Cytoskeleton-based regulation of microtubule motors in intracellular transport.

细胞的形态和机械特性是由细胞骨架（一组生物聚合丝）维持的。细胞骨架是高度动态的，不断重塑以允许细胞形状的改变，这是细胞分裂和运动等基本功能所必需的。同时，细胞骨架细丝作为运动蛋白携带囊泡货物到细胞内不同位置的轨迹。定向运输是至关重要的，特别是对于像人类神经元这样的长细胞，可以延伸到1米。这些双重结构和运输角色对细胞骨架提出了相互竞争的要求——同时作为固体来维持细胞形态，同时作为液体来允许囊泡货物和细胞器通过细胞空间有效运动。为了满足各种各样的需求，细胞骨架细丝不断地聚合和解聚合，交联蛋白在细丝和细胞膜之间形成动态键，从而允许机械性能在不同的时间和长度尺度上进行调整。**细胞骨架不是简单地作为运动蛋白的被动跟踪，而是通过其网络结构、翻译后修饰和细胞骨架相关蛋白主动调节运动蛋白的活性。这些细胞骨架线索指导细胞内交通，以维持适当的空间和时间定位的蛋白质，mRNA，细胞器和信号分子。**本发现项目旨在(1)识别和表征允许细胞骨架动态适应细胞机械特性的分子机制，以及(2)了解细胞骨架如何调节运动蛋白功能以实现细胞内货物的定向运输。在提出的发现计划中，我们将开发先进的光学捕获技术，对细胞环境的频率依赖性粘弹性特性进行高分辨率测量，并检查干扰细胞骨架交联剂动力学对动态机械响应的影响。此外，我们将使用新的体外重构分析来检查分离的内源性货物在天然细胞骨架网络上的运动性。通过该项目，两名博士生和两名本科生将成为生物物理学、细胞生物学和分析技术方面的专家。**建议的研究将导致对细胞骨架如何动态调整其机械反应和调节运动蛋白以指导细胞内运输的新见解，导致我们对细胞生物学的基本理解的进步，以及设计由生物运动蛋白驱动的工程纳米级设备的能力。

The morphology and mechanical properties of the cell are maintained by the cytoskeleton, a set of biopolymeric filaments. The cytoskeleton is highly dynamic, constantly remodeling to allow changes in cell shape required for essential functions like cell division and motility. At the same time, cytoskeletal filaments serve as tracks along which motor proteins carry vesicular cargoes to different locations in the cell. Directed transport is critical, particularly for long cells like neurons that can extend up to 1 m in humans. These dual structural and trafficking roles place competing demands on the cytoskeleton - to simultaneously act as a solid to maintain cell morphology, and as a fluid to allow efficient movement of vesicular cargoes and organelles through the cellular space. To meet a diverse array of demands, cytoskeletal filaments continually polymerize and depolymerize and crosslinking proteins form dynamic bonds between filaments and to the cell membrane, allowing mechanical properties to be tuned across various time and length scales.**The cytoskeleton does not simply act as a passive track for motor proteins, instead it actively modulates their activity through its network architecture, post-translational modifications, and cytoskeletal-associated proteins. These cytoskeletal cues direct intracellular traffic to maintain the proper spatial and temporal localization of proteins, mRNA, organelles, and signaling molecules.**This Discovery program seeks to (1) identify and characterize the molecular mechanisms that allow the cytoskeleton to dynamically adapt the cell's mechanical properties, and (2) understand how the cytoskeleton modulates motor protein function to achieve the directed transport of intracellular cargoes. In the proposed Discovery program, we will develop advanced optical trapping techniques to perform high-resolution measurements of the frequency-dependent viscoelastic properties of the cellular environment, and examine the effect of perturbing the kinetics of cytoskeletal crosslinkers on the dynamic mechanical response. In addition, we will use novel in vitro reconstitution assays to examine the motility of isolated endogenous cargoes on native cytoskeletal networks. Through this program, two PhD students and two undergraduate students will become experts in biophysical, cell biological, and analytical techniques.**The proposed studies will result in novel insights into how the cytoskeleton dynamically tunes its mechanical response and regulates motor proteins to direct intracellular transport, leading to advances in our fundamental understanding of cell biology, and the ability to design engineered nano-scale devices powered by biological motor proteins.