrids generated by fertilizing Xenopus tropicalis eggs with X. laevis sperm, and identified incompatibility between a subset of paternal centromeres and maternal cytoplasm as one underlying cause. We will elucidate the molecular basis of inter-species conflicts that impact cell division and contribute to reproductive isolation. 3) What is the molecular basis of mitotic chromosome condensation? We have developed a novel approach using optical tweezers to measure the dynamics of single DNA molecules in real-time in Xenopus egg extracts with high spatial and temporal precision and will use this system to dissect the roles of key factors in driving mitotic chromosome assembly. Absolute and relative size of biological entities varies widely, both within and among species at all levels of organization above the atomic/molecular: the organism, the cells that make up the organism, and the cellular components. How does scaling occur so that everything fits and functions properly? Correct scaling inside cells is crucial for cell function, architecture, and division, but until recently the control systems that a cell uses to regulate the size of its internal structures were virtually unknown. We have established assays to elucidate mechanisms of intracellular scaling between different-sized frog species and during the rapid, reductive cell divisions of early embryogenesis. We are further developing these systems to ask: 1) What scales mitotic chromosome size to cell size? We are testing the hypothesis that a surface area to volume sensor acting on the interphase nucleus and the mitotic spindle also coordinately adjusts mitotic chromosomes to cell size during Xenopus development. 2) What are the connections between genome size, cell size, physiology, and development? Cell size correlates strongly with genome size across evolution, but underlying mechanisms are unknown. We will utilize different ploidy frog embryos to address how altering genome size affects gene expression, and a variety of species including the dodecaploid frog Xenopus longipes to investigate relationships between genome size, cell division mechanisms, development, and physiology. The means to address these fundamental cell biological questions is enabled by powerful experimental systems based on cytoplasmic extracts and functional in vivo assays in vertebrate (Xenopus) embryos. We have established productive collaborations and apply diverse techniques including high-resolution microscopy, single molecule assays, genomics, proteomics, microfluidics and computational modeling to fill important conceptual gaps in an innovative, rigorous, and interdisciplinary manner. Our research will continue to provide novel insight into cell division and size control, processes essential for viability and development, and defective in human diseases including cancer. Although introduced as distinct topics, cell division and size control are intimately linked. We are increasingly focused on how cross-species comparisons can elucidate molecular mechanisms underlying cell division and size control, as well as how biological constraints related to these processes have shaped evolution. Together, these projects uniquely advance our understanding of long-standing questions in biology.

项目概要 爪蟾有丝分裂和大小控制机制 我实验室的研究主要集中在两个主要领域： 细胞分裂可以说是细胞生命中最引人注目的事件。染色体浓缩，细胞器 囊泡状，微管细胞骨架重新排列成双极纺锤体，附着在染色体上 它们的着丝粒并将完整的基因组分离到每个子细胞中。虽然形态上 有丝分裂过程中发生的变化在一个多世纪前就被首次观察到，但我们仍然不明白 这些动态事件是如何编排的。已确定许多因素会影响主轴 组装和功能，但确保有丝分裂的分子和生物物理机制和相互作用 保真度仍不清楚。我们当前的项目解决了突出的问题，包括 1) 什么是分子 不同主轴架构的基础和功能后果？主轴尺寸及机构 不同的细胞类型和生物体差异很大，并且已知影响这些参数的因素在不同的环境中发生改变 许多癌症，但是特定的纺锤体特征是如何建立的及其对染色体分离的影响 和细胞分裂知之甚少。我们将一起利用形态学和系统发育比较 通过生化和功能测定来研究纺锤体结构发生的巨大变化 爪蟾和海鞘的卵母细胞减数分裂和早期发育的有丝分裂之间的关系 肠杆菌。我们将阐明具体因素在这一转变中的作用，并研究其后果 改变胚胎细胞分裂的纺锤体结构。 2）细胞分裂机制的缺陷是什么？ 物种形成？我们观察到非洲爪蟾受精产生的不可存活杂种中存在染色体错误分离 热带丝虫卵与非洲丝虫精子，并鉴定出父本着丝粒子集和 母体细胞质是根本原因之一。我们将阐明物种间冲突的分子基础 影响细胞分裂并有助于生殖隔离。 3）有丝分裂染色体的分子基础是什么 缩合？我们开发了一种使用光镊来测量单个粒子动力学的新颖方法 非洲爪蟾卵提取物中的 DNA 分子具有高空间和时间精度，并将使用此技术 系统来剖析驱动有丝分裂染色体组装的关键因素的作用。 生物实体的绝对和相对大小在各个层次的物种内部和物种之间差异很大。 原子/分子之上的组织：有机体、组成有机体的细胞和细胞 成分。如何进行缩放以使一切都适合并正常运行？内部正确缩放 细胞对于细胞功能、结构和分裂至关重要，但直到最近，细胞的控制系统 细胞用于调节其内部结构大小的功能实际上是未知的。我们已经建立了化验 阐明不同大小的青蛙物种之间以及快速、 早期胚胎发生的还原性细胞分裂。我们正在进一步开发这些系统来询问：1）什么规模 有丝分裂染色体大小与细胞大小的关系？我们正在测试以下假设：表面积与体积传感器的作用 间期核和有丝分裂纺锤体也根据细胞大小协调调整有丝分裂染色体 在爪蟾发育期间。 2) 基因组大小、细胞大小、生理学和 发展？细胞大小与整个进化过程中的基因组大小密切相关，但潜在的机制是 未知。我们将利用不同倍性的青蛙胚胎来解决改变基因组大小如何影响基因 表达，以及包括十二倍体青蛙 Xenopus longipes 在内的多种物种来研究关系 基因组大小、细胞分裂机制、发育和生理学之间的关系。 解决这些基本细胞生物学问题的方法是通过强大的实验系统实现的 基于脊椎动物（非洲爪蟾）胚胎的细胞质提取物和功能性体内测定。我们有 建立了富有成效的合作并应用了多种技术，包括高分辨率显微镜、单 分子分析、基因组学、蛋白质组学、微流体学和计算模型来填补重要的概念 以创新、严谨和跨学科的方式弥补差距。我们的研究将继续提供新颖的见解 细胞分裂和大小控制、生存和发育所必需的过程以及人类的缺陷 疾病，包括癌症。尽管作为不同的主题引入，但细胞分裂和大小控制密切相关 已链接。我们越来越关注跨物种比较如何阐明分子机制 潜在的细胞分裂和大小控制，以及与这些过程相关的生物限制如何影响 形演化。这些项目共同促进了我们对长期存在的问题的理解 生物学。