Wnt Signalling Promotes Actin Dynamics during Axon Remodelling through the Actin-Binding Protein Eps8.
Wnt Signalling Promotes Actin Dynamics during Axon Remodelling through the Actin-Binding Protein Eps8.
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DOI：
10.1371/journal.pone.0134976
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Salinas PC
中科院分区：
文献类型：
--
作者：
Stamatakou E;Hoyos-Flight M;Salinas PC
在到达它们的突触目标后，轴突在突触前终扣组装之前减慢它们的生长并广泛地重塑。Wnt蛋白是靶源性分泌因子，促进轴突重塑和突触组装。在发育中的脊髓中，运动神经元分泌的Wnt促进NT-3反应性背根神经节神经元的轴突重塑。由Wnts诱导的轴突重塑的特征在于生长锥暂停和扩大，这些过程依赖于微管的重组。然而，肌动蛋白细胞骨架的贡献仍然没有探索。在这里，我们证明了Wnt 3a通过支架蛋白Dishevelled-1（Dvl 1）和丝氨酸-苏氨酸激酶Gsk 3 β快速诱导啮齿动物DRG神经元生长锥中的F-actin积累来调节肌动蛋白细胞骨架。重要的是，肌动蛋白细胞骨架的这些变化发生在生长锥明显扩大之前。延时成像显示Wnt 3a增加了板层突起和丝状伪足速度。此外，肌动蛋白组装的药理学抑制表明Wnt 3a增加肌动蛋白动力学。通过酵母双杂交筛选，我们确定肌动蛋白结合蛋白Eps 8作为Dvl 1的直接相互作用物，Dvl 1是Wnt信号通路的关键支架蛋白。Eps 8功能的获得模拟Wnt介导的轴突重塑，而Eps 8沉默阻断Wnt 3a的轴突重塑活性。重要的是，阻断Dvl 1-Eps 8相互作用完全消除了Wnt 3a介导的轴突重塑。这些发现表明，Wnt-Dvl 1信号通过Eps 8在轴突重塑的调节中的新作用。
Upon arrival at their synaptic targets, axons slow down their growth and extensively remodel before the assembly of presynaptic boutons. Wnt proteins are target-derived secreted factors that promote axonal remodelling and synaptic assembly. In the developing spinal cord, Wnts secreted by motor neurons promote axonal remodelling of NT-3 responsive dorsal root ganglia neurons. Axon remodelling induced by Wnts is characterised by growth cone pausing and enlargement, processes that depend on the re-organisation of microtubules. However, the contribution of the actin cytoskeleton has remained unexplored. Here, we demonstrate that Wnt3a regulates the actin cytoskeleton by rapidly inducing F-actin accumulation in growth cones from rodent DRG neurons through the scaffold protein Dishevelled-1 (Dvl1) and the serine-threonine kinase Gsk3β. Importantly, these changes in actin cytoskeleton occurs before enlargement of the growth cones is evident. Time-lapse imaging shows that Wnt3a increases lamellar protrusion and filopodia velocity. In addition, pharmacological inhibition of actin assembly demonstrates that Wnt3a increases actin dynamics. Through a yeast-two hybrid screen, we identified the actin-binding protein Eps8 as a direct interactor of Dvl1, a scaffold protein crucial for the Wnt signalling pathway. Gain of function of Eps8 mimics Wnt-mediated axon remodelling, whereas Eps8 silencing blocks the axon remodelling activity of Wnt3a. Importantly, blockade of the Dvl1-Eps8 interaction completely abolishes Wnt3a-mediated axonal remodelling. These findings demonstrate a novel role for Wnt-Dvl1 signalling through Eps8 in the regulation of axonal remodeling.