Collaborative Research: Understanding and controlling force generation by a centrin-based contractile system

力量的产生是许多与生命最相关的过程的基础：运动、生长和繁殖。纤毛虫疑似螺旋口的超快收缩代表了自然界最强大的生物力量生成。这种极端的运动被认为是由一种被称为肌丝的收缩蛋白网络驱动的，但人们对此知之甚少。该项目将结合实验和计算模型来阐明肌丝收缩的机制，以期揭示生物力产生的新原理。这些发现将为设计合成细胞的力产生系统奠定基础，例如控制细胞的形状和运动。该团队成立于2019年NSF Ideas Lab，致力于构建合成细胞，作为生活规则倡议的一部分。这项工作的更广泛的影响包括它的内在本质，揭示了可能是已知的最强大的生物马达的机械细节。其他活动将包括对高中、本科生、研究生和博士后学者进行多学科培训。每年有1000多名科学家和公众参观的海洋生物实验室(MBL)将展出一个永久性的“看到细胞”展览，该展览也将扩大。展示的一个主题是，细胞中的给定功能可以通过不同类型的细胞中的不同机制来实现。该项目将催化关于这一主题的新讨论，合成细胞的设计，以及它们可能对社会的影响，这些讨论的想法将被纳入展览并翻译成在线格式，以接触到广泛的观众。肌丝收缩由钙触发，肌丝由Centin EF-Hand蛋白质和Sfi1支架蛋白质组成。与研究良好的由ATP驱动的肌动球蛋白收缩系统相比，人们对肌球蛋白如何产生力量知之甚少。在多个尺度上的研究将产生定量的综合模型，解释分子构象变化如何在整个有机体中产生力量。使这项研究得以实现的一个关键进展是，该团队在体外通过仅由中心素和Sfi1组成的细丝重建了钙诱导的收缩。该项目将检验这一假设，即在分子水平上肌丝蛋白的特定构象变化是由钙触发的，以驱动这种生物体在毫米尺度上的超快收缩。其目的是1)确定调节体外组装和力产生的因素，2)在分子水平上阐明收缩的结构基础，3)确定肌丝网络、钙动力学、微管及其周围环境的相互作用如何产生整个生物体的超快收缩。从长远来看，这项工作将使人们能够以新的方式理解产生超快力量的独立生物学机制，这种机制可以被用来在体外和体内操纵生物材料。该项目由分子生物学和分子与细胞生物科学部门的系统和合成生物学计划共同资助。该奖项反映了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。

Force generation underlies many of the processes most associated with life: movement, growth, and reproduction. The ultrafast contraction of the ciliate Spirostomum ambiguum represents the most powerful biological force generation in nature. This extreme movement is thought to be driven by a contractile protein-based network known as myonemes, which is poorly understood. The project will combine experiments and computational modeling to elucidate the mechanism of myoneme contraction with a view toward revealing new principles of biological force generation. The findings will set the stage to engineer force-generating systems for synthetic cells, for example to control cell shape and movement. The team was established at a 2019 NSF Ideas Lab on building synthetic cells as part of the Rules of Life initiative. The Broader Impact of the work includes its intrinsic nature in revealing the mechanistic details of what may be the most powerful biologic motor known. Additional activities will include the multidisciplinary training of high school, undergraduate, graduate students, and post-doctoral scholars. A permanent exhibit on “Seeing Cells” at the Marine Biological Laboratory (MBL), where 1000+ scientists and members of the public visit each year, will also be expanded upon. A theme of the exhibit is that a given function in a cell can be accomplished through different mechanisms in different types of cells. The project will catalyze new discussions on this theme, the design of synthetic cells, and their possible impact on society, and ideas from these discussions will be incorporated into the exhibit and translated to an online format to reach a wide audience.Myoneme contraction is triggered by calcium, and myonemes are composed of centrin EF-hand proteins and Sfi1 scaffold proteins. In contrast to the well-studied ATP-driven actomyosin contractile system, little is known about how myonemes generate force. Studies at multiple scales will produce quantitative integrative models that explain how molecular conformational changes produce force in the whole organism. A key advance enabling the studies is the team’s reconstitution of calcium-induced contraction by filaments composed of only centrin and Sfi1 in vitro. The project will test the hypothesis that specific conformational changes of myoneme proteins at the molecular level are triggered by calcium to drive the ultrafast contraction at the millimeter scale in this organism. The aims are to 1) determine the factors that modulate assembly and force generation in vitro, 2) elucidate the structural bases of contraction at the molecular level, and 3) determine how the interplay of the myoneme network, calcium dynamics, microtubules, and their surroundings produce ultrafast contraction of the whole organism. In the long term, this work will enable novel understanding of an independent biological mechanism for ultrafast force generation, which can be harnessed to manipulate biological materials, both in vitro and in vivo.This project was co-funded by the Molecular Biophysics and the Systems and Synthetic Biology programs in the Division of Molecular and Cellular Biosciences.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.