University of North Carolina Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; Howard Hughes Medical Institute, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
University of North Carolina Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Dev Cell. 2015 Jan 12;32(1):54-67. doi: 10.1016/j.devcel.2014.10.026. Epub 2014 Dec 24.
Cells contain multiple F-actin assembly pathways, including the Arp2/3 complex, formins, and Ena/VASP, which have largely been analyzed separately. They collectively generate the bulk of F-actin from a common pool of G-actin; however, the interplay and/or competition between these pathways remains poorly understood. Using fibroblast lines derived from an Arpc2 conditional knockout mouse, we established matched-pair cells with and without the Arp2/3 complex. Arpc2(-/-) cells lack lamellipodia and migrate more slowly than WT cells but have F-actin levels indistinguishable from controls. Actin assembly in Arpc2(-/-) cells was resistant to cytochalasin-D and was highly dependent on profilin-1 and Ena/VASP but not formins. Profilin-1 depletion in WT cells increased F-actin and Arp2/3 complex in lamellipodia. Conversely, addition of exogenous profilin-1 inhibited Arp2/3 complex actin nucleation in vitro and in vivo. Antagonism of the Arp2/3 complex by profilin-1 in cells appears to maintain actin homeostasis by balancing Arp2/3 complex-dependent and -independent actin assembly pathways.
细胞内包含多种 F-肌动蛋白组装途径，包括 Arp2/3 复合物、formin 和 Ena/VASP，这些途径在很大程度上是分别进行分析的。它们共同从一个常见的 G-肌动蛋白池中产生大部分 F-肌动蛋白；然而，这些途径之间的相互作用和/或竞争仍知之甚少。使用源自 Arpc2 条件性敲除小鼠的成纤维细胞系，我们建立了具有和不具有 Arp2/3 复合物的配对细胞。Arpc2(-/-)细胞缺乏片状伪足，迁移速度比 WT 细胞慢，但 F-肌动蛋白水平与对照无明显差异。Arpc2(-/-)细胞中的肌动蛋白组装对细胞松弛素 D 具有抗性，并且高度依赖于原肌球蛋白-1 和 Ena/VASP，但不依赖于formin。WT 细胞中原肌球蛋白-1 的耗竭增加了 F-肌动蛋白和片状伪足中的 Arp2/3 复合物。相反，外源性原肌球蛋白-1 的添加抑制了体外和体内的 Arp2/3 复合物肌动蛋白成核。细胞中 Arp2/3 复合物被原肌球蛋白-1 拮抗似乎通过平衡依赖于 Arp2/3 复合物和独立的肌动蛋白组装途径来维持肌动蛋白的动态平衡。
