Local phosphatidylinositol 3,4,5-trisphosphate accumulation recruits Vav2 and Vav3 to activate Rac1/Cdc42 and initiate neurite outgrowth in nerve growth factor-stimulated PC12 cells
Local phosphatidylinositol 3,4,5-trisphosphate accumulation recruits Vav2 and Vav3 to activate Rac1/Cdc42 and initiate neurite outgrowth in nerve growth factor-stimulated PC12 cells
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DOI：
10.1091/mbc.e04-10-0904
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发表时间：
2005-05-01
影响因子：
3.3
通讯作者：
Matsuda, M
中科院分区：
文献类型：
--
作者：
Aoki, K;Nakamura, T;Matsuda, M
轴突生长是神经元网络形成的重要过程。Rho家族的Rho家族成员rac1和CDc42通过重组肌动蛋白细胞骨架，对突起的延伸起到积极的调节作用。在这里，我们研究了在神经生长因子(NGF)诱导的PC12细胞突起生长过程中，rac1/CDc42和磷脂酰肌醇3-激酶(P13-Kinase)之间的动态联系。应用荧光共振能量转移探针的活性成像显示，在加入NGF后，P13-激酶和rac1/CDc42在细胞外围的广泛区域瞬间被激活。随后，在突起部位观察到P13-激酶和rac1/CDc42的局部和重复激活。RNA干扰使Vav2和Vav3的缺失显著抑制了rac1/CDc42的激活和导致轴突生长的短突起的形成。在NGF诱导的突起中，局部3，4，5-三磷酸磷脂酰肌醇的聚集招募了Vav2和Vav3来激活rac1和CDc42，反之，P13-激酶的局部激活则需要Vav2和Vav3。这些观察结果首次证明，Vav2和Vav3是由P13-激酶和rac1/CDc42组成的正反馈环的重要组成部分，并伴随着形态变化而局部循环。
Neurite outgrowth is an important process in the formation of neuronal networks. Rac1 and Cdc42, members of the Rho-family GTPases, positively regulate neurite extension through reorganization of the actin cytoskeleton. Here, we examine the dynamic linkage between Rac1/Cdc42 and phosphatidylinositol 3-kinase (P13-kinase) during nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Activity imaging using fluorescence resonance energy transfer probes showed that P13-kinase as well as Rac1/Cdc42 was transiently activated in broad areas of the cell periphery immediately after NGF addition. Subsequently, local and repetitive activation of P13-kinase and Rac1/Cdc42 was observed at the protruding sites. Depletion of Vav2 and Vav3 by RNA interference significantly inhibited both Rac1/Cdc42 activation and the formation of short processes leading to neurite outgrowth. At the NGF-induced protrusions, local phosphatidylinositol 3,4,5-trisphosphate accumulation recruited Vav2 and Vav3 to activate Rac1 and Cdc42, and conversely, Vav2 and Vav3 were required for the local activation of P13-kinase. These observations demonstrated for the first time that Vav2 and Vav3 are essential constituents of the positive feedback loop that is comprised of P13-kinase and Rac1/Cdc42 and cycles locally with morphological changes.