Neurofibromatosis type 1 (NF1) tumor suppressor, neurofibromin, regulates the neuronal differentiation of PC12 cells via its associating protein, CRMP-2
Neurofibromatosis type 1 (NF1) tumor suppressor, neurofibromin, regulates the neuronal differentiation of PC12 cells via its associating protein, CRMP-2
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DOI：
10.1074/jbc.m708206200
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发表时间：
2008-04-04
影响因子：
4.8
通讯作者：
Araki, Norie
中科院分区：
文献类型：
--
作者：
Patrakitkomjorn, Siriporn;Kobayashi, Daiki;Araki, Norie
1 型神经纤维瘤病 (NF1) 肿瘤抑制基因产物神经纤维蛋白的部分功能是作为 Ras-GAP（Ras 的负调节因子）。神经纤维蛋白与 NF1 患者的神经元异常有关；然而，神经纤维蛋白的精确细胞功能尚未阐明。使用蛋白质组学策略，我们鉴定了一组神经纤维蛋白相关细胞蛋白，包括轴突调节因子 CRMP-2（塌陷反应介质蛋白-2）。 CRMP-2 直接与神经纤维蛋白的 C 端结构域结合，并且这种结合受到 CRMP-2 磷酸化方式的调节。在神经生长因子刺激的 PC12 细胞中，神经纤维蛋白和 CRMP-2 共同定位，特别是在延伸神经突的远端和分支上。使用 NF1 小干扰 RNA 抑制神经纤维蛋白可显着抑制这种神经突生长，并通过 CDK5、GSK-3b 和 Rho 激酶上调一系列 CRMP-2 磷酸化。 NF1-RAS-GAP 相关结构域的过度表达挽救了这些 NF1 小干扰 RNA 诱导的事件。我们的结果表明，神经纤维蛋白通过发挥一种或多种互补作用来调节神经元分化。首先，神经纤维蛋白通过复合物的形成直接调节 CRMP-2 磷酸化的可及性。此外，神经纤维蛋白似乎通过其 Ras-GAP 功能抑制 CRMP-2 磷酸化激酶级联，从而间接调节 CRMP-2 活性。我们的研究表明，神经纤维蛋白和 CRMP-2 的功能关联对于神经细胞分化至关重要，神经纤维蛋白表达缺乏或异常调节可能导致神经细胞功能受损，这可能是 NF1 相关发病机制的一个因素。
Neurofibromatosis type 1 (NF1) tumor suppressor gene product, neurofibromin, functions in part as a Ras-GAP, a negative regulator of Ras. Neurofibromin is implicated in the neuronal abnormality of NF1 patients; however, the precise cellular function of neurofibromin has yet to be clarified. Using proteomic strategies, we identified a set of neurofibromin-associating cellular proteins, including axon regulator CRMP-2 (Collapsin response mediator protein-2). CRMP-2 directly bound to the C-terminal domain of neurofibromin, and this association was regulated by the manner of CRMP-2 phosphorylation. In nerve growth factor-stimulated PC12 cells, neurofibromin and CRMP-2 co-localized particularly on the distal tips and branches of extended neurites. Suppression of neurofibromin using NF1 small interfering RNA significantly inhibited this neurite outgrowth and up-regulated a series of CRMP-2 phosphorylations by kinases identified as CDK5, GSK-3b, and Rho kinase. Overexpression of the NF1-RAS-GAP-related domain rescued these NF1 small interfering RNA-induced events. Our results suggest that neurofibromin regulates neuronal differentiation by performing one or more complementary roles. First, neurofibromin directly regulates CRMP-2 phosphorylation accessibility through the complex formation. Also, neurofibromin appears to indirectly regulate CRMP-2 activity by suppressing CRMP-2-phosphorylating kinase cascades via its Ras-GAP function. Our study demonstrates that the functional association of neurofibromin and CRMP-2 is essential for neuronal cell differentiation and that lack of expression or abnormal regulation of neurofibromin can result in impaired function of neuronal cells, which is likely a factor in NF1-related pathogenesis.