Mechanisms of Septin Assembly and Dynamics

智能优点细胞骨架是由蛋白质聚合物组成的，这种聚合物是细胞正常结构和功能所必需的。这些聚合物网络充当分离细胞成分、改变细胞形状的机器和轨道，并充当蛋白质和膜支架。间隔蛋白是细胞骨架中一种高度保守的成分，对细胞分裂、膜间隔和分泌等过程非常重要。Septin蛋白在真菌和哺乳动物中广泛表达。间隔蛋白在细胞中聚集在一起，形成更高层次的结构，如环、杆和纤维。我们使用一种名为Ashbya Cotsypii的遗传易驯化的丝状真菌模型来研究Septin，因为它的持久、动态和基于Septin的大型组装尤其适合显微镜和生化分析。该项目的目标是解决目前尚不了解的Septin结构的三个关键方面：1.确定活细胞中Septin细丝如何聚合和连接在一起；2.确定哪些因素控制Septin在细胞中组装在一起的位置；以及3.评估Septin细丝的动力学。这项研究采用了新颖的单分子和偏振荧光成像、基因突变分析和体外生化方法相结合的方法。这个项目中的实验解决了一个普遍的细胞工程问题：如何在细胞中的特定位置建立持久的、但动态的、大小恒定的结构。这个项目将发现细胞在空间中组织自己执行生命功能(如生长和分裂)的基本规则。更广泛的影响国际协会致力于培养本科生和研究生，并在社区中交流科学。作为本科生细胞生物学的课程主任，PI将扩展她和她的研究小组以Ashbya为模型系统开发的基于探究的实验单元。在撰写并获得了在生物学课堂上使用“点击器”的资助后，她开发了一系列问题，将以讲座为基础的细胞生物学课程转变为一个积极的学习空间，并将在未来的课程中继续扩展和与其他教师分享这个工具包。在过去的五年里，PI已经在实验室培养了七名本科生，其中包括两名数学和计算机科学专业的女性和一名计算机科学专业的男性。2010年夏天，一名本科生在伍兹霍尔的海洋生物实验室与PI一起工作，获得了REU奖。达特茅斯大学的两名本科生今年在PI的实验室完成了荣誉论文，现在他们的作品都作为第一作者发表了。她曾指导过两名女性博士生、一名女性博士后和一名女性技术员，并有五名女性轮换学生，其中两名是拉丁裔。PI在校园的许多场所谈论女性在科学领域面临的问题，以及平衡科学和母性的问题，包括各种女生联谊会教职员工晚宴、生物学本科生社团和本科生女性在科学领域的项目。他在2010年美国细胞生物学学会本科生年会特别会议上发表了主旨演讲。该协会已指导了两名教师-学者博士后研究员，通过与他们共同授课并每周就他们的教学提供反馈，促进他们作为教师的发展。这位PI在她所在社区的晚间研讨会上谈到了基础科学研究的重要性，并在中学科学教室里谈到了作为一名科学家的感觉。PI实验室的研究生定期向当地高中生和老师讲授光学显微镜的原理和我们实验室的研究。PI将通过继续这些描述的活动，将培训、指导和当地社区参与编织到她的研究计划中。

Intellectual meritThe cytoskeleton is made of protein-based polymers that are required for proper cell structure and function. These polymer networks serve as machines and tracks to segregate cell components, change cell shape, and act as protein and membrane scaffolds. The septins are a highly conserved component of the cytoskeleton important for processes such as cell division, membrane compartments, and secretion. The septin proteins are widely expressed from fungi to mammals. Septins come together in cells to build higher-order structures such as rings, bars, and fibers. We use a genetically tractable model filamentous fungus called Ashbya gossypii to study septins because its persistent, dynamic and large septin-based assemblies are especially amenable to microscopy and biochemical analysis. The objectives of this project address three key aspects of septin architecture that are presently not understood: 1. Determination of how septin filaments polymerize and link together in living cells; 2. Identification of what factors control where septins are assembled together in cells; and 3. Evaluation of the dynamics of septin filaments. The research employs a combination of novel single molecule and polarized fluorescence imaging, genetic mutant analysis and in vitro biochemical approaches. The experiments in this project tackle a general cellular engineering problem: how to erect persistent, yet dynamic, structures of a constant size at specific locations in the cell. This project will discover basic rules cells use to organize themselves in space to carry out functions of life such as growth and division. Broader ImpactsThe PI is committed to training undergraduates and graduate students and communicating science in the community. As the course director for undergraduate Cell Biology, the PI will expand on an inquiry-based lab unit developed by her and her research group using Ashbya as a model system. After writing and receiving a grant for "clickers" to use in Biology classes, she has developed a series of questions to change the lecture based cell biology class into an active learning space and will continue to expand and share this tool kit with other faculty in future offerings. The PI has trained seven undergraduates in the lab in the last five years, including two female math and computer science majors and one male computer science major. One undergraduate was awarded an REU in the summer of 2010 to work with the PI at the Marine Biological Lab in Woods Hole. Two Dartmouth undergraduates completed honors theses in the PI's lab this year and both now have had their work published as first authors. She has mentored two female PhD students, one female post-doc and one female technician and has had five female rotation students, two of whom are Latina. The PI speaks about issues facing women in science and balancing science and motherhood at many venues on campus including various sorority faculty dinners, at the Biology undergraduate society, and the undergraduate Women in Science Program. The PI was the keynote speaker at the American Society for Cell Biology Annual Meeting Special Session for undergraduate participants in 2010. The PI has mentored two teacher-scholar post-doctoral fellows to facilitate their development as teachers by co-teaching with them and providing weekly feedback on their teaching. The PI speaks at evening seminars in her community about the importance of basic science research and has spoken to middle school science classrooms about what it is like to be a scientist. Graduate students in the PI's lab regularly teach local high school students and teachers about the principles of light microscopy and the research in our lab. The PI will weave training, mentoring and local community involvement into her research program through the continuation of these described activities.