Septin Organization in Multinucleated Cells

多核细胞中的 Septin 组织

• 批准号: 0719126
• 负责人: Amy Gladfelter
• 金额: $ 66.3万
• 依托单位: Dartmouth College
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0719126&HistoricalAwards=false
• 关键词: Septin; Organization; Multinucleated; Cells

Proteins at the cell cortex link the cell interior and exterior by transmitting and responding to a variety of signals. Thus, the cell cortex is an information processing center where cells can receive and react to diverse stimuli in their environment or within the cell. A conserved family of proteins called septins organizes and regulates structures and processes at the cortex of eukaryotic cells. In addition to performing essential functions specific to certain cell types, septins are also important for normal cell division; they can act as "molecular fences" in membranes to keep proteins in specific locations and they are thought to act as protein "scaffolds" to assemble specific groups of proteins at the cell cortex. In most cells, septin proteins self-assemble into filaments that further assemble into complex higher-order structures such as rings and fibers. Despite the widespread distribution and varied functions of septins, little is known about the molecular mechanisms that direct septin assembly into filaments and the higher order organization of these filaments. A major goal of this project is to determine how septins assemble into an array of morphologically different forms within a single cell. The PI and her colleagues will investigate how septin structures are assembled, reorganized and regulated in the multinucleated, filamentous fungus, Ashbya gossypii. In this filamentous fungus, septins assemble into many morphologically distinct and dynamic structures, making it an excellent model for understanding the molecular controls of complex septin organization. The simple lifestyle and small genome of the fungus facilitates generation and analysis of mutants in regulatory proteins. Additionally, septin proteins can be visualized in living cells by linking the septin proteins to fluorescent proteins that are visible under the microscope. The specific goals of this project are to: 1. Identify the composition and dynamics of different septin complexes that coexist in one A. gossypii cell. 2. Establish the regulatory pathways that control the assembly of different septin rings. 3. Determine if the nuclear division cycle directs changes in the septin cortex in a multinucleated cell. These goals will be achieved using a combination of time-lapse fluorescence microscopy, electron microscopy and biochemistry for mutant analysis. This work is significant because the septins are a nearly ubiquitous, highly conserved protein family, yet major gaps exist in our knowledge of how they assemble and function in complexes in living cells. Furthermore, little is known about how internal or external signals lead to the assembly of elaborate types of septin structures. Knowledge gained from this work will provide a mechanistic foundation for understanding septin organization in other eukaryotic cells. The project will have substantial broader impact on education and training. The PI will mentor graduate and undergraduate students from multiple programs in research projects; undergraduate students will perform experiments, attend weekly lab meetings and present their data at Dartmouth's undergraduate research symposia.

细胞皮层的蛋白质通过传递和响应各种信号将细胞内部和外部连接起来。因此，细胞皮层是一个信息处理中心，细胞可以在这里接受环境或细胞内的各种刺激并作出反应。一个被称为septin的保守蛋白家族组织和调节真核细胞皮层的结构和过程。除了执行特定细胞类型的基本功能外，septin对正常细胞分裂也很重要；它们可以作为细胞膜上的“分子栅栏”，将蛋白质保持在特定的位置，它们被认为是蛋白质的“支架”，在细胞皮层组装特定的蛋白质群。在大多数细胞中，septin蛋白自组装成细丝，细丝进一步组装成复杂的高阶结构，如环和纤维。尽管septin分布广泛，功能多样，但人们对septin组装成细丝的分子机制以及这些细丝的高阶组织知之甚少。该项目的一个主要目标是确定septin如何在单个细胞内组装成一系列形态不同的形式。PI和她的同事将研究在多核丝状真菌Ashbya gossypii中septin结构是如何组装、重组和调节的。在这种丝状真菌中，septin组装成许多形态上不同的动态结构，使其成为理解复杂septin组织的分子控制的绝佳模型。真菌的简单生活方式和小基因组有利于调控蛋白突变体的产生和分析。此外，通过将septin蛋白与显微镜下可见的荧光蛋白连接，可以在活细胞中看到septin蛋白。该项目的具体目标是：1。鉴定一个棉蚜细胞中共存的不同septin复合物的组成和动态。2. 建立控制不同septin环组装的调控途径。3. 确定核分裂周期是否指导多核细胞中隔质皮层的变化。这些目标将通过结合延时荧光显微镜、电子显微镜和生物化学进行突变体分析来实现。这项工作意义重大，因为septin是一个几乎无处不在的高度保守的蛋白质家族，但我们对它们如何在活细胞复合体中组装和功能的了解存在重大空白。此外，对于内部或外部信号如何导致复杂类型的septin结构的组装知之甚少。从这项工作中获得的知识将为理解其他真核细胞中的septin组织提供机制基础。该项目将对教育和培训产生重大而广泛的影响。PI将指导来自多个专业的研究生和本科生进行研究项目；本科生将进行实验，参加每周的实验室会议，并在达特茅斯大学的本科生研究研讨会上展示他们的数据。