Liu Xiujie, Wang Yonggang, Zhang Yong, Zhu Wei, Xu Xiaohui, Niinobe Michio, Yoshikawa Kazuaki, Lu Changlin, He Cheng
Institute of Neuroscience and Key Laboratory of Molecular Neurobiology, Ministry of Education, SMMU, Shanghai 200433, China.
Mol Cell Neurosci. 2009 May;41(1):51-61. doi: 10.1016/j.mcn.2009.01.009. Epub 2009 Feb 6.
Nogo-A has been identified in the central nervous system as an inhibitor for axonal regeneration. Previous works have mainly focused on Nogo-A in oligodendrocytes and the roles of neuronal intracellular Nogo-A remain elusive. To gain deep insight into the physiological functions of Nogo-A, a yeast two-hybrid screening was performed with Nogo-66 as bait. We identified a new interaction between Nogo-66 and necdin. Mutagenesis analysis revealed that the central region of necdin was indispensable for the interaction of necdin with Nogo-66. The interaction was further confirmed by co-immunoprecipitation in neural tissues and cultured cortical neurons. Morphological evidence showed that Nogo-A and necdin highly colocalized in rat cortical and dorsal root ganglia neurons. Ectopic expression of Nogo-A in HEK293 cells led to retention of necdin from the nucleus to the cytoplasm. Furthermore, overexpression of Nogo-A in PC12 cells and cultured cortical neurons inhibited necdin-accelerated neurite outgrowth. Meanwhile, necdin was found to be significantly sequestered in the cytoplasm of PC12 cells stably overexpressing Nogo-A. Together, these data suggest that Nogo-A is a novel necdin binding protein and inhibits necdin-accelerated neuronal neurite outgrowth by sequestering necdin in the cytoplasm.
Nogo-A在中枢神经系统中被鉴定为轴突再生的抑制剂。以往的研究主要集中在少突胶质细胞中的Nogo-A，而神经元细胞内Nogo-A的作用仍不清楚。为了深入了解Nogo-A的生理功能，以Nogo-66为诱饵进行了酵母双杂交筛选。我们鉴定出Nogo-66与神经生长抑制因子之间存在新的相互作用。诱变分析表明，神经生长抑制因子的中央区域对于其与Nogo-66的相互作用是不可或缺的。通过神经组织和培养的皮质神经元中的免疫共沉淀进一步证实了这种相互作用。形态学证据表明，Nogo-A和神经生长抑制因子在大鼠皮质和背根神经节神经元中高度共定位。Nogo-A在HEK293细胞中的异位表达导致神经生长抑制因子从细胞核滞留到细胞质中。此外，Nogo-A在PC12细胞和培养的皮质神经元中的过表达抑制了神经生长抑制因子促进的神经突生长。同时，发现神经生长抑制因子在稳定过表达Nogo-A的PC12细胞的细胞质中被显著隔离。总之，这些数据表明Nogo-A是一种新型的神经生长抑制因子结合蛋白，并通过将神经生长抑制因子隔离在细胞质中来抑制神经生长抑制因子促进的神经元神经突生长。
