Differential regulation of ceramide in lipid-rich microdomains (rafts): Antagonistic role of palmitoyl: Protein thioesterase and neutral sphingomyelinase 2
Differential regulation of ceramide in lipid-rich microdomains (rafts): Antagonistic role of palmitoyl: Protein thioesterase and neutral sphingomyelinase 2
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DOI：
10.1002/jnr.20549
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发表时间：
2005-07-15
影响因子：
4.2
通讯作者：
Dawson, G
中科院分区：
文献类型：
--
作者：
Goswami, R;Ahmed, M;Dawson, G
细胞分化和髓鞘形成涉及停滞和程序性细胞死亡之间的微妙平衡，但调控这一点的基因尚未明确定义。因此，我们研究了两个关键的基因产物参与少突胶质细胞质膜脂质代谢和神经酰胺介导的细胞死亡信号的拮抗作用。棕榈酰：蛋白硫酯酶（PPT 1;通过V5标记的蛋白质的蛋白质印迹和增加的酶活性验证）的过表达导致相对于胆固醇和鞘磷脂（SM）的洗涤剂抗性微结构域（DRM或筏）中的神经酰胺减少。该PPT 1过表达还导致对由星形孢菌素或C-2-神经酰胺诱导的细胞死亡的保护。相反，中性鞘磷脂酶2（NSMase 2;通过FLAG标记的蛋白质的蛋白质印迹和增加的酶活性验证）的过表达导致膜NSMase增加和相对于胆固醇和SM的筏中神经酰胺增加。SM和神经酰胺周转率的差异通过[H-3]棕榈酸脉冲追踪标记进行定量。此外，当NBD-SM被添加到细胞中时，它被NSMase转染的细胞水解，其速率是未转染细胞的两倍以上。与PTT 1过表达的保护作用相反，NSMase 2过表达增强了星形孢菌素或C-2-神经酰胺诱导的细胞死亡。筏中存在一部分PPT 1和NSMase 2及其底物（分别为棕榈酰化蛋白和SM），表明某些蛋白质通过NSMase激活控制细胞死亡的动态棕榈酰化/脱棕榈酰化机制。(c)2005 Wiley-Liss，Inc.
Cell differentiation and myelination involve a fine balance between stasis and programmed cell death, yet the genes that regulate this have not been clearly defined. We therefore studied two key gene products involved in oligodendrocyte plasma membrane lipid metabolism and their antagonistic role in ceramide-mediated cell death signaling. Overexpression of palmitoyl:protein thioesterase (PPT1; verified by Western blot of the V5-tagged protein and increased enzyme activity) resulted in decreased ceramide in the detergent-resistant microdomain (DRM, or raft) relative to cholesterol and sphingomyelin (SM). This PPT1 overexpression also resulted in protection against cell death induced by either staurosporine or C-2-ceramide. In contrast, overexpression of neutral sphingomyelinase 2 (NSMase2; verified by Western blot of the FLAG-tagged protein and increased enzyme activity) resulted in increased membrane NSMase and increased ceramide in rafts relative to cholesterol and SM. The difference in SM and ceramide turnover was quantitated by [H-3]palmitate pulse-chase labeling. Furthermore, when NBD-SM was added to cells, it was hydrolyzed by NSMase-transfected cells at more than twofold the rate in untransfected cells. NSMase2 overexpression enhanced cell death induced by staurosporine or C-2-ceramide, in contrast to the protective effect of PTT1 overexpression. The presence of a fraction of both PPT1 and NSMase2 in rafts together with their substrates (palmitoylated proteins and SM, respectively) suggests a mechanism for dynamic palmitoylation/depalmitoylation of certain proteins in controlling cell death via NSMase activation. (c) 2005 Wiley-Liss, Inc.