Transitions: A unified cellular and in vitro approach to discover molecular mechanisms of microtubule dynamics and regulation

通过“过渡”奖，首席研究员将获得现代遗传学和成像方法方面的培训，并将这些技能应用于研究被称为微管的动态细胞聚合物，微管为真核细胞提供结构和形状。该项目的目标是在聚合亚基中发现新的、机制定义的突变，并了解微管的细胞功能如何响应聚合动力学的选择性扰动。这项工作的长期目标是通过结合体外和细胞研究，结合机制特异性微管蛋白突变，对微管动力学建立更全面的理解，将分子与表型联系起来。从这项工作中产生的结果和材料将为一个具有重要意义的基本过程提供新的见解。除了工作本身的影响之外，该项目更广泛的影响还包括为PI提供科学动画方面的新培训，这将使PI的家乡机构能够通过新的纳米课程传播这一日益重要的学科。研究生和本科生也将通过参与研究接受跨学科的培训，PI将继续在当地公立学校开展以显微镜为基础的推广活动。微管的动态特性是其功能的核心，它们源于单个αβ-微管蛋白亚基的结构和生化特性及其在聚合物中的相互作用。理解微管动力学和调控背后的分子机制一直具有挑战性，因为这样做需要将“一次一个分子”的生物化学和结构观点与跨越不同长度和时间尺度的集体行为的低分辨率测量相结合。在项目的专业发展阶段，PI将接受现代酵母遗传学和成像方面的培训，通过执行抑制和新生筛选来识别干扰微管动力学的新突变，并通过定义其对细胞功能的影响。该项目的研究部分将包括两个互补的方向：(i)使用更大规模的遗传筛选来识别干扰微管动力学的新隐藏突变，以及（ii）通过表达可变数量的机制特异性微管蛋白突变来确定“重新调整”细胞微管动力学的纺锤体组装和染色体分离的后果。这些新的方法和方向将克服当前的瓶颈，并允许PI进行更广泛的研究，将机制研究与细胞生理学联系起来。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Through this “Transitions” award, the principal investigator will obtain training in modern genetics and imaging methods and apply these skills to the study of the dynamic cellular polymers called microtubules, which provide structure and shape to eukaryotic cells. The goals of this projects are to discover new, mechanism-defining mutations in the polymerizing subunits and to understand how cellular functions of microtubules respond to selective perturbations of polymerization dynamics. The long-term goal of the work is to create a more wholistic understanding of microtubule dynamics that bridges molecules to phenotypes using a combination of in vitro and in cell studies, unified with mechanism-specific tubulin mutants. The results and materials arising from this work will provide new insights into a fundamental process of high significance. In addition to the impact of the work itself, broader impacts of the project include new training for the PI in scientific animation that will allow dissemination of this increasingly important discipline at the PI’s home institution through a new nanocourse. Graduate and undergraduate students will also receive interdisciplinary training by taking part in the research, and the PI will continue microscopy-based outreach activities at local public schools.The dynamic properties of microtubules are central to their function, and they derive from the structural and biochemical properties of individual αβ-tubulin subunits and their interactions within the polymer. Understanding the molecular mechanisms underlying microtubule dynamics and regulation has been challenging because doing so requires integrating ‘one molecule at a time’ views of biochemistry and structure with lower-resolution measurements of collective behavior spanning different length and time scales. During the professional development phase of the project, the PI will receive training in modern yeast genetics and imaging by performing suppressor and de novo screens to identify new mutations that perturb microtubule dynamics and by defining their consequences for cellular function. The research portion of the project will encompass two complementary directions: (i) using larger scale genetic screening to identify new buried mutations that perturb microtubule dynamics, and (ii) determining the consequences for spindle assembly and chromosome segregation of ‘re-tuning’ cellular microtubule dynamics by expressing variable amounts of mechanism-specific tubulin mutants. These new methods and directions will overcome current bottlenecks and allow the PI to undertake a broader scope of research to connect mechanistic studies to cellular physiology.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.