Role of down-regulated neutral ceramidase during all-trans retinoic acid induced neuronal differentiation in SH-SY5Y neuroblastoma cells
Role of down-regulated neutral ceramidase during all-trans retinoic acid induced neuronal differentiation in SH-SY5Y neuroblastoma cells
复制标题
下调中性神经酰胺酶在全反式维A酸诱导SH-SY5Y神经母细胞瘤细胞神经元分化中的作用
DOI：
10.1093/jb/mvs033
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Kouji Tanaka
中科院分区：
文献类型：
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作者：
Kanamori E;Itoh M;Tojo N;Koyama T;Nara N;Tohda S;Kouji Tanaka
神经酰胺是一种重要的生物活性物质，神经酰胺酶是控制神经酰胺代谢的关键酶，决定着细胞的命运。已报道全反式视黄酸（ATRA）诱导神经元分化和细胞周期停滞[Lopez-Carballo，Moreno，Masia，Perez，and Barettino（Activation of the phosphatidylinositol 3-kinase/Akt signaling pathway by retinoic acid is required for neural differentiation of SH-SY 5 Y human neuroblastoma cells.J Biol Chem 2002：277：25297-304.）]。在这项研究中，我们观察到ATRA诱导的细胞神经酰胺积累，细胞生长停滞和分化伴随着NCD酶在SH-SY 5 Y细胞中的下调，而没有减少鞘氨醇或鞘氨醇1-磷酸。我们研究了NCD酶的下调是否参与神经酰胺和细胞分化的增加。ATRA可下调NCD酶的mRNA、蛋白和酶活性。有趣的是，加塔-2也被ATRA处理后降低，并且使用其表达载体和siRNA的实验以及染色质免疫沉淀分析证实加塔-2作为NCD酶基因表达的转录因子。通过建立NCDase表达和活性降低的稳定转染子，我们阐明了NCDase下调对ATRA诱导的神经元分化的意义。这些亚克隆表现出增加的细胞神经酰胺和减少的细胞生长以及神经元分化表型。这些结果表明，通过ATRA诱导的加塔-2减少下调NCD酶在诱导神经酰胺积累和神经元分化中起重要作用。
Neutral ceramidase (NCDase) is considered to be a critical enzyme for controlling the turnover of ceramide, an important bioactive lipid, which determines cell's fate. All-transretinoic acid (ATRA) has been reported to induce neuronal differentiation and cell-cycle arrest [Lopez-Carballo, Moreno, Masia, Perez, and Barettino (Activation of the phosphatidylinositol 3-kinase/Akt signalling pathway by retinoic acid is required for neural differentiation of SH-SY5Y human neuroblastoma cells.J Biol Chem2002:277:25297–304.)]. In this study, we observed that ATRA-induced cellular ceramide accumulation, cell-growth arrest and differentiation accompanied with down-regulation of NCDase in SH-SY5Y cells, without a decrease in sphingosine or sphingosine 1-phosphate. We examined whether the down-regulation of NCDase was involved in the increase in ceramide and cell differentiation. ATRA was found to down-regulate mRNA, protein and the enzyme activity of NCDase. Interestingly, GATA-2 was also decreased with ATRA treatment, and experiments using its expression vector and siRNA and chromatin immunoprecipitation assay demonstrated GATA-2 acted as transcription-factor of NCDase gene expression. By establishing stable transfectants with decreased NCDase expression and activity, we clarified the significance of NCDase down-regulation for ATRA-induced neuronal differentiation. Those sub-clones showed both increased cellular ceramide and reduced cell growth as well as neuronal differentiation phenotypes. These results demonstrate that down-regulation of NCDase through ATRA-induced GATA-2 decrease plays an important role in induction of ceramide accumulation and neuronal differentiation in SH-SY5Y cells.