Role of down-regulated neutral ceramidase during all-trans retinoic acid-induced neuronal differentiation in SH-SY5Y neuroblastoma cells

• Published in: Journal of biochemistry (Tokyo) 2012-06, Vol.151 (6), p.611-620
• Main Authors: Tanaka, Kouji, Tamiya-Koizumi, Keiko, Hagiwara, Kazumi, Ito, Hiromi, Takagi, Akira, Kojima, Tetsuhito, Suzuki, Motoshi, Iwaki, Soichiro, Fujii, Satoshi, Nakamura, Mitsuhiro, Banno, Yoshiko, Kannagi, Reiji, Tsurumi, Tatsuya, Kyogashima, Mamoru, Murate, Takashi
• Format: Article
• Language: English
• Subjects: Cell Differentiation - drug effects; Cell Proliferation - drug effects; Down-Regulation - drug effects; Drug Screening Assays, Antitumor; Humans; Neuroblastoma - pathology; Neurons - drug effects; Neurons - pathology; Neutral Ceramidase - biosynthesis; Neutral Ceramidase - genetics; Neutral Ceramidase - metabolism; Structure-Activity Relationship; Tretinoin - pharmacology
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvs033

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-trans retinoic 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 Chem 2002:277:25297-304.)]. 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.