IntBIO: Linking genome to phenome to understand the function of Masticatory Myosin

肌肉是动物产生力量、运动和与环境相互作用的主要手段。肌肉功能是从分子到整个肌肉和肌肉/骨骼系统的层次结构的结果。然而，对肌肉如何工作的深入理解受到没有完全跨越这些尺度的研究的限制。研究基因如何编码蛋白质，数万亿不同种类的蛋白质如何相互作用，肌肉质量和形状如何影响收缩，以及骨骼几何形状如何调节运动的速度和力度的多尺度研究是实现对肌肉性能的新见解的最佳途径。该项目利用了最先进的多尺度实验技术和肌肉数学建模的协同作用，以了解啮齿动物模型的咬伤性能。啮齿类动物有一系列的咬合策略、下颌几何形状和肌肉蛋白质组成，包括一种独特的肌肉运动蛋白肌球蛋白，称为咀嚼肌球蛋白。先前的研究表明，咀嚼肌球蛋白具有特殊的特性，可以为肌肉如何产生咬合力提供新的见解。该项目将测试咬合性能是否主要受肌肉大小和形状、骨骼几何形状或咀嚼肌球蛋白的存在影响。从这一综合补充研究中收集的见解将为肌肉组织特征如何控制肌肉功能的未来研究提供信息。在这个项目更广泛的影响活动中，生物功能“不仅仅是其各部分的总和”的原则将通过一个定制编码的电脑游戏来介绍，这个游戏将向来自STEM中代表性不足的群体的中学生讲授肌肉的功能。如果没有本项目中使用的多尺度收敛方法，理解像肌肉这样的复杂生物系统是具有挑战性的。肌肉生理学的一个基本概念是力量和速度之间的权衡。然而，肌蛋白肌球蛋白的咀嚼异构体被认为既有力又快速。连接肌球蛋白功能域的两个分子环序列的独特机械化学可以解释这种矛盾的现象。这一想法将通过C2C12肌肉细胞的靶向诱变、体外运动测定和单纤维实验来测试，这些实验可以测量不同肌球蛋白聚集体的峰值和负载速度。另一种观点认为，咀嚼肌球蛋白作用下颌骨肌肉的快速和强壮表现是由于器官和生物体在肌肉大小、几何形状、咬类型和生物力学方面的差异，这些差异可以缓冲肌球蛋白的力量或速度限制。这一想法将在实验中得到验证，在咬咬具有控制大小和不同硬度的食物时，使用肌电图、声学测量法、微力扣和x射线运动形态重建来测量肌肉激活、张力、力和杠杆动力学。这些方法将允许测定实现的咬合力-速度。啮齿动物是确定力-速度调节的跨尺度机制基础的理想选择，因为它们的表型具有不同的肌球蛋白异构体表达，但具有共同的咬伤生物力学。跨生物尺度（肌动球蛋白、肌纤维、完整肌肉和整个喂养系统）的力-速度关系将使用减法方法进行耦合，多尺度肌肉数学模型将用于确定具有或缺乏咀嚼肌球蛋白的几何相似或不同啮齿动物的紧急表现的机制基础。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。

Muscles provide the primary means for animals to produce force, move, and interact with their environment. Muscle function is a consequence of hierarchical structure from molecular to whole muscle and muscle/skeletal systems. However, deep understanding of how muscles work is limited by research that does not fully cross these scales. Multiscale studies that investigate how genes encode proteins, how trillions of proteins of different kinds interact, how muscle mass and shape influence contraction, and how skeletal geometry tunes the speed and forcefulness of movements are the best avenue for enabling new insights into muscle performance. This project leverages the synergy of state-of-the-art, multiscale experimental techniques, and mathematical modeling of muscle, to understand bite performance in rodent models. Rodents have a range of bite strategies, jaw geometries, and muscle protein compositions, including a unique type of the muscle motor-protein myosin called masticatory myosin. Previous studies have suggested that masticatory myosin has exceptional properties that could provide new insights into how muscles produce bite forces. This project will test whether bite performance is influenced primarily by muscle size and shape, skeletal geometry, or the presence of masticatory myosin. Insights gathered from this integrative complement of studies will inform future studies of how muscle function is controlled by features of muscle tissue. In the broader impact activities of this project, the principle that biological function is “more than the sum of its parts” will be introduced through a custom-coded computer game that will teach secondary school students from groups underrepresented in STEM about how muscles function.Understanding complex biological systems like muscle is challenging without multiscale convergence approaches, such as those used in this project. A fundamental idea in muscle physiology is the trade-off between force and velocity. However, a masticatory isoform of the myoprotein myosin has been suggested to be both forceful and fast. Unique mechanochemistry of two molecular loop sequences connecting myosin functional domains could explain this paradoxical phenomenon. This idea will be tested using targeted mutagenesis of C2C12 muscle cells, in-vitro motility assays, and single-fiber experiments that measure peak and loaded velocity of diverse myosin aggregates. An alternate idea is that fast and strong performance of jaw muscles with masticatory myosin is due to organ- to organism-scale differences in muscle size, geometry, bite type, and biomechanics that can buffer myosin force or speed limitations. This idea will be tested in experiments where muscle activation, strain, force, and leverage dynamics are measured using electromyography, sonomicrometry, micro force-buckles, and X-ray Reconstruction of Moving Morphology during biting on food items with controlled size and varying hardness. These methods will permit determination of the realized force-velocity of biting. Rodents are ideal for determining the cross-scale mechanistic bases of force-velocity modulation, as phenotypes exist with distinct myosin isoform expression but shared bite biomechanics. Force-velocity relationships across biological scales (actomyosin, myofibers, intact muscles, and whole feeding systems) will be coupled using a subtractive approach, and multiscale mathematical muscle modeling will be used to determine the mechanistic bases for emergent performance in geometrically similar or different rodents that either possess or lack masticatory myosin.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.