GluRδ2 Assembles Four Neurexins into Trans-Synaptic Triad to Trigger Synapse Formation
GluRδ2 Assembles Four Neurexins into Trans-Synaptic Triad to Trigger Synapse Formation
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DOI：
10.1523/jneurosci.5584-11.2012
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发表时间：
2012-03-28
影响因子：
5.3
通讯作者：
Mishina, Masayoshi
中科院分区：
文献类型：
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作者：
Lee, Sung-Jin;Uemura, Takeshi;Mishina, Masayoshi
阐明突触形成的分子机制是理解神经线路、高级大脑功能和精神障碍的先决条件。突触后谷氨酸受体 delta 2 (GluR delta 2) 和突触前神经毒素 (NRXN) 通过小脑蛋白前体蛋白 1 (Cbln1) 的跨突触相互作用介导小脑体内突触形成。在这里，我们询问跨突触三联体如何诱导突触形成。天然 GluR delta 2 作为四聚体存在于膜中，而 GluR delta 2 的 N 末端结构域 (NTD) 形成稳定的同二聚体。当与培养的小鼠小脑颗粒细胞 (GC) 一起孵育时，二聚体 GluR delta 2-NTD 和 Cbln1 对 GC 轴突中巴松管和囊泡谷氨酸转运蛋白 1 点状免疫染色信号的积累几乎没有影响。然而，四聚化的 GluR delta 2-NTD 刺激了这些突触前蛋白在轴突中的积累。对 Cbln1 突变体的分析表明，Cbln1 上的 GluR delta 2 和 NRXN1 beta 的结合位点是不同的。此外，GluR delta 2-NTD 和 NRXN1 beta 的胞外结构域 (ECD) 之间不存在与 Cbln1 的竞争。因此，GluR delta 2 和 Cbln1 彼此相互作用，而与 Cbln1-NRXN1 beta 相互作用无关，反之亦然。凝胶过滤和等温滴定量热分析一致表明，二聚体 GluR delta 2-NTD 和六聚体 Cbln1 以 1:1 的比例组装，而六聚体 Cbln1 和 NRXN1 beta -ECD 的层粘连蛋白-神经释放蛋白-性激素结合球蛋白结构域以 1:2 的比例组装。因此，突触发生三联体由四聚体 GluR delta 2、六聚体 Cbln1 和单体 NRXN 以 1:2:4 的比例组装而成。这些结果表明，GluR delta 2 通过三联体形成聚集四个 NRXN 来触发突触形成。
Elucidation of molecular mechanisms of synapse formation is a prerequisite for the understanding of neural wiring, higher brain functions, and mental disorders. The trans-synaptic interaction of postsynaptic glutamate receptor delta 2 (GluR delta 2) and presynaptic neurexins (NRXNs) through cerebellin precursor protein 1 (Cbln1) mediates synapse formation in vivo in the cerebellum. Here, we asked how the trans-synaptic triad induces synapse formation. Native GluR delta 2 existed as a tetramer in the membrane, whereas the N-terminal domain (NTD) of GluR delta 2 formed a stable homodimer. When incubated with cultured mouse cerebellar granule cells (GCs), dimeric GluR delta 2-NTDand Cbln1 exerted little effect on the accumulation of punctate immunostaining signals for Bassoon and vesicular glutamate transporter 1 in GC axons. However, tetramerized GluR delta 2-NTD stimulated the accumulation of these presynaptic proteins in the axons. Analysis of Cbln1 mutants suggested that the binding sites of GluR delta 2 and NRXN1 beta on Cbln1 are differential. Furthermore, there was no competition in the binding to Cbln1 between GluR delta 2-NTD and the extracellular domain (ECD) of NRXN1 beta. Thus, GluR delta 2 and Cbln1 interacted with each other rather independently of Cbln1-NRXN1 beta interaction and vice versa. Gel filtration and isothermal titration calorimetry analyses consistently showed that dimeric GluR delta 2-NTD and hexameric Cbln1 assembled in the 1:1 ratio, whereas hexameric Cbln1 and the laminin-neurexin-sex hormone-binding globulin domain of NRXN1 beta -ECD assembled in the 1:2 ratio. Thus, the synaptogenic triad is assembled from tetrameric GluR delta 2, hexameric Cbln1, and monomeric NRXN in the ratio of 1:2:4. These results suggest that GluR delta 2 triggers synapse formation by clustering four NRXNs through triad formation.