Uncovering how the dynamic architecture of a layered contractile ring induces furrow ingression

揭示分层收缩环的动态结构如何引起沟槽侵入

• 批准号: 10004677
• 负责人: Caroline Laplante
• 金额: $ 30.82万
• 依托单位: NORTH CAROLINA STATE UNIVERSITY RALEIGH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10004677
• 关键词: Abnormal Cell; Actin-Binding Protein; Actins; Actomyosin; Address; Affect; Alleles; Animal Model; Architecture; Biological Models; Biology; Bundling; Cell division; Cell membrane; Cells; Complex; Congenital Abnormality; Contractile Proteins; Cytokinesis; Cytoplasm; Data; Defect; Development; Disease; Event; Fission Yeast; Fluorescence; Foundations; Generations; Genes; Genetic Materials; Human; Immune System Diseases; Individual; Kinetics; Knowledge; Lasers; Lead; Life; Light; Malignant Neoplasms; Measures; Mechanics; Membrane; Methodology; Microfilaments; Microscopy; Microsurgery; Modeling; Molecular; Mothers; Motor; Mutation; Myosin ATPase; Organism; Outcome; Pathogenesis; Plus End of the Actin Filament; Process; Production; Proteins; Publishing; Research; Research Proposals; Role; Speed; Structure; System; Testing; Work; Yeasts; base; cell cortex; confocal imaging; constriction; daughter cell; frontier; fungus; imaging genetics; innovation; molecular dynamics; nanometer resolution; nervous system disorder; novel; photoactivation; public health relevance; transmission process

Project description: 30 lines Cytokinesis, the separation of a mother cell into two daughter cells, is one of life's most fundamental processes; it is central to the making of multicellular organisms and the transmission of genetic material across generations. The core machinery of cytokinesis is a contractile ring of actin, myosin motors and other proteins and is conserved from fungi to human. The contractile ring is connected to the inside of the cell cortex and its constriction leads the ingression of the plasma membrane into a furrow between the two cytoplasmic compartments that will become individual cells after the completion of cell division. Furrow ingression requires the cooperation between two main mechanisms: 1) constriction of actin filaments by the action of myosin motors and 2) the transmission of this contractile force to the plasma membrane via anchor proteins that connect the actomyosin bundle to the plasma membrane. We found that the proteins of the contractile ring occupy distinct layers with plasma membrane-interacting proteins in the outer layer adjacent to the cortex and the force- producing myosin motors in the inner layer. The next frontier in understanding the mechanism of cytokinesis is to determine how these proteins are organized into complex structures, how these structures move within the contractile ring and are removed from the ring during constriction, and how this dynamic architecture governs the force-generation function of the contractile rings. The objective of this application is to determine the anchoring role of the outer ring and the tension-force producing role of the inner ring by uncovering their dynamic architecture and effects on mechanics of constriction. Our hypothesis is that furrow ingression results from contractile forces produced in the inner layer of the contractile ring, conveyed to the plasma membrane via anchoring achieved by proteins in the outer layer. We plan to test our central hypothesis with the following Specific Aims: 1) determine how cytokinetic node proteins anchor the contractile ring and transmit contractile forces to the plasma membrane during furrow ingression and 2) determine how the molecular architecture of the inner ring governs the constriction of the contractile ring during furrow ingression. The proposed research in this application is innovative because we will use a unique combination of high-speed Fluorescence Photoactivation Localization Microscopy (hsFPALM) in live cells to determine protein organization and its dynamics with laser microsurgery to probe the mechanics of this tension-force producing machine in live cells. The proposed research in this application is significant as it will result in the identification of previously unknown parameters for new molecular and functional models of the contractile ring. As the proteins of the contractile ring are conserved from yeast to human, we expect that the core mechanism of cytokinesis elucidated in fission yeast will act as a template for understanding the mechanism of cytokinesis and even perhaps other non-musclecontractile cellular machines in other species.

项目描述：30行 胞质分裂，即将一个母细胞分裂成两个子细胞，是生命最基本的过程之一； 它对多细胞生物体的形成和遗传物质的世代传播至关重要。 胞质分裂的核心机制是由肌动蛋白、肌球蛋白马达和其他蛋白质组成的收缩环，是 从真菌到人类都保存着。收缩环连接到细胞皮质的内侧，其 收缩导致质膜进入两个细胞质之间的沟槽。 细胞分裂完成后将成为单个细胞的隔室。沟渠渗入需要 两个主要机制之间的协作：1)肌球蛋白马达的作用使肌动蛋白微丝收缩 2)这种收缩力量通过连接细胞膜的锚定蛋白传递到质膜。 肌动球蛋白结合到质膜上。我们发现，收缩环上的蛋白质占据了明显的位置 与皮层相邻的外层含有质膜相互作用的蛋白质层，力- 在内层产生肌球蛋白马达。理解细胞质分裂机制的下一个前沿是 为了确定这些蛋白质是如何组织成复杂结构的，这些结构如何在 收缩环，并在收缩过程中从环上移除，以及这种动态架构如何管理 收缩环的力生成函数。此应用程序的目标是确定 揭示外环的锚定作用和内环的张力产生作用 结构和对收缩力学的影响。我们的假设是，沟进是由 在收缩环的内层产生的收缩力通过 由外层的蛋白质实现锚定。我们计划通过以下几点来验证我们的中心假设 具体目标：1)确定细胞运动节点蛋白如何锚定收缩环并传递收缩 在沟进过程中对质膜的作用力和2)决定了质膜的分子结构 内环控制着沟槽进入时收缩环的收缩。建议在这方面进行的研究 应用程序是创新的，因为我们将使用一种独特的组合高速荧光光活化 利用激光在活细胞中定位显微镜(HsFPALM)确定蛋白质组织及其动力学 在活细胞中探索这种张力产生机器的机制的显微外科手术。建议数 对这一应用的研究具有重要意义，因为它将导致识别先前未知的参数 收缩环的新分子和功能模型。因为收缩环的蛋白质是保守的 从酵母到人类，我们期望在分裂酵母中阐明的胞质分裂的核心机制将作为一种 用于了解胞质分裂甚至其他非肌肉收缩细胞机制的模板 其他物种中的机器。