Intracellular Ca2+-dependent formation of N-acyl-phosphatidylethanolamines by human cytosolic phospholipase A2ε
Intracellular Ca2+-dependent formation of N-acyl-phosphatidylethanolamines by human cytosolic phospholipase A2ε
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DOI：
10.1016/j.bbalip.2019.158515
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发表时间：
2019-12-01
影响因子：
4.8
通讯作者：
Ueda, Natsuo
中科院分区：
文献类型：
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作者：
Mustafiz, Smriti Sultana Binte;Uyama, Toru;Ueda, Natsuo
已知N-酰基-磷脂酰乙醇胺（NAPE）是生物活性N-酰基乙醇胺（NAE）的前体，包括内源性大麻素花生四烯酸乙醇酰胺（anandamide）和抗炎棕榈酰乙醇酰胺。在哺乳动物中，NAPE由N-酰基转移酶产生，其将酰基链从甘油磷脂的sn-1位转移到磷脂酰乙醇胺（PE）的氨基。最近发现胞浆磷脂酶A（2）（cPLA（2）β）是一种钙离子依赖的N-酰基转移酶。然而，人们对哪种类型的磷脂在活细胞中充当底物知之甚少。本研究建立了人胚肾293细胞系，并在此细胞系中建立了强力霉素诱导人cPLA（2）降解的高效液相色谱-串联质谱（LC-MS）分析方法。当用多西环素和Ca 2+离子载体处理时，细胞产生各种各样的二酰基和烯酰基类型的NAPE以及大量的NAES。此外，二酰基和烯酰基型的PE和二酰基磷脂酰胆碱（PC）的水平下降，而那些溶血磷脂酰乙醇胺和溶血磷脂酰胆碱增加。这些结果表明，cPLA（2）通过利用内源性二酰基型和烯酰基型PE作为酰基受体，二酰基型PC和二酰基型PE作为酰基供体，依赖Ca ~（2+）产生NAPE。
N-Acyl-phosphatidylethanolamines (NAPEs) are known to be precursors of bioactive N-acylethanolamines (NAEs), including the endocannabinoid arachidonoylethanolamide (anandamide) and anti-inflammatory palmitoylethanolamide. In mammals, NAPEs are produced by N-acyltransferases, which transfer an acyl chain from the sn-1 position of glycerophospholipid to the amino group of phosphatidylethanolamine (PE). Recently, the epsilon isoform of cytosolic phospholipase A(2) (cPLA(2)epsilon) was found to be Ca2+ dependent N-acyltransferase. However, it was poorly understood which types of phospholipids serve as substrates in living cells. In the present study, we established a human embryonic kidney 293 cell line, in which doxycycline potently induces human cPLA(2)epsilon, and used these cells to analyze endogenous substrates and products of cPLA(2)epsilon with liquid chromatography-tandem mass spectrometry. When treated with doxycycline and Ca2+ ionophore, the cells produced various species of diacyl- and alkenylacyl-types of NAPEs as well as NAEs in large quantities. Moreover, the levels of diacyl- and alkenylacyl-types of PEs and diacyl-phosphatidylcholines (PCs) decreased, while those of lysopho-sphatidylethanolamines and lysophosphatidylcholines increased. These results suggested that cPLA(2)epsilon Ca2+-dependently produces NAPEs by utilizing endogenous diacyl- and alkenylacyl-types of PEs as acyl acceptors and diacyl-type PCs and diacyl-type PEs as acyl donors.