The Biochemical Basis for the Mechanics of Cytokinesis

细胞分裂机制的生化基础

基本信息

批准号：
8000107
负责人：
DOUGLAS N ROBINSON
金额：
$ 9.97万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-01-14 至 2010-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8000107
关键词：
Actins Affect Aneuploidy Antineoplastic Agents Behavior Biochemical Cell Adhesion Cell Cycle Cell Shape Cell division Cell physiology Cells Computer Architectures Crosslinker Cytokinesis Cytoskeletal Modeling Data Daughter Dependency Disease Dissection Drosophila polo protein Drug Delivery Systems Event Evolution Failure Genes Genetic Goals Grant Human Image Lead Life Link Malignant Neoplasms Measures Mechanical Stress Mechanics Mediating Microtubules Modeling Molecular Mothers Myosin Type II Nocodazole Normal Cell Pathway interactions Plasmids Process Property Proteins RNA-Binding Proteins Race Regulatory Pathway Shapes Signal Transduction Source Specificity Stress Structure System Testing Tetraploidy Tumor Suppressor Genes Work aurora kinase base cancer cell cancer therapy cell growth chemical genetics clinical application crosslink daughter cell gene function in vivo inhibitor/antagonist inner centromere protein interest loss of function mutant novel particle physical property public health relevance reconstitution response small molecule tumor

项目摘要

DESCRIPTION (provided by applicant): Cytokinesis, the separation of a mother cell into two daughters, is an essential life process. Cytokinesis failure leads to tetraploidy then aneuploidy, an early event in tumor formation. We have been striving to understand how cells use proteins to generate the relevant cellular physical properties that drive cytokinesis contractility. We are also interested in developing small molecule inhibitors to aid in gene function identification and pathway dissection, but with the ultimate goal that some of these small molecule inhibitors will have clinical applications. In this proposal, we will build on the analytical framework that we initiated in the first cycle of the grant, but we will also expand our effort in several ways. In Aim 1, we will measure the lifetimes of myosin-II and various actin crosslinkers in different mutant backgrounds where the mechanics are known in order to assess the consequences of mechanical strain on crosslinker lifetimes. We will test our understanding of the molecular control of cytokinesis mechanics and dynamics by measuring cortical mechanics of dividing mutant strains where we have specific predictions of the mechanics. We will also begin studying lower hierarchical levels of cytoskeletal function by reconstituting crosslinked actin networks, applying mechanical strain to them, and studying the behavior of the crosslinkers and the network. We will use either FRAP or single particle analysis to assess the strain dependency of crosslinker lifetimes; we have several predictions based on our in vivo studies. In Aim 2, we will draw upon our observations that RacE is responsible for generating resistive stresses during cytokinesis and for restricting the mechanosensory system that we discovered to cytokinesis. In this Aim, we will identify RacE effectors to flesh out this pathway. We will study 14-3-3, which was identified as a suppressor of nocodazole. Genetic interactions between RacE and 14-3-3 further point towards a pathway in which microtubules regulate RacE and/or 14-3-3, which in turn regulate global actin crosslinkers to control the dynamics and mechanics of cytokinesis contractility. In Aim 3, we will expand our molecular inquiry by identifying the affected genes in the REMI mutants we have already recovered. PUBLIC HEALTH RELEVANCE: Of great importance for normal cell growth and disease processes such as cancer, cytokinesis has the promise of providing a rich source of new anti-cancer drug targets. We are striving to understand how cytokinesis works at a fundamental level and how the cell uses proteins to generate the physical features of contractility. Ultimately, with a rigorous understanding and a complete molecular handle on the process, it should be possible to develop better cancer therapies that are tailored to the properties of specific types of cancer cells.

描述（由申请人提供）：细胞分裂，即母细胞分裂成两个子细胞，是一个重要的生命过程。细胞分裂失败导致四倍体，然后导致非整倍体，这是肿瘤形成的早期事件。我们一直在努力了解细胞如何利用蛋白质产生驱动胞质分裂收缩性的相关细胞物理特性。我们也有兴趣开发小分子抑制剂来帮助基因功能识别和通路剖析，但最终目标是其中一些小分子抑制剂将具有临床应用。在这项提案中，我们将建立在我们在第一个赠款周期中启动的分析框架的基础上，但我们也将以多种方式扩大我们的努力。在目标 1 中，我们将在已知力学的不同突变体背景下测量肌球蛋白-II 和各种肌动蛋白交联剂的寿命，以评估机械应变对交联剂寿命的影响。我们将通过测量分裂突变株的皮质力学来测试我们对胞质分裂力学和动力学的分子控制的理解，其中我们对力学有具体的预测。我们还将开始通过重建交联肌动蛋白网络、对其施加机械应变以及研究交联剂和网络的行为来研究细胞骨架功能的较低层次。我们将使用 FRAP 或单粒子分析来评估交联剂寿命的应变依赖性；根据我们的体内研究，我们有一些预测。在目标 2 中，我们将利用我们的观察结果，即 RacE 负责在胞质分裂期间产生抵抗应力，并限制我们发现的胞质分裂的机械感觉系统。在这个目标中，我们将鉴定 RacE 效应子来充实这一途径。我们将研究 14-3-3，它被确定为诺考达唑的抑制剂。 RacE 和 14-3-3 之间的遗传相互作用进一步指向微管调节 RacE 和/或 14-3-3 的途径，进而调节全局肌动蛋白交联剂以控制胞质分裂收缩性的动力学和机制。在目标 3 中，我们将通过识别我们已经恢复的 REMI 突变体中受影响的基因来扩展我们的分子探究。公共健康相关性：胞质分裂对于正常细胞生长和癌症等疾病过程非常重要，有望提供丰富的新抗癌药物靶点来源。我们正在努力了解胞质分裂如何在基本水平上发挥作用以及细胞如何利用蛋白质产生收缩性的物理特征。最终，通过对这一过程的严格理解和完整的分子处理，应该有可能开发出更好的针对特定类型癌细胞特性的癌症疗法。