Biochemical Regulation of Actin Cytoskeleton Assembly

描述(申请人提供)：Palladin是一种新的92kD蛋白质， 似乎在基于肌动蛋白的结构蛋白的组装中起着关键作用 数组。我们假设Palladin是一种关键的支架蛋白，它为 调节组装和更高顺序的多个蛋白质的对接位置 肌动蛋白细丝的组织。由于肌动蛋白细胞骨架提供了 许多基本细胞过程的结构框架(包括 传播、粘连和迁移)，重要的是要准确地了解 Palladin具有调节细胞骨架组织的功能。为此， 将解决以下问题：(1)什么是结构和 与Palladin结合的细胞骨架的调节蛋白？为了这些 研究，我们将以共享的序列同源性为指导 Palladin和已知的细胞骨架蛋白，指导我们进行潜在的结合 合伙人。(2)Palladin是如何监管的？为了回答这个问题，我们将采取 观察到当细胞四舍五入时Palladin被修饰的优点 才能进入有丝分裂。我们将研究它的生化性质。 修饰，我们将探索有丝分裂特异性的修饰是如何 Palladin会影响它与通常伴侣的结合能力。(3)什么 Palladin结构域是其结合作用所必需的吗？对接地点在 Palladin将被映射到特定的氨基酸，这样我们就可以设计Palladin 缺乏提供某些结合位点的能力的突变体。 然后，这些帕拉丁突变体将在活细胞中表达，以探索 对细胞骨架组装的影响。总而言之，我们认为帕拉丁是一种 独特而强大的工具，用来研究细胞机制 控制细胞骨架在生活中的组装、维护和调节 细胞。细胞的扩散、黏附和迁移对于这样的过程至关重要 如伤口愈合、炎症反应、轴突生长和癌症 转移；因此，更好地理解了基于肌动蛋白的调节 结构可能在许多不同的领域具有潜在的健康益处。

DESCRIPTION (provided by applicant): Palladin is a novel 92 kD protein that appears to play a critical role in the assembly of actin-based structural arrays. We hypothesize that palladin is a key scaffold protein that provides docking sites for multiple proteins that regulate the assembly and higher-order organization of actin filaments. Since the actin cytoskeleton provides the structural framework for many fundamental cellular processes (including spreading, adhesion, and migration), it is important to understand exactly how palladin functions to modulate cytoskeletal organization. To this end, the following questions will be addressed: (1) What are the structural and regulatory proteins of the cytoskeleton that bind to palladin? For these studies, we will be guided by sequence homologies that are shared between palladin and known cytoskeletal proteins, to direct us to potential binding partners. (2) How is palladin regulated? To answer this question, we will take advantage of the observation that palladin becomes modified when cells round up to enter mitosis. We will investigate the biochemical nature of this modification, and we will explore how mitosis-specific modifications of palladin impact upon its ability to bind to its usual partners. (3) What domains of palladin are required for its binding interactions? Docking sites on palladin will be mapped to specific amino acids, so that we can design palladin mutants that are deficient in the ability to provide certain binding sites. These palladin mutants will then be expressed in living cells, to explore the effects on cytoskeletal assembly. In summary, we believe that palladin is a unique and powerful tool with which to study the cellular mechanisms that govern how the cytoskeleton is assembled, maintained and regulated in living cells. Cell spreading, adhesion and migration are critical for processes such as wound healing, inflammation reaction, neurite outgrowth and cancer metastasis; thus, a better understanding of the regulation of actin-based structures could have potential health benefits in many different fields.