TRACK Regulates Cytokinesis in Trypanosoma brucei

TRACK 调节布氏锥虫的细胞分裂

• 批准号: 6844717
• 负责人: Lawrence Ruben
• 金额: $ 32.47万
• 依托单位: SOUTHERN METHODIST UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-02-01 至 2009-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6844717
• 关键词: A kinase anchoring protein; RNA interference; Trypanosoma brucei; affinity chromatography; biological signal transduction; cell cycle; cell growth regulation; cytoskeletal proteins; flow cytometry; genetic library; genetic screening; laboratory mouse; laboratory rat; nucleic acid sequence; phenotype; phosphorylation; protein binding; protein quantitation /detection; protein signal sequence; protein structure function; site directed mutagenesis; two dimensional gel electrophoresis; yeast two hybrid system

DESCRIPTION (provided by applicant): Cytokinesis is the mechanical process by which a cell divides in two. The process is essential for propagation of unicellular parasites. Little is known about cytokinesis in African trypanosomes, but current information suggests that the process is different from that of the mammalian host. My laboratory has a strong interest in cytokinesis because it is clearly tied to signaling events. In particular, it utilizes temporal information to be triggered only at a specific time in the division cycle; and spatial information to form a cleavage furrow only in the appropriate site. One way to provide spatial and temporal information to the signal process is by means of signal anchor proteins. We have recently identified a putative signal anchor protein called TRACK with strong similarity to mammalian and yeast RACK1/Cpc2. We report that TRACK can correct morphological and growth related defects in cpc2 null mutants of S. pombe. In T. brucei, knock-down of TRACK with RNAi demonstrates that it is essential for progression through a mid-stage of cytokinesis. Cells that become "stuck" in cytokinesis continue through multiple rounds of partial division, and thus accumulate multiple nuclei, and multiple cytoplasmic extensions, each with attached flagellum. The observations are significant since they are the first to demonstrate that once cytokinesis is initiated, it is discontinuous and uses a different set of proteins at mid-stage than at the outset. Since no other RACK1/Cpc2 homologue has been implicated in cytokinesis, these data also underscore the unique aspects of the process in T. brucei. The disruption of TRACK blocks cell division, and indicates that novel components within the cytokinesis pathway may be useful as targets for therapy design. The current project utilizes TRACK as a tool to specifically probe molecular events at the mid-stage of cytokinesis. The project has three specific aims. The first aim examines the role of TRACK in cytokinesis through an examination of its binding partners. The second aim investigates proteins downstream of TRACK whose phosphorylation state changes in TRACKi knock-down cells. The third aim is to screen RNAi libraries for the unique phenotypic changes that accompany defective cytokinesis. In this way TRACK binding partners and other components of this essential process will be identified.

描述（由申请人提供）：细胞因子是细胞分为两部分的机械过程。该过程对于单细胞寄生虫的传播至关重要。关于非洲锥虫中细胞因子的知之甚少，但目前的信息表明，该过程与哺乳动物宿主的过程不同。我的实验室对细胞因子有浓厚的兴趣，因为它显然与信号事件有关。特别是，它利用时间信息仅在分区周期中的特定时间触发；和空间信息仅在适当的位点形成乳沟。向信号过程提供空间和时间信息的一种方法是信号锚蛋白。我们最近确定了一种称为轨道的推定信号锚固蛋白，与哺乳动物和酵母架1/cpc2具有很强的相似性。我们报告说，轨道可以纠正S. pombe的CPC2无效突变体中与形态和生长相关的缺陷。在T. Brucei中，用RNAi敲门表明，这对于通过细胞因子的中期进展至关重要。在细胞因子中“卡住”的细胞继续通过部分分裂的多个回合，从而积累了多个核和多个细胞质延伸，每个延伸均带有附着的鞭毛。这些观察结果很重要，因为它们是第一个证明一旦启动细胞因子，它是不连续的，并且在中段时就使用了一组不同的蛋白质。由于没有其他RACK1/CPC2同源物与细胞因子有关，因此这些数据也强调了T. Brucei过程的独特方面。轨道阻滞细胞分裂的破坏，并表明细胞因子途径中的新成分可能是治疗设计的目标。当前的项目利用轨道作为一种工具，可以在细胞动物的中期特别探测分子事件。该项目具有三个特定的目标。第一个目的通过检查其结合伙伴来检查轨道在细胞因子中的作用。第二个目标调查了轨道下游的蛋白质，其磷酸化状态在跟踪敲低细胞中的变化。第三个目的是筛选RNAi库，以伴随有缺陷的细胞因子的独特表型变化。以这种方式，将确定该基本过程的绑定伙伴和其他组件。