Analysis of a RhoGEF, KIAA380, in neurite retraction

RhoGEF、KIAA380 在神经突回缩中的分析

• 批准号: 12680646
• 负责人: NAGATA Koh-ichi
• 金额: $ 2.5万
• 依托单位: AICHI CANCER CENTER
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12680646/
• 关键词: Rho; RhoGEF; Neurite retraction; KIAA380; Plexin B1; 神経実起; アクチン; グアニンヌクレオチド交換因子

Semaphorins are repulsive guidance cues that regulate cell migration in vivo and in cultured neurons lead to growth cone collapse in vitro. Growth cone collapse is characterised by a disappearance offilopodial and lamellipodial actin structures in the growth cone. Small GTPases of the Rho family regulate the actin cytoskeleton in response to extracellular stimuli. Previously, we had found evidence for an effect ofa Rho-specific GEF, KIAA0380, on neurite retraction of neuronal culture cells, Neuro2A. Then we carried out yeast two hybrid screeing for binding partners of KIAA0380.Consequently, we isolated plexin-B1 as a KIAA380-binding protein. By further analyses, we determined thatthe PDZ domain ofKIAA380 bind with the C terminus ofplexin B1. We next described that multimerisation of plexin-B1 led to activation of Rho, as observed by stress fiber formation. Interestingly a mutant KIAA0380 which lacks DH domain effectively inhibited the multimerization dependent Rho activation in Swiss 3T3 cells.We decided to study furtherthe mechanism bywhich plexins cause collapse.We present evidence foractivation of Rho by plexins via direct association with a Rho specific guanine nucleotide exchange factor (GEF), KIAA0380. The role of this RhoGEF in the growth cone collapse induced by semaphorins is currently being studied.

信号素是一种排斥性的引导信号，在体内调节细胞的迁移，在体外培养的神经元中会导致生长锥体崩溃。生长锥塌陷的特征是生长锥内的足底和板脂肌动蛋白结构消失。Rho家族的小GTP酶调节肌动蛋白细胞骨架以响应细胞外刺激。在此之前，我们已经发现了Rho特异性的全环基金KIAA0380对神经元培养细胞Neuro2A轴突回缩的影响的证据。然后进行酵母双杂交筛选KIAA0380的结合伙伴，从而分离到与KIAA380结合的蛋白Plexin-B1。通过进一步的分析，我们确定KIAA380的PDZ结构域与丛B1的C末端结合。接下来，我们描述了Plexin-B1的多聚体导致Rho的激活，这是通过应力纤维形成观察到的。有趣的是，一个缺乏水解域的突变体KIAA0380有效地抑制了瑞士3T3细胞中依赖多聚化的Rho的激活。我们决定进一步研究丛蛋白导致崩溃的机制。我们提供了丛蛋白通过与Rho特异的鸟嘌呤核苷酸交换因子KIAA0380直接关联来激活Rho的证据。目前正在研究这种Rhogef在信号素诱导的生长锥体塌陷中的作用。

项目成果

Nakamujra, Y., Hashimoto, R., Amano, M., Nagata, K., Matsumoto, N., Goto, H., Fukusho, E., Mori, H., Kashiwagi, Y., Kudo, T., Inagaki, M., and Takeda, M.: "Localized phosphorylation of vimentin by Rho-kinase in neuroblastoma N2a cells"Genes to Cells. 5. 8

Nakamujra，Y.，桥本，R.，天野，M.，永田，K.，松本，N.，后藤，H.，福所，E.，森，H.，柏木，Y.，工藤，T.，

Togashi, H., Nagata, K., Takagishi, M., Saitoh, N., and Inagaki, M.: "Functions of a Rho-specific guanine nucletoride-exchange factor, in neurite retraction : Possible involvement of proline-rich motif of KIAA0380 in the localiztion"J. Giol. Chem.. 275. 2

Togashi, H.、Nagata, K.、Takagishi, M.、Saitoh, N. 和 Inagaki, M.：“Rho 特异性鸟嘌呤核苷交换因子在神经突收缩中的功能：可能涉及富含脯氨酸的基序

Inada, H., Nagata, K., Goto, H., and In agaki M.: "Regulation of intermediate filament dynamics : A novel approach using site-and phosphorylation state-specific antibodies"Cytoskeleton : Signalling and Cell Regulation : A Practical Approach (The practial

Inada, H.、Nagata, K.、Goto, H. 和 In agaki M.：“中间丝动力学的调节：一种使用位点和磷酸化状态特异性抗体的新方法”细胞骨架：信号传导和细胞调节：实用

Nagata, K. et al.: "A decade of site-and phophorylation state-specific anubodies"Genes to Cells. 6. 653-664 (2001)

Nagata, K. 等人：“位点和磷酸化状态特异性无抗体的十年”基因到细胞。

Nakamura, Y., et al.: "Localized phosphorylation of vimentin by Rho-kinase in neuroblastoma N2a cells"Genes to Cells. 5. 823-837 (2000)

Nakamura, Y. 等人：“神经母细胞瘤 N2a 细胞中 Rho 激酶对波形蛋白的局部磷酸化”基因到细胞。