MOLECULAR MODEL OF COFILIN REG IN ACTIN FILAMENT DYNAMICS & CELL FUNCTIONS

以肌动蛋白为基础的细胞骨架在细胞内起着重要的作用 运动 当细胞响应外部信号而移动时， 细胞骨架不断重塑：肌动蛋白丝网络， 形成于前板，附着在下面的 通过局部粘连，从而提供粘合剂 用于缩回单元体所需的牵引力的触点。 为了使这一过程继续下去，肌动蛋白亚基必须不断地被 再循环到前沿。 尽管肌动蛋白的组装率在 体外都是极其迅速的，尖端解体速率恒定 在体外测量的（约1 s-1）太慢而不能再循环 单体，以使这一进程继续下去。 如果发生净解聚 仅通过一个螺旋铣削机制，据估计， 将需要更接近~180 s-1以保持30 mm/min的运动。 提高拆卸率最明显的方法是切断 细丝暴露新的倒刺末端， ADP-肌动蛋白亚基的常数要高得多（> 7 s-1）。 的 cofilin/ADF蛋白家族被认为是最明显的 候选人这样做，因为他们明显的能力， F-肌动蛋白无帽。 我们用电子低温显微镜， 螺旋重建以确定其在肌动蛋白上结合位点 细丝。 我们的结构显示cofilin通过以下方式协同结合F-肌动蛋白： 桥接两个与细胞相关的肌动蛋白亚基。 的结合 该位点轴向居中于下部肌动蛋白亚基的亚结构域2， 在上肌动蛋白亚结构域1和3之间的裂缝处径向延伸 亚单位 我们的工作揭示了一个完全出乎意料的（和独特的） cofilin的性质，即其改变长丝捻度的能力。 作为 这种捻度变化的结果是， cofilin具有短得多的肌动蛋白交叉（大约75%的那些 通常在F-肌动蛋白结构中观察到）。 尽管其约束力 位点不同，cofilin与鬼笔环肽竞争F-肌动蛋白 约束力 这是第一次证明肌动蛋白结合蛋白 其通过改变长丝的捻度来竞争结合。 改变 通过cofilin/ADF的F-肌动蛋白结构似乎是一种新的机制 当细胞进入时， 响应外部信号。

The actin-based cytoskeleton plays an important role in cell locomotion. As cells move in response to external signals, the cytoskeleton is continuously remodeled: actin filament networks are formed at the frontal lamella, which attach to the underlying substratum through focal adhesions, thereby providing adhesive contacts for the traction necessary for retraction of the cell body. For this process to continue, actin subunits must be continuously recycled to the leading edge. Although the assembly rates of actin in vitro are extremely rapid, the pointed end disassembly rate constant measured in vitro (about 1 s-1) is far too slow to recycle the monomers for this process to continue. If net depolymerization occurs only by a treadmilling mechanism, it has been estimated that this rate would need to be nearer ~180 s-1 to maintain motion at 30 mm/min. The most obvious way to increase the disassembly rate is to sever filaments to expose new barbed ends, where the dissociation rate constant for ADP-actin subunits is much higher (> 7 s-1). The cofilin/ADF family of proteins were thought to be the most obvious candidates to do this because of their apparent ability to sever F-actin without capping. We have used electron cryomicroscopy and helical reconstruction to identify its binding site on actin filaments. Our structure shows cofilin binds F-actin cooperatively by bridging two longitudinally-associated actin subunits. The binding site is centered axially at subdomain 2 of the lower actin subunit and radially at the cleft between subdomains 1 and 3 of the upper actin subunit. Our work has revealed a totally unexpected (and unique) property of cofilin, namely, its ability to change filament twist. As a consequence of this change in twist, filaments decorated with cofilin have much shorter actin crossovers' (about 75% of those normally observed in F-actin structures). Although their binding sites are distinct, cofilin competes with phalloidin for F-actin binding. This is the first demonstration of an actin-binding protein which competes for binding by changing filament twist. Alteration of F-actin structure by cofilin/ADF appears to be a novel mechanism through which the actin cytoskeleton may be remodeled as cells move in response to external signals.