Chan Chikio, Beltzner Christopher C, Pollard Thomas D
Department of Molecular Cellular, Yale University, PO Box 208103, New Haven, CT 06520-8103, USA.
Curr Biol. 2009 Apr 14;19(7):537-45. doi: 10.1016/j.cub.2009.02.060.
Actin-based cellular motility requires spatially and temporally coordinated remodeling of a network of branched actin filaments. This study investigates how cofilin and Arp2/3 complex, two main players in the dendritic nucleation model, interact to produce sharp spatial transitions between densely branched filaments and long, unbranched filaments.
We found that cofilin binding reduces both the affinity of actin filaments for Arp2/3 complex and the stability of branches. We used fluorescence spectroscopy to measure the kinetics of cofilin association with filaments and the resulting dissociation of Arp2/3 complex and TIRF microscopy to visualize filament severing and the loss of actin filament branches. Cofilin severs filaments optimally when few actin subunits are occupied but dissociates branches rapidly only at higher occupancies. Effective debranching is nevertheless achieved, as a result of cooperative binding and reduced affinity of Arp2/3 complex for the filament, at cofilin concentrations below those required for direct competition.
Cofilin rapidly dissociates Arp2/3 complex and branches by direct competition for binding sites on the actin filament and by propagation of structural changes in the actin filament that reduce affinity for Arp2/3 complex.
基于肌动蛋白的细胞运动需要分支肌动蛋白丝网络在空间和时间上的协调重塑。本研究探讨了树突状成核模型中的两个主要参与者——丝切蛋白和Arp2/3复合物如何相互作用，以在密集分支的丝和长的、无分支的丝之间产生明显的空间转变。
我们发现丝切蛋白的结合降低了肌动蛋白丝对Arp2/3复合物的亲和力以及分支的稳定性。我们使用荧光光谱法测量丝切蛋白与丝结合的动力学以及由此导致的Arp2/3复合物的解离，并使用全内反射荧光显微镜（TIRF显微镜）观察丝的切断和肌动蛋白丝分支的丢失。当很少有肌动蛋白亚基被占据时，丝切蛋白能最佳地切断丝，但只有在更高的占据率下才会迅速解离分支。然而，由于协同结合以及Arp2/3复合物对丝的亲和力降低，在低于直接竞争所需浓度的丝切蛋白浓度下仍能实现有效的去分支。
丝切蛋白通过直接竞争肌动蛋白丝上的结合位点以及通过肌动蛋白丝中结构变化的传播来降低对Arp2/3复合物的亲和力，从而迅速解离Arp2/3复合物和分支。
