Structural bases for CRMP function in plexin-dependent semaphorin3A signaling
Structural bases for CRMP function in plexin-dependent semaphorin3A signaling
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DOI：
10.1038/sj.emboj.7600021
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发表时间：
2004-01-14
期刊：
影响因子：
11.4
通讯作者：
Strittmatter, SM
中科院分区：
文献类型：
--
作者：
Deo, RC;Schmidt, EF;Strittmatter, SM
坍缩反应介质蛋白（CRMPs）是参与神经元分化和轴突引导的细胞质磷蛋白。CRMP2先前被证明介导Sema3A对轴突的排斥作用，并参与轴突规范。小鼠CRMP1的x射线晶体结构以2.1埃的分辨率确定，并证明CRMP1是一种形成四聚体组装的双叶“肺形”蛋白质。利用基于结构的表面暴露残留物诱变来绘制功能域。作为CRMP的快速检测方法，我们在表达受体成分Neuropilin1和PlexinA1 （NP1/PlexA1）的COS-7细胞中构建了一个重组的Sema3A信号系统。在这些细胞中，CRMP和PlexA1形成一种物理复合物，其数量被NP1减少，但被Sema3A/NP1增强。此外，CRMP加速sema3a诱导的细胞收缩。CRMP1的一个结构域的丙氨酸取代产生一个组成活性蛋白，导致不依赖sema3a的COS-7收缩。该突变体CRMP模拟了Sema3A对DRG神经突生长的抑制作用，并减少了Sema3A诱导的轴突排斥。这些数据提供了在plex依赖的Sema3A信号中CRMP功能的结构视图。
Collapsin response mediator proteins (CRMPs) are cytosolic phosphoproteins involved in neuronal differentiation and axonal guidance. CRMP2 was previously shown to mediate the repulsive effect of Sema3A on axons and to participate in axonal specification. The X-ray crystal structure of murine CRMP1 was determined at 2.1 Angstrom resolution and demonstrates that CRMP1 is a bilobed 'lung-shaped' protein forming a tetrameric assembly. Structure-based mutagenesis of surface-exposed residues was employed to map functional domains. As a rapid assay for CRMP, we exploited a reconstituted Sema3A signaling system in COS-7 cells expressing the receptor components Neuropilin1 and PlexinA1 (NP1/PlexA1). In these cells, CRMP and PlexA1 form a physical complex that is reduced in amount by NP1 but enhanced by Sema3A/NP1. Furthermore, CRMP accelerates Sema3A-induced cell contraction. Alanine substitutions in one domain of CRMP1 produce a constitutively active protein that causes Sema3A-independent COS-7 contraction. This mutant CRMP mimics the DRG neurite outgrowth-inhibiting effects of Sema3A and reduces Sema3A-induced axonal repulsion. These data provide a structural view of CRMP function in Plex-dependent Sema3A signaling.