Surface-bound Regulation of Actin Polymerization

描述(由申请人提供)：细胞形状的变化对于抵抗病原体的免疫防御和正常的组织发育，包括细胞迁移，是重要的。异常调控在某些疾病过程中是一个关键点，包括癌症的转移。因为肌动蛋白皮质负责这些细胞形状的变化，我们的长期目标是了解它的调节和它改变形状的机制。我们开创了新的生物物理方法和新的纳米技术来探索肌动蛋白皮质的机械功能。皮质调节经常发生在膜结合因子控制含有许多倒刺末端的皮质斑块的情况下。通过研究Arp2/3和Ena/Vasp依赖的反应，我们将探讨膜-表面催化在调节皮质肌动蛋白的两种模式中的物理化学约束：片层和丝状足突。从本质上讲，我们建立了一个细胞前沿的模型，我们的长期目标是重建这些具有代表性的反应，以便在纳米颗粒表面上直接可视化。即使没有可视化，表面吸附也已经揭示了溶液研究中意想不到的生化相互作用。Arp2/3的表面激活揭示了一种新的覆盖蛋白质的机械作用，而Ena/Vasp的表面激活表明某些所提出的生化结合不是加速突起所必需的。为了实现我们的前两个具体目标，理解这两个效应还需要生物物理测量来监控机械性能。此外，了解这两个具有代表性的过程将为指导可视化皮质动力学的第三个更长期目标的发展提供合理的基础。

DESCRIPTION (provided by applicant): Changes in cell shape are important for immunological defense against pathogens and for normal tissue development, including cell migration. Aberrant regulation is a key point in some disease processes, including metastases of cancer. Because the actin cortex is responsible for these changes in cell shape, our long-term goal is to understand its regulation and its mechanisms for changing shape. We have pioneered new biophysical approaches and novel nanotechnology for probing the mechanical functions of the actin cortex. Cortical regulation often occurs with membrane-bound factors controlling a patch of cortex containing many barbed ends. By examining ARP2/3 and Ena/VASP-dependent reactions, we will probe the physicochemical constraints of membrane-surface catalysis in two modes of regulating cortical actin: lamellae and filopodia protrusions. Essentially building a model of the leading edges of cells, our longer-term Aim is to reconstitute these representative reactions for direct visualization on nanofabricated surfaces. Even without visualization, surface-adsorption already reveals biochemical interactions unexpected from solution studies. Surface-activation of ARP2/3 reveals a novel mechanical role for capping protein, whereas surface-activation of Ena/VASP suggests that certain proposed biochemical associations are not required to speed protrusion. Constituting our first two Specific Aims, understanding these two effects also requires biophysical measurements to monitor mechanical properties. In addition, understanding these two representative processes will provide a rational basis to guide development of the third, longer-term Aim of visualizing cortical dynamics.