Molecular Cell Biology Division, Kanagawa Cancer Center Research Institute, Yokohama 241-0815, Japan.
Cell Signal. 2010 Mar;22(3):510-8. doi: 10.1016/j.cellsig.2009.11.005.
Membrane targeting of WAVE2 along microtubules is mediated by a motor protein kinesin and requires Pak1, a downstream effector of Rac1. However, the mechanism by which WAVE2 targeting to the leading edge is directionally controlled remains largely unknown. Here we demonstrate that EB1, a microtubule plus-end-binding protein, constitutively associates with stathmin, a microtubule-destabilizing protein, in human breast cancer cells. Stimulation of the cells with insulin-like growth factor I (IGF-I) induced Pak1-dependent binding of the EB1-stathmin complex to microtubules that bear WAVE2 and colocalization of the complex with WAVE2 at the leading edge. Depletion of EB1 by small interfering RNA (siRNA) abrogated the IGF-I-induced WAVE2 targeting and stathmin binding to microtubules. On the other hand, chemotaxis chamber assays indicated that the IGF-I receptor (IGF-IR) was locally activated in the region facing toward IGF-I. In addition, IGF-I caused phosphatidylinositol 3-kinase (PI 3-kinase)-dependent production of phosphatidylinositol 3,4,5-triphosphate (PIP3) near activated IGF-IR and WAVE2 colocalization with it. Collectively, WAVE2-membrane targeting is directionally controlled by binding of the EB1-stathmin complex to WAVE2-bearing microtubules and by the interaction between WAVE2 and PIP3 produced near IGF-IR that is locally activated by IGF-I.
WAVE2 沿着微管向细胞膜的靶向运输是由运动蛋白驱动蛋白介导的，并且需要 Rac1 的下游效应物 Pak1。然而，WAVE2 靶向前沿的定向控制机制在很大程度上仍然未知。在这里，我们证明了微管末端结合蛋白 EB1 与微管不稳定蛋白 stathmin 在人乳腺癌细胞中持续结合。用胰岛素样生长因子 I（IGF-I）刺激细胞会诱导 Pak1 依赖性 EB1-stathmin 复合物与承载 WAVE2 的微管结合，并且该复合物与 WAVE2 在前沿处共定位。用小干扰 RNA（siRNA）耗尽 EB1 会破坏 IGF-I 诱导的 WAVE2 靶向和 stathmin 与微管的结合。另一方面，趋化性室测定表明，IGF-IR 在面向 IGF-I 的区域局部激活。此外，IGF-I 导致在活化的 IGF-IR 附近产生磷脂酰肌醇 3-激酶（PI 3-kinase）依赖性的磷脂酰肌醇 3,4,5-三磷酸（PIP3），并且 WAVE2 与它共定位。总之，WAVE2 向膜的靶向运输是通过 EB1-stathmin 复合物与承载 WAVE2 的微管的结合以及 WAVE2 与局部由 IGF-I 激活的 IGF-IR 附近产生的 PIP3 之间的相互作用来定向控制的。
