The Toca-1-N-WASP Complex Links Filopodial Formation to Endocytosis
The Toca-1-N-WASP Complex Links Filopodial Formation to Endocytosis
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DOI：
10.1074/jbc.m805940200
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发表时间：
2009-04-24
影响因子：
4.8
通讯作者：
Ahmed, Sohail
中科院分区：
文献类型：
--
作者：
Bu, Wenyu;Chou, Ai Mei;Ahmed, Sohail
作为体外Cdc42驱动的肌动蛋白聚合的关键辅因子，Cdc42依赖性肌动蛋白组装的转导蛋白（Toca - 1）-N - WASP复合物被分离出来。Toca - 1包含一个N端F - BAR结构域，接着是一个Cdc42结合位点（HR1结构域）和一个SH3结构域（N - WASP相互作用位点）。N - WASP是通过Arp2/3复合物激活肌动蛋白成核的因子。本研究的目的是探究Toca - 1 - N - WASP复合物的细胞功能。我们报道在N1E115神经母细胞瘤细胞中，Toca - 1和N - WASP一样可诱导丝状伪足和轴突。Toca - 1需要F - BAR结构域、Cdc42结合位点和SH3结构域来诱导丝状伪足。Toca - 1和N - WASP相互需要对方来诱导丝状伪足。Toca - 1和N - WASP的表达影响Rab5阳性膜的分布、大小和数量。通过福斯特共振能量转移可见，Toca - 1在丝状伪足和Rab5膜中与N - WASP直接相互作用。因此，Toca - 1 - N - WASP复合物定位于丝状伪足和内吞小泡并诱导其形成。最后，三种内吞作用抑制剂，即动力蛋白 - K44A、Eps15 Δ95/295和网格蛋白重链RNA干扰，可阻断Toca - 1诱导的丝状伪足形成。综上所述，这些数据表明Toca - 1 - N - WASP复合物可将丝状伪足形成与内吞作用联系起来。
The transducer of Cdc42-dependent actin assembly (Toca-1)-N-WASP complex was isolated as an essential cofactor for Cdc42-driven actin polymerization in vitro. Toca-1 consists of an N-terminal F-BAR domain, followed by a Cdc42 binding site (HR1 domain) and an SH3 domain, (the N-WASP interacting site). N-WASP is an activator of actin nucleation through the Arp2/3 complex. The aim of the present study was to investigate the cellular function of the Toca-1-N-WASP complex. We report that Toca-1 induces filopodia and neurites as does N-WASP in N1E115 neuroblastoma cells. Toca-1 requires the F-BAR domain, Cdc42 binding site, and SH3 domain to induce filopodia. Toca-1 and N-WASP both require each other to induce filopodia. The expression of Toca-1 and N-WASP affects the distribution, size, and number of Rab5 positive membranes. Toca-1 interacts directly with N-WASP in filopodia and Rab5 membrane as seen by Forster resonance energy transfer. Thus the Toca-1-N-WASP complex localizes to and induces the formation of filopodia and endocytic vesicles. Last, three inhibitors of endocytosis, Dynamin-K44A, Eps15 Delta 95/295, and clathrin heavy chain RNA interference, block Toca-1-induced filopodial formation. Taken together, these data suggest that the Toca-1-N-WASP complex can link filopodial formation to endocytosis.