MOLECULAR ANALYSIS OF CYTOKINESIS IN DICTYOSTELIUM

盘基网柄菌细胞分裂的分子分析

• 批准号: 6841677
• 负责人: ARTURO DE LOZANNE
• 金额: $ 33.22万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-01-01 至 2006-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6841677
• 关键词: MOLECULAR; ANALYSIS; CYTOKINESIS; DICTYOSTELIUM

EXCEEDTHESPACE PROVIDED. Despite recent advances, cytokinesis remains the least understood aspect of the cell cycle. In animal cells, cytokinesis requires the coordination of multiple cellular components including the microtubule cytoskeleton, the actin cytoskeleton and membrane traffic. Today, it is not clear how these components are regulated and how they interact with each other. The long-term goal of this project is to define the elements involved in cytokinesis and to understand how these elements integrate to accomplish cytokinesis. Toward that goal, two proteins required for cytokinesis in Dictyostelium have been identified. These proteins provide a handle on two different pathways that come together during cytokinesis. The objective of this grant is to dissect the contribution of these pathways during cytokinesis: 1) The role of LvsA in osmorequlation and cytokinesis will be defined. LvsA is a member of the novel beach family of membrane trafficking proteins and is required for cytokinesis and for the function of the contractile vacuole. This provides the opportunity to dissect the contribution of membrane traffic to cytokinesis in a simple model system. The functional contribution of each of the LvsA domains to its localization and function in vivo will be ascertained. Binding- partners for LvsA will be identified and their requirement for cytokinesis will be tested. Genetic suppressors of LvsA will be isolated and the mechanism of suppression will be determined. These studies will illuminate our understanding of membrane processes in cytokinesis and also how mutations in a human homologue of LvsA (LYST) cause a human disorder, the Chediak-Higashi Syndrome. 2) The signaling pathway that requires RacE durinq cytokinesis will be defined. The small GTPase RacE is essential for the development of cortical tension and for the ingression of the cleavage furrow during cytokinesis. A combination of approaches will be used to dissect the mechanisms by which racE controls cytokinesis. A model that explains the role of RacE during cytokinesis is proposed based on recent data from several laboratories on proteins required in cytokinesis. Several predictions made by this model will be tested, among them: a) that RacE controls the organization of Cortexillins at the cleavage furrow though the activation of specific IQGAPs; b) that the GAP domain of DdRacGAP1 modulates the activity of RacE in vivo. In addition, a method to purify RacE has been developed and will be used to identify novel binding partners for racE. The role of these proteins in cytokinesis will be determined by biochemical and genetic approaches. PERFORMANCESITE( ========================================Section End===========================================

超出所提供的空间。尽管最近取得了进展，胞质分裂仍然是细胞周期中人们最不了解的方面。在动物细胞中，胞质分裂需要多种细胞组分的协调，包括微管细胞骨架、肌动蛋白细胞骨架和膜运输。目前尚不清楚这些成分是如何调节的，以及它们如何相互作用。这个项目的长期目标是确定参与胞质分裂的元素，并了解这些元素如何整合完成胞质分裂。为了实现这一目标，已经鉴定了网骨藻中胞质分裂所需的两种蛋白质。这些蛋白质为胞质分裂期间聚集在一起的两种不同途径提供了一个手柄。本基金的目的是剖析这些途径在胞质分裂过程中的作用：1）LvsA在细胞周期调节和胞质分裂中的作用将被定义。LvsA是膜运输蛋白的新型海滩家族的成员，并且是胞质分裂和收缩泡的功能所需的。这提供了在一个简单的模型系统中剖析膜交通对胞质分裂的贡献的机会。将确定每个LvsA结构域对其体内定位和功能的功能贡献。将鉴定LvsA的结合配偶体，并检测其对胞质分裂的要求。将分离LvsA的遗传抑制因子，并确定抑制机制。这些研究将阐明我们对胞质分裂中膜过程的理解，以及LvsA（LYST）的人类同源物突变如何导致人类疾病Chediak-Higashi综合征。2)将定义胞质分裂期间需要RacE的信号传导途径。小的GTTRase RacE是必不可少的皮质张力的发展和细胞质分裂过程中的卵裂沟的侵入。一个组合的方法将被用来解剖的机制，其中racE控制胞质分裂。根据最近几个实验室对胞质分裂所需蛋白质的数据，提出了一个解释胞质分裂过程中RacE作用的模型。将测试由该模型做出的几个预测，其中包括：a）RacE通过特定IQGAP的激活控制皮质素在切割沟处的组织; B）DdRacGAP 1差距域调节RacE的体内活性。此外，已经开发了一种纯化RacE的方法，并将用于鉴定RacE的新型结合伴侣。这些蛋白质在胞质分裂中的作用将通过生物化学和遗传学方法来确定。表演现场（=