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中发挥着重要作用。 移动。随着细胞对外部信号的响应而移动， 细胞骨架不断重塑：肌动蛋白细丝网络 在额板上形成，附着在下面的 通过焦点粘合形成衬底，从而提供粘合剂 触点，用于拉回细胞体所需的牵引。 为了继续这一过程，肌动蛋白亚基必须连续 回收利用到了领先的边缘。尽管肌动蛋白在体内的组装速率 体外均极快，尖端拆解速率恒定 在体外测量(约1 S-1)太慢了，无法再循环 单体使这一过程得以继续。如果发生网状解聚 仅通过跑步机机制，这一速度已被估计 需要更接近~180 S-1才能保持30毫米/分钟的运动速度。这个 提高拆卸率最明显的方法是切断 花丝露出新的带刺末端，在那里解离率 腺苷二磷酸-肌动蛋白亚基的常数要高得多(&gt；7 S-1)。这个 Cofilin/ADF家族被认为是最明显的 候选人这样做是因为他们明显有能力 F-肌动蛋白，不封顶。我们使用了电子冷冻显微镜和 螺旋重建确定其在肌动蛋白上的结合位置 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 连接两个纵向相关的肌动蛋白亚基。装订 位点位于较低的肌动蛋白亚基的亚区2的轴向中心，并且 在上游肌动蛋白1和3亚区之间的裂隙处呈放射状 亚单位。我们的工作揭示了一种完全意想不到的(和独特的) 胶丝蛋白的性质，即它改变细丝缠绕的能力。AS 这种扭曲的变化的结果是，长丝上装饰着 Cofilin具有更短的肌动蛋白交叉(约75% 通常在F-肌动蛋白结构中观察到)。尽管它们的绑定 位点是不同的，cofilin与phalloidin竞争F-肌动蛋白 有约束力的。这是肌动蛋白结合蛋白的首次演示。 它通过改变长丝的扭度来竞争结合。更改 通过cofilin/ADF构建F-肌动蛋白似乎是一种新的机制 当细胞移入时，肌动蛋白细胞骨架可以通过它来重塑 对外部信号的响应。

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.