Dynamics Control and Analysis of Biopolymer Networks

描述（由申请人提供）： 该项目的目标是开发新技术，用全息激光镊子阵列分析和控制生物聚合物网络微结构，这是一种功能强大的新仪器，其潜力刚刚开始探索。 该工具将允许复杂的空间分布的扰动和动态生物聚合物网络中的力的测量。 虽然这些技术将在肌动蛋白上开发，但它们应该适合于研究广泛的网络形成生物分子中的力和前沿不稳定性。 网络对扰动的响应应该揭示聚合前沿动力学和网络的机械性质之间的相互关系。 理解这种相互关系对于理解各种细胞骨架蛋白的聚合动力学如何影响细胞功能至关重要，例如，在与细胞移动性相关的形态学变化中。 了解和控制细胞的流动性对人类健康的许多方面都有影响，从增强损伤后的神经再生到减少转移性癌细胞的流动性。 具体目标1：使用全息镊子阵列以亚微米分辨率控制肌动蛋白聚合前沿。 聚合前沿的空间扩展扰动将通过热加热或通过影响聚合的分子的局部释放用镊子阵列引入。 从聚合前沿对扰动的响应中，研究人员可以提取特征长度尺度和时间尺度，并研究不稳定性。 具体目标2：开发使用全息镊子阵列测量力的空间分布的技术。 皮科牛顿力将与被困珠和被困扩展对象进行测量。 他们将测量聚合肌动蛋白前沿产生的力以及这些力之间的相关性。 通过这种技术，他们将确定聚合力如何取决于前沿的形状，生长速度和其他参数。 具体目标3：用多个激光陷阱使含有肌动蛋白的囊泡变形，以探测网络特性和机械不稳定性。 用多个激光镊子固定和变形膜将使我们能够探测并可能控制突起和不规则膜形状的发展，这些突起和不规则膜形状类似于在活细胞中观察到的板状伪足或伪足。 这项工作将为研究细胞运动的新方法奠定技术和分析基础。

DESCRIPTION (provided by applicant): The goal of this project is to develop new techniques to analyze and control biopolymer network microstructures with a holographic laser tweezer array, a powerful new instrument whose potential is just beginning to be explored. This tool will permit complex spatially distributed perturbations and measurements of forces in dynamic biopolymer networks. While the techniques will be developed on actin, they should be suitable to investigate forces and front instabilities in a broad range of network forming biomolecules. The response of the networks to perturbations should reveal the interrelationship between the polymerization front dynamics and the mechanical properties of the network. Understanding this inter-relationship is crucial for understanding how the polymerization dynamics of various cytoskeletal proteins affect cell function, for example, in the morphological changes associated with cell mobility. Understanding and controlling cell mobility has implications for many aspects of human health, from enhancing nerve regeneration after injury to reducing the mobility of metastatic cancer cells. Specific Aim 1: Control actin polymerization fronts with submicron resolution using a holographic tweezer array. Spatially extended perturbations of polymerization fronts will be introduced with the tweezer array by thermal heating or by localized uncaging of molecules that affect the polymerization. From the response of the polymerization front to the perturbation the investigators can extract characteristic length scales and timescales, and investigate instabilities. Specific Aim 2: Develop technology to measure the spatial distribution of forces using the holographic tweezer array. Pico Newton forces will be measured with trapped beads and trapped extended objects. They will measure the forces generated by a front of polymerizing actin and the correlations between those forces. With this technique they will determine how the polymerization force depends on the shape of the front, growth velocity, and other parameters. Specific Aim 3: Deform actin-containing vesicles with multiple laser traps to probe network properties and mechanical instabilities. Holding and deforming the membrane with multiple laser tweezers will permit us to probe and possibly control the development of protrusions and of irregular membrane shapes similar to the lamellipodia or pseudopodia observed in living cells. This work will lay the technical and analytical foundations for a new approach to investigating cell motility.