Structure and functions of the actin cytoskeleton

项目总结/摘要 肌动蛋白细胞骨架是细胞的主要组成部分，在细胞的各个方面都具有关键功能 生理学，包括细胞运动、形状和力学、细胞和组织形态发生、细胞-细胞和细胞-基质 膜细胞器的相互作用和动力学。肌动蛋白细胞骨架结构和功能的异常 和/或动力学对人类病理学，特别是癌症和神经变性， 免疫和心血管疾病。肌动蛋白细胞骨架在细胞中起着不可或缺的作用， 以许多不同的组合产生大的推力、拉力和阻力。执行这些 肌动蛋白丝具有多种功能，通过多种辅助蛋白被组织成不同的结构阵列。 尽管进行了广泛的研究，但这些基于肌动蛋白的分子机器的确切组织通常是 未知实现这一目标的主要障碍是难以在高水平上解析肌动蛋白细胞骨架结构。 单灯丝水平。如果不了解结构， 不完整我的实验室使用一种独特的方法来解决这个问题。我们利用铂金 复制电子显微镜（PREM），这是唯一能够结合联合收割机高分辨率成像的 细胞骨架的完整覆盖，并有效地将细胞骨架结构与肝脏相关。 细胞动力学在PREM的帮助下，我的实验室为理解 在一系列的通用和专门的细胞类型的细胞骨架功能。在本申请中，我们在 在今后五年中，我们将努力解决以下四个主要类别的问题： 肌动蛋白细胞骨架功能：（1）蛋白原-微管如何调节肌动蛋白的降解活性 定向迁移的细胞骨架;（2）收缩-非肌肉肌球蛋白II之间的相互作用 旁系同源物调节收缩力的极化亚细胞分布;（3）细胞力学-差异如何 在肌动蛋白皮质的分子结构中， （4）膜动力学-分支肌动蛋白网络如何促进网格蛋白包被的细胞内陷 膜结构域我们在PREM方面的专业知识，以及广泛的其他细胞生物学，成像， 功能，生物化学和分子生物学方法使我们处于独特的地位， 我们对肌动蛋白细胞骨架功能的理解。反过来，这些知识可能会提供重要的新见解， 如何对抗与肌动蛋白细胞骨架功能障碍有关的人类疾病。

PROJECT SUMMARY/ABSTRACT The actin cytoskeleton is a major cellular component with key functions in virtually every aspect of cell physiology including cell motility, shape and mechanics, cell and tissue morphogenesis, cell-cell and cell-matrix interactions, and dynamics of membrane organelle. Aberrations in actin cytoskeleton structure, functions and/or dynamics contribute significantly to human pathologies, especially to cancer and neurodegenerative, immune and cardiovascular diseases. The actin cytoskeleton plays indispensable roles in cells due to its ability to generate large pushing, pulling and resistance forces in many different combinations. To perform these diverse functions, actin filaments are organized into diverse structural arrays by multiple accessory proteins. Despite extensive research, the exact organization of these actin-based molecular machineries is frequently unknown. The main barrier toward this goal is the difficulty of resolving actin cytoskeleton architecture at a single-filament level. Without knowledge of the structure, functional understanding of the machinery is incomplete. My lab uses a distinctive approach to overcome this problem. We take advantage of platinum replica electron microscopy (PREM), which is uniquely able to combine high resolution imaging of the cytoskeleton with full coverage of the whole cell and to efficiently correlate the cytoskeleton structure with live cell dynamics. With help of PREM, my lab has made multiple fundamental contributions toward understanding of cytoskeleton functions in a range of generic and specialized cell types. In this application, we propose in the course of the next five years to address the following questions representing each of the four major categories of actin cytoskeleton functions: (1) Protrusion – How microtubules regulate protrusive activity of the actin cytoskeleton for directional migration; (2) Contraction – How an interplay between nonmuscle myosin II paralogs regulates polarized subcellular distribution of contractile forces; (3) Cell mechanics – How differences in the molecular architecture of the actin cortex are linked to different mechanical properties of normal and cancer cells; (4) Membrane dynamics – How branched actin networks promote invagination of clathrin-coated membrane domains. Our expertise in PREM in addition to a broad range of other cell biological, imaging, functional, biochemical, and molecular biological methods puts us in unique position to significantly advance our understanding of actin cytoskeleton functions. In turn, this knowledge may provide important new insights into how to combat human diseases associated with actin cytoskeleton malfunctions.