BIOPHYSICAL STUDIES OF CYTOSKELETAL FUNCTION

拟议工作的长期目标是大致了解如何 细胞的功能是机械的，更具体地说，是什么力量和 结构决定了它们的形状和变形能力。方法是 基于细胞形状和变形能力受 细胞骨架。由于细胞骨架是系统相互作用的聚合物 细丝，它的粘弹性和细胞的粘弹性都是可以理解的 在聚合物化学方面。要达到这种程度的理解，就必须 有必要对细胞进行定量测量，因此细胞骨架， 粘弹性。为此，一种测量细胞的NEV方法 粘弹性基于确定细胞对 已开发出局部压痕。此方法将用于 确定三种细胞骨架系统对 通过单独干扰细胞的变形能力来控制细胞的变形能力 每个系统的结构。此外，它的机械功能 细胞核和细胞-底物以及细胞-细胞的相互作用 检查过了。肌球蛋白的机械功能将在突变体中进行检测 完全缺乏这种蛋白质的Dictyostatus细胞。机械的 分泌细胞的生理激活引起的变化和 白细胞将从细胞骨架的角度进行测定和解释 流程。为了支持这项关于细胞的工作，还将对 纯化细胞骨架蛋白在简化重组中的相互作用 系统。例如，荧光漂白回收将被用于 描述α-肌动蛋白与肌动蛋白细丝的相互作用。

The long range goal of the proposed work is to understand generally how cells function mechanically and, more specifically, what forces and structures determine their shapes and deformabilities. The approach is based on the hypothesis that cell shape and deformability are governed by the cytoskeleton. Since the cytoskeleton is a system interacting polymer filaments, both its and the cell's viscoelasticity should be understandable in terms of polymer chemistry. To achieve this level of understanding it is necessary to measure quantitatively cellular, and therefore cytoskeletal, viscoelasticity. Toward this end a nev method for measuring cellular viscoelasticity based on determining the resistance of the cell to localized indentation has been developed. This method will be used to determine the relative contributions of the three cytoskeletal systems to the control of cellular deformability by interfering separately with the structure of each of the systems. In addition the mechanical function of the nucleus and of cell-substrate and cell-cell interactions will be examined. The mechanical functions of myosin will be examined in mutant Dictyostelium cells which entirely lack this protein. The mechanical changes resulting from physiological activation of secretory cells and leukocytes will be determined and interpreted in terms of cytoskeletal processes. Supporting this work on cells there will also be studies of the interactions of purified cytoskeletal proteins in simplified reconstituted systems. For example, fluorescence photobleaching recovery will be used to characterize the interactions of alpha-actinin with actin filaments.