Molecular Dissection of the "Pacman-Flux" Machinery Used to Move Chromosomes

用于移动染色体的“Pacman-Flux”机器的分子解剖

• 批准号: 8000101
• 负责人: DAVID James SHARP
• 金额: $ 10.05万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-14 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8000101
• 关键词: Anaphase; Biochemical; Cell division; Cells; Centrosome; Chromatids; Chromosome Segregation; Chromosomes; Complement; Congenital Abnormality; Defect; Disease; Dissection; Drosophila genus; Drosophila melanogaster; Embryo; Enzymes; Etiology; Goals; Homologous Gene; Human; Imagery; Kinesin; Kinetochores; Lead; Life; Malignant Neoplasms; Microscopic; Microtubule Depolymerization; Microtubules; Minus End of the Microtubule; Mitosis; Mitotic spindle; Modeling; Molecular; Motion; Motor; Pathway interactions; Phosphorylation; Plus End of the Microtubule; Process; Protein Array; Proteins; Recruitment Activity; Regulation; Sister Chromatid; System; Techniques; Testing; Therapeutic; Tubulin; Work; base; cell motility; depolymerization; embryo cell; insight; katanin; polymerization; protein function; spastin; tissue/cell culture; tumorigenesis

DESCRIPTION (provided by applicant): The ability of chromosomes to move towards opposite poles of the mitotic spindle is fundamental for proper cell division and defects in this process are believed to be an initial step in tumorigenesis. Poleward chromosome motility occurs by a "Pacman-flux" mechanism: Chromosomes induce the depolymerization of attached microtubule plus-ends, termed "Pacman", while being reeled into spindle poles by poleward tubulin flux" stimulated by depolymerization of microtubule minus-ends. The goal of the studies outlined in this proposal is to elucidate the molecular machinery that stimulates and controls "Pacman-flux"-based chromosome motility. The central hypothesis of this work is that chromosome motility is controlled by a mechanistically diverse array of proteins which employ distinct targeting and mechanisms-of-action to control the polymerization state of microtubule ends. There are two specific aims: Aim 1) Elucidate the molecular pathway that stimulates and controls the velocity of microtubule minus-end depolymerization and poleward flux. Studies in this aim evaluate the flux-related functions of microtubule severing proteins, multiple kinesin-13s and kinesin-13 phosphorylation. Aim 2) Elucidate the molecular pathway by which chromosomes induce the depolymerization of microtubule plus-ends. Studies in this aim evaluate the Pacman-related functions of CLIP-170/190, microtubule severing proteins, and multiple kinesin-13s. The fruit fly Drosophila melanogaster is the primary experimental system used in these studies. Live cell techniques for visualizing spindle and chromosome dynamics have been optimized in Drosophila S2 cells and embryos and thus these cells provide an ideal context within which to study how the manipulation of protein function impacts "Pacman-Flux". Live cell microscopic studies of mitosis will be complemented by biochemical and molecular approaches to provide an in-depth understanding of whether and how specific classes of proteins drive chromosome segregation. Additional studies are performed in human cells to determine whether aspects of our proposed pathways are evolutionary conserved. Defects in chromosome segregation lead to human maladies such as birth defects and cancer. An understanding of how this process occurs normally should provide insights into the molecular etiology of these diseases and suggest therapeutic strategies for their treatment.

描述（由申请人提供）：染色体向有丝分裂纺锤体相反两极移动的能力是正确细胞分裂的基础，并且该过程中的缺陷被认为是肿瘤发生的第一步。向极方向的染色体运动是通过“Pacman-flux”机制发生的：染色体诱导附着的微管正端解聚，称为“Pacman”，同时通过微管负端解聚刺激的向极向微管蛋白通量“卷入纺锤体极。本提案中概述的研究的目的是阐明分子机制 刺激和控制基于“Pacman-flux”的染色体运动。这项工作的中心假设是，染色体运动是由机械上多样化的蛋白质阵列控制的，这些蛋白质采用不同的靶向和作用机制来控制微管末端的聚合状态。有两个具体目标： 目标 1) 阐明刺激和控制速度的分子途径 微管负端解聚和极向通量。该目标的研究评估了微管切断蛋白、多种驱动蛋白 13 和驱动蛋白 13 磷酸化的通量相关功能。目标 2) 阐明染色体诱导微管正端解聚的分子途径。该目的的研究评估了 Pacman 相关功能 CLIP-170/190、微管切断蛋白和多个驱动蛋白-13。果蝇 Drosophila melanogaster 是这些研究中使用的主要实验系统。用于可视化纺锤体和染色体动力学的活细胞技术已在果蝇 S2 细胞和胚胎中得到优化，因此这些细胞为研究蛋白质功能的操纵如何影响“Pacman-Flux”提供了理想的环境。直播 有丝分裂的细胞显微镜研究将得到生化和分子方法的补充，以深入了解特定类别的蛋白质是否以及如何驱动染色体分离。在人类细胞中进行了其他研究，以确定我们提出的途径的各个方面是否是进化保守的。染色体分离缺陷会导致人类疾病，例如出生缺陷和癌症。了解此过程如何正常发生 应该提供对这些疾病的分子病因学的见解，并提出治疗策略。

项目成果

Drosophila katanin is a microtubule depolymerase that regulates cortical-microtubule plus-end interactions and cell migration.

• DOI: 10.1038/ncb2206
• 发表时间: 2011-04
• 期刊: Nature cell biology
• 影响因子: 21.3
• 作者: Zhang D;Grode KD;Stewman SF;Diaz-Valencia JD;Liebling E;Rath U;Riera T;Currie JD;Buster DW;Asenjo AB;Sosa HJ;Ross JL;Ma A;Rogers SL;Sharp DJ
• 通讯作者: Sharp DJ

Motor domain phosphorylation and regulation of the Drosophila kinesin 13, KLP10A.

• DOI: 10.1083/jcb.200902113
• 发表时间: 2009-08-24
• 期刊: The Journal of cell biology
• 作者: Mennella V;Tan DY;Buster DW;Asenjo AB;Rath U;Ma A;Sosa HJ;Sharp DJ
• 通讯作者: Sharp DJ

Three microtubule severing enzymes contribute to the "Pacman-flux" machinery that moves chromosomes.

• DOI: 10.1083/jcb.200612011
• 发表时间: 2007-04-23
• 期刊: The Journal of cell biology
• 作者: Zhang D;Rogers GC;Buster DW;Sharp DJ
• 通讯作者: Sharp DJ