Functional analysis of protein kinase C(PKC), a molecule involved in plasticity, development and cell death in central nervous system, using gene-manipulated mice.

使用基因操作小鼠对蛋白激酶 C (PKC)（一种参与中枢神经系统可塑性、发育和细胞死亡的分子）进行功能分析。

• 批准号: 16500243
• 负责人: SAKAI Norio
• 金额: $ 2.3万
• 依托单位: HIROSHIMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16500243/
• 关键词: protein phosphorylation; protein kinase C; Purkinje cell; transgenic mice; Propagation on neuronal excitation

1)We developed brain region specific and inducible transgenic mice expressing gammaPKC-GFP using tet-regulated system. In these mice, gammaPKC-GFP was abundantly expressed in Purkinje cells. We prepared the cerebellar slices from these mice and attempted to observe gammaPKC-GFP translocation in living Purkinje cells. Trans-synaptic stimulation of parallel fiber triggered gammaPKC-GFP translocation along the dendritic shafts. This translocation propagated from peripheral to soma of the cells, indicating that neural activity can control gammaPKC translocation. This phenomenon provides the novel insight for developing the methods to monitor the neural activity in a protein level.2)Recently it was reported that the hereditary spinocerebellar ataxia type 14 (SCA14) is caused by missense mutations of gammaPKC. To elucidate the molecular mechanism underlying the SCA14 pathogenesis, we introduced SCA14 mutations into human gammaPKC cDNA, fused it with GFP cDNA and expressed mutant gammaPKC-GFP in CHO cells. The aggregation of mutant gammaPKC was obviously seen in the expressing cells. Moreover, gammaPKC-GFP was frequently aggregated during the process of its receptor-mediated translocation. These results suggest that the aggregate formation of mutant gammaPKC is involved in the SCA14 pathogenesis.3)We performed PCR-based screening of genomic DNA prepared from undefined Japanese SCA patients. We found new Japanese SCA14 family and proved that SCA14 is very rare in Japanese SCA population.

1)我们使用tet调节系统开发了表达gammaPKC-GFP的脑区域特异性和可诱导转基因小鼠。在这些小鼠中，gammaPKC-GFP 在浦肯野细胞中大量表达。我们制备了这些小鼠的小脑切片，并尝试观察活浦肯野细胞中的 gammaPKC-GFP 易位。平行纤维的跨突触刺激触发了沿树突轴的 gammaPKC-GFP 易位。这种易位从细胞外周传播到细胞体，表明神经活动可以控制 gammaPKC 易位。这一现象为开发在蛋白质水平上监测神经活动的方法提供了新的见解。2)最近报道遗传性脊髓小脑共济失调14型(SCA14)是由γPKC错义突变引起的。为了阐明 SCA14 发病机制的分子机制，我们将 SCA14 突变引入人 gammaPKC cDNA 中，将其与 GFP cDNA 融合，并在 CHO 细胞中表达突变的 gammaPKC-GFP。在表达细胞中明显观察到突变型γPKC的聚集。此外，gammaPKC-GFP在其受体介导的易位过程中频繁聚集。这些结果表明，突变型γPKC的聚集形成参与了SCA14的发病机制。3)我们对从未明确的日本SCA患者制备的基因组DNA进行了基于PCR的筛查。我们发现了新的日本SCA14家族，并证明SCA14在日本SCA人群中非常罕见。

项目成果

Superoxide production at phacosomal cup/phagosime through b1 protein kinase C during DcgR-mediated phagocutosis in microglia.

在小胶质细胞 DcgR 介导的吞噬过程中，通过 b1 蛋白激酶 C 在吞噬体杯/吞噬体中产生超氧化物。

• 发表时间: 2004
• 期刊: J.Immunol. 173
• 作者: Ueyama;T.
• 通讯作者: T.

Ceramide-induced apoptosis by translocation, phosphorylation and activation of protein kinase Cdelta at Golgi complex.

神经酰胺通过高尔基复合体蛋白激酶 Cdelta 的易位、磷酸化和激活诱导细胞凋亡。

• 发表时间: 2004
• 期刊: Journal of Biological Chemistry 279・13
• 作者: Kajimoto;T.
• 通讯作者: T.

Ceramide-induced apoptosis by translocation, phosphorylation and activation of protein kinase Cδ at Golgi complex.

神经酰胺通过高尔基复合体蛋白激酶 Cδ 的易位、磷酸化和激活诱导细胞凋亡。

• 发表时间: 2004
• 期刊: J.Biol.Chem. 279
• 作者: Kajimoto;T.
• 通讯作者: T.

Embryonic stem cel1-derived neuron models of Parkinson's disease exhibit delayed neuronal death.

帕金森病的胚胎干细胞衍生的神经元模型表现出延迟的神经元死亡。

• 发表时间: 2006
• 期刊: J. Neurochem. (in press)
• 作者: Yamashita;H.
• 通讯作者: H.

Mutant protein kinase C gamma found in spinocerebellar ataia type 14 is susceptible to aggregation and causes cell death.

14 型脊髓小脑 ataia 中发现的突变蛋白激酶 C gamma 易于聚集并导致细胞死亡。

• 发表时间: 2005
• 期刊: J.Biol.Chem. 280
• 作者: Seki;T.
• 通讯作者: T.