Department of Physiology and Biochemistry and Molecular Biophysics Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA.
Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY, 10065, USA.
Nat Commun. 2022 Feb 2;13(1):628. doi: 10.1038/s41467-022-28112-2.
Positive feedback loops involving signaling and actin assembly factors mediate the formation and remodeling of branched actin networks in processes ranging from cell and organelle motility to mechanosensation. The Arp2/3 complex inhibitor Arpin controls the directional persistence of cell migration by interrupting a feedback loop involving Rac-WAVE-Arp2/3 complex, but Arpin's mechanism of inhibition is unknown. Here, we describe the cryo-EM structure of Arpin bound to Arp2/3 complex at 3.24-Å resolution. Unexpectedly, Arpin binds Arp2/3 complex similarly to WASP-family nucleation-promoting factors (NPFs) that activate the complex. However, whereas NPFs bind to two sites on Arp2/3 complex, on Arp2-ArpC1 and Arp3, Arpin only binds to the site on Arp3. Like NPFs, Arpin has a C-helix that binds at the barbed end of Arp3. Mutagenesis studies in vitro and in cells reveal how sequence differences within the C-helix define the molecular basis for inhibition by Arpin vs. activation by NPFs.
正反馈环涉及信号转导和肌动蛋白组装因子，介导从细胞和细胞器运动到机械感觉的分支肌动蛋白网络的形成和重塑。Arp2/3 复合物抑制剂 Arpin 通过中断涉及 Rac-WAVE-Arp2/3 复合物的反馈环来控制细胞迁移的定向持久性，但 Arpin 的抑制机制尚不清楚。在这里，我们描述了 Arpin 与 Arp2/3 复合物在 3.24-Å 分辨率下的冷冻电镜结构。出乎意料的是，Arpin 与 Arp2/3 复合物的结合方式类似于激活复合物的 Wiskott-Aldrich 综合征蛋白（ WASP ）家族成核促进因子（ NPFs ）。然而，NPFs 结合到 Arp2/3 复合物的两个位点，即 Arp2-ArpC1 和 Arp3 ，而 Arpin 仅结合到 Arp3 的位点。与 NPFs 一样，Arpin 具有一个 C 螺旋，该螺旋结合在 Arp3 的游离端。体外和细胞内的突变研究揭示了 C 螺旋内的序列差异如何定义了 Arpin 抑制与 NPFs 激活的分子基础。
