Glycerol-3-phosphate acyltransferase-1 upregulation by O-GlcNAcylation of Sp1 protects against hypoxia-induced mouse embryonic stem cell apoptosis via mTOR activation.
Glycerol-3-phosphate acyltransferase-1 upregulation by O-GlcNAcylation of Sp1 protects against hypoxia-induced mouse embryonic stem cell apoptosis via mTOR activation.
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DOI：
10.1038/cddis.2015.410
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发表时间：
2016-03-24
影响因子：
9
通讯作者：
Han HJ
中科院分区：
文献类型：
--
作者：
Lee HJ;Ryu JM;Jung YH;Lee KH;Kim DI;Han HJ
氧信号对于干细胞调节是至关重要的，并且氧化应激诱导的干细胞凋亡降低了干细胞治疗的效率。缺氧激活干细胞的O-连接的β-N-乙酰葡糖胺化（O-GlcNAc化），这有助于调节细胞代谢以及细胞命运。本研究探讨了葡萄糖胺修饰O-GlcNAc在缺氧诱导的小鼠胚胎干细胞凋亡中的保护作用。缺氧可使mESCs凋亡呈时间依赖性增加。此外，缺氧也轻微增加O-GlcNAc水平。氨基葡萄糖治疗进一步提高了O-GlcNAc水平，并防止缺氧诱导的mESC凋亡，这是由O-GlcNAc转移酶抑制剂抑制。此外，缺氧调节几种脂质代谢酶，而葡萄糖胺增加甘油-3-磷酸酰基转移酶-1（GPAT 1）的表达，这是一种产生溶血磷脂酸（LPA）的脂质代谢酶。此外，葡糖胺增加了Sp1的O-GlcNAc化，随后导致Sp1核转位和GPAT 1表达。通过gpat 1 siRNA转染沉默GPAT 1减少了mESC中葡糖胺介导的抗凋亡，并减少了哺乳动物雷帕霉素靶蛋白（mTOR）磷酸化。事实上，LPA阻止了mESC经历缺氧诱导的凋亡，并增加了mTOR及其底物（S6 K1和4 EBP 1）的磷酸化。此外，雷帕霉素（mTOR抑制剂）使mTOR失活增加促凋亡蛋白的表达和mESC凋亡。此外，在小鼠皮瓣模型中，非靶向siRNA和葡糖胺处理的mESC的移植增加了细胞存活并抑制了皮瓣坏死。相反，GPAT 1表达的沉默逆转了这些葡萄糖胺的作用。总之，通过葡糖胺增强Sp1的O-GlcNAc化刺激GPAT 1表达，这导致通过mTOR激活抑制缺氧诱导的mESC凋亡。
Oxygen signaling is critical for stem cell regulation, and oxidative stress-induced stem cell apoptosis decreases the efficiency of stem cell therapy. Hypoxia activates O-linked β-N-acetyl glucosaminylation (O-GlcNAcylation) of stem cells, which contributes to regulation of cellular metabolism, as well as cell fate. Our study investigated the role of O-GlcNAcylation via glucosamine in the protection of hypoxia-induced apoptosis of mouse embryonic stem cells (mESCs). Hypoxia increased mESCs apoptosis in a time-dependent manner. Moreover, hypoxia also slightly increased the O-GlcNAc level. Glucosamine treatment further enhanced the O-GlcNAc level and prevented hypoxia-induced mESC apoptosis, which was suppressed by O-GlcNAc transferase inhibitors. In addition, hypoxia regulated several lipid metabolic enzymes, whereas glucosamine increased expression of glycerol-3-phosphate acyltransferase-1 (GPAT1), a lipid metabolic enzyme producing lysophosphatidic acid (LPA). In addition, glucosamine-increased O-GlcNAcylation of Sp1, which subsequently leads to Sp1 nuclear translocation and GPAT1 expression. Silencing of GPAT1 by gpat1 siRNA transfection reduced glucosamine-mediated anti-apoptosis in mESCs and reduced mammalian target of rapamycin (mTOR) phosphorylation. Indeed, LPA prevented mESCs from undergoing hypoxia-induced apoptosis and increased phosphorylation of mTOR and its substrates (S6K1 and 4EBP1). Moreover, mTOR inactivation by rapamycin (mTOR inhibitor) increased pro-apoptotic proteins expressions and mESC apoptosis. Furthermore, transplantation of non-targeting siRNA and glucosamine-treated mESCs increased cell survival and inhibited flap necrosis in mouse skin flap model. Conversely, silencing of GPAT1 expression reversed those glucosamine effects. In conclusion, enhancing O-GlcNAcylation of Sp1 by glucosamine stimulates GPAT1 expression, which leads to inhibition of hypoxia-induced mESC apoptosis via mTOR activation.