The Drosophila Arf1 homologue Arf79F is essential for lamellipodium formation
The Drosophila Arf1 homologue Arf79F is essential for lamellipodium formation
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DOI：
10.1242/jcs.108092
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发表时间：
2012-12-01
影响因子：
4
通讯作者：
Koronakis, Vassilis
中科院分区：
文献类型：
--
作者：
Humphreys, Daniel;Liu, Tao;Koronakis, Vassilis
WAVE调节复合物（WRC）驱动位于质膜下的肌动蛋白丝聚合，以产生对细胞结构和运动至关重要的片状伪足。通过在膜上重建WRC依赖的肌动蛋白组装，我们最近发现几类Arf家族GTP酶直接招募和激活细胞提取物中的WRC，并且Arf与Rac1合作触发肌动蛋白聚合。在这里，我们证明了1类Arf1同源Arf79F与WRC在动态板状伪足共定位。我们报告说，Arf79F是所需的板状伪足形成在果蝇S2R+细胞，只表达一个Arf亚型的每一类。通过显性负性Arf表达或通过Arf双链RNA干扰（dsRNAi）介导的敲低来阻碍Arf功能，揭示了Arf依赖性板状伪足形成对Arf 79F是特异性的，从而确立了1类Arf，而不是2类或3类Arf，对板状伪足是至关重要的。在Arf79F沉默的细胞中，通过表达哺乳动物Arf 1而不是通过组成型活性Rac 1来恢复板状伪足的形成，表明Arf 79F不通过Rac 1起作用。取消板状伪足形成Arf79F沉默的细胞是不是由于高尔基体破坏。用鸟嘌呤核苷酸交换因子抑制剂阻断Arf79F活化损害了WRC定位于质膜和伴随的片状伪足的产生。我们的数据表明，I类Arf GTdR是WRC驱动的板状伪足形成的核心组成部分。
The WAVE regulatory complex (WRC) drives the polymerisation of actin filaments located beneath the plasma membrane to generate lamellipodia that are pivotal to cell architecture and movement. By reconstituting WRC-dependent actin assembly at the membrane, we recently discovered that several classes of Arf family GTPases directly recruit and activate WRC in cell extracts, and that Arf cooperates with Rac1 to trigger actin polymerisation. Here, we demonstrate that the Class 1 Arf1 homologue Arf79F colocalises with the WRC at dynamic lamellipodia. We report that Arf79F is required for lamellipodium formation in Drosophila S2R+ cells, which only express one Arf isoform for each class. Impeding Arf function either by dominant-negative Arf expression or by Arf double-stranded RNA interference (dsRNAi)-mediated knockdown uncovered that Arf-dependent lamellipodium formation was specific to Arf79F, establishing that Class 1 Arfs, but not Class 2 or Class 3 Arfs, are crucial for lamellipodia. Lamellipodium formation in Arf79F-silenced cells was restored by expressing mammalian Arf1, but not by constitutively active Rac1, showing that Arf79F does not act via Rac1. Abolition of lamellipodium formation in Arf79F-silenced cells was not due to Golgi disruption. Blocking Arf79F activation with guanine nucleotide exchange factor inhibitors impaired WRC localisation to the plasma membrane and concomitant generation of lamellipodia. Our data indicate that the Class I Arf GTPase is a central component in WRC-driven lamellipodium formation.