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Mechanism of accelerating death of newborn neurons by ss-amyloid

ss-淀粉样蛋白加速新生神经元死亡的机制

基本信息

批准号：
22500347
负责人：
UCHIDA Yoko
金额：
$ 3.08万
依托单位：
Tokyo Metropolitan Geriatric Hospital and Institute of Gerontology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2010
资助国家：
日本
起止时间：
2010 至 2012
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-22500347/
关键词：
β-アミロイド神経新生

项目摘要

In Alzheimer's disease (AD), immature neuronal marker proteins increased in brain. However, the increased neurogenesis is not sufficient to repair the nervous system because progressive neuronal loss occurs in AD brain. Alteration of microenvironment in AD brain may affect the fate of immature neurons. Ss-amyloid (Ass) plays an important role in the early pathogenesis of AD. It is reasonable to speculate Ass alters the brain microenvironment to make it toxic to newborn neurons. To address these issues, we tried to identify the genes to inhibit survival of newborn neurons in Ass-induced genes. Among 7 Ass-induced genes, only Chat (Cas/HEF1-associated signal transducer) accelerated death of newborn neurons. Other 6 genes promoted neuronal survival or did not affect on neuronal survival. Next, we focused on Chat gene and tried to clarify toxic mechanism to newborn neurons. The results are follows :(1) C-terminal portion of Chat is necessary to induce toxicity in newborn neurons.(2) C-terminal portion of Chat have the binding domain to Cas protein family. However, co-transfection of Cas proteins (p130Cas and NEDD9) and Chat into newborn neurons did not accelerate neuronal death. The mutant Chat, which does not bind to Cas proteins, also accelerated neuronal death. These results suggests that the binding of unknown proteins to Chat C-terminal portion (Chat C) other than Cas binding site might be necessary to induce toxicity in newborn neurons. Finally we tried to identify the proteins that bind to Chat C and accelerate death of newborn neurons. A GST-Chat C fusion protein was constructed and purified. The lysate from cultured cortical neurons was incubated with a GST-Chat C fusion protein. Pull downed proteins were subjected to SDS-PAGE and analyzed with nanoLC-MS/MS. We could not identify any binding proteins to GST-Chat C fusion protein. It could not rule out that homopolymer formation of Chat C would induce neurotoxicity to newborn neurons.

在阿尔茨海默病 (AD) 中，大脑中不成熟的神经元标记蛋白增加。然而，增加的神经发生不足以修复神经系统，因为AD大脑中发生进行性神经元损失。 AD大脑微环境的改变可能会影响未成熟神经元的命运。 Ss-淀粉样蛋白（Ass）在 AD 的早期发病机制中发挥着重要作用。有理由推测 Ass 改变了大脑微环境，使其对新生神经元有毒。为了解决这些问题，我们试图找出抑制 Ass 诱导基因中新生神经元存活的基因。在 7 个 Ass 诱导基因中，只有 Chat（Cas/HEF1 相关信号转导器）加速新生神经元的死亡。其他6个基因促进神经元存活或不影响神经元存活。接下来，我们关注Chat基因，试图阐明对新生神经元的毒性机制。结果如下：(1)Chat的C端部分是诱导新生神经元毒性所必需的。(2)Chat的C端部分具有Cas蛋白家族的结合域。然而，将 Cas 蛋白（p130Cas 和 NEDD9）和 Chat 共转染到新生神经元中并不会加速神经元死亡。 Chat 突变体不与 Cas 蛋白结合，也会加速神经元死亡。这些结果表明，除了 Cas 结合位点之外，未知蛋白与 Chat C 末端部分（Chat C）的结合可能是诱导新生神经元毒性所必需的。最后，我们试图识别与 Chat C 结合并加速新生神经元死亡的蛋白质。构建并纯化了 GST-Chat C 融合蛋白。将培养的皮质神经元的裂解物与 GST-Chat C 融合蛋白一起孵育。对下拉的蛋白质进行 SDS-PAGE 并使用 nanoLC-MS/MS 进行分析。我们无法识别任何 GST-Chat C 融合蛋白的结合蛋白。不能排除Chat C的均聚物形成会对新生神经元产生神经毒性。