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

通过这一“转变”奖，首席研究人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被称为微管的动态细胞聚合物的研究，微管为真核细胞提供结构和形状。该项目的目标是发现聚合亚基中新的、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建立对微管动力学的更完整的理解，使用体外和细胞研究相结合的方法，结合机制特定的微管蛋白突变体，将分子与表型联系起来。这项工作的结果和材料将为一个具有重要意义的基本进程提供新的见解。除了工作本身的影响外，该项目的更广泛的影响包括对国际和平机构进行新的科学动画培训，这将允许通过一个新的纳米课程在国际和平机构的本土机构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培训，PI将继续在当地公立学校开展基于显微镜的推广活动。微管的动态特性是其功能的核心，它们来自于单个αβ-微管蛋白亚基的结构和生化特性以及它们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是具有挑战性的，因为要做到这一点，需要将生物化学和结构的“一次一个分子”的观点与跨越不同长度和时间尺度的集体行为的较低分辨率测量相结合。在该项目的专业开发阶段，PI将接受现代酵母遗传学和成像方面的培训，方法是进行抑制子和从头筛查，以确定扰乱微管动力学的新突变，并确定它们对细胞功能的影响。该项目的研究部分将包括两个相辅相成的方向：(I)使用更大规模的遗传筛选来确定扰乱微管动力学的新的隐藏突变，以及(Ii)通过表达不同数量的机制特定微管突变体来确定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目前的瓶颈，并允许PI承担更广泛的研究范围，将机械学研究与细胞生理学联系起来。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势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.