Aggregate formation of mutant protein kinase C gamma found in spinocerebellar ataxia type 14 impairs ubiquitin-proteasome system and induces endoplasmic reticulum stress
Aggregate formation of mutant protein kinase C gamma found in spinocerebellar ataxia type 14 impairs ubiquitin-proteasome system and induces endoplasmic reticulum stress
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DOI：
10.1111/j.1460-9568.2007.05933.x
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发表时间：
2007-12-01
影响因子：
3.4
通讯作者：
Sakai, Norio
中科院分区：
文献类型：
--
作者：
Seki, Takahiro;Takahashi, Hideyuki;Sakai, Norio
在脊髓小脑性共济失调14型（SCA 14）中发现了蛋白激酶C γ（gamma PKC）基因的几个致病性错义突变。我们以前证明，突变体γ PKC SCA 14中发现的是两种类型的聚集，细胞质点样和核周大量聚集，并导致细胞死亡，在中国仓鼠卵巢细胞。长时程成像显示，突变体γ PKC-绿色荧光蛋白（GFP）首先聚集成点状聚集体，逐渐形成核周块状聚集体，随后细胞死亡。然而，目前仍不清楚突变型γ PKC的聚集体形成如何导致细胞死亡。在本研究中，我们研究了这些突变体聚集是否影响泛素-蛋白酶体系统（UPS）和内质网（ER）的压力。两个突变体γ PKC-GFPs（S119 P和G128 D）被强烈泛素化，这些突变体的点样聚集体是泛素阳性的，并与蛋白酶体20 S共定位。此外，蛋白酶体活性在细胞聚集，特别是大规模的，显着下降。突变体γ PKC-GFP的聚集体形成诱导PERK（PKR样ER激酶）的磷酸化和CHOP（C/EBP同源蛋白）的核表达，这是ER应激的标志，随后激活半胱天冬酶-3。这些结果表明，在SCA 14中发现的突变体γ PKC的聚集体形成损害UPS并诱导ER应激，导致凋亡性细胞死亡。
Several causal missense mutations in protein kinase C gamma (gamma PKC) gene have been found in spinocerebellar ataxia type 14 (SCA14), an autosomal dominant neurodegenerative disease. We previously demonstrated that mutant gamma PKC found in SCA14 is susceptible to two types of aggregation, cytoplasmic dot-like and perinuclear massive aggregation, and causes cell death in Chinese hamster ovary cells. Long-term time-lapse imaging revealed that firstly accumulated dot-like aggregation of mutant gamma PKC-green fluorescent protein (GFP) gradually formed perinuclear massive aggregations, followed by cell death. However, it remains unclear how aggregate formation of mutant gamma PKC causes cell death. In the present study, we examined whether these mutant aggregations affect the ubiquitin-proteasome system (UPS) and endoplasmic reticular (ER) stress. Two mutant gamma PKC-GFPs (S119P and G128D) were strongly ubiquitinated, and dot-like aggregations of these mutants were ubiquitin-positive and colocalized with proteasome 20S. Furthermore, proteasome activity in cells with aggregates, especially massive ones, was significantly decreased. Aggregate formation of mutant gamma PKC-GFP induced phosphorylation of PERK (PKR-like ER kinase) and nuclear expression of CHOP (C/EBP homologous protein), hallmarks of ER stress and subsequently activated caspase-3. These results indicate that aggregate formation of mutant gamma PKC found in SCA14 impairs UPS and induces ER stress, leading to apoptotic cell death.