title: Generation of N-acylphosphatidylethanolamine by members of the phospholipase A/acyltransferase (PLA/AT) family.
abstract: Bioactive N-acylethanolamines (NAEs), including N-palmitoylethanolamine, N-oleoylethanolamine, and N-arachidonoylethanolamine (anandamide), are formed from membrane glycerophospholipids in animal tissues. The pathway is initiated by N-acylation of phosphatidylethanolamine to form N-acylphosphatidylethanolamine (NAPE). Despite the physiological importance of this reaction, the enzyme responsible, N-acyltransferase, remains molecularly uncharacterized. We recently demonstrated that all five members of the HRAS-like suppressor tumor family are phospholipid-metabolizing enzymes with N-acyltransferase activity and are renamed HRASLS1-5 as phospholipase A/acyltransferase (PLA/AT)-1-5. However, it was poorly understood whether these proteins were involved in the formation of NAPE in living cells. In the present studies, we first show that COS-7 cells transiently expressing recombinant PLA/AT-1, -2, -4, or -5, and HEK293 cells stably expressing PLA/AT-2 generated significant amounts of [(14)C]NAPE and [(14)C]NAE when cells were metabolically labeled with [(14)C]ethanolamine. Second, as analyzed by liquid chromatography-tandem mass spectrometry, the stable expression of PLA/AT-2 in cells remarkably increased endogenous levels of NAPEs and NAEs with various N-acyl species. Third, when NAPE-hydrolyzing phospholipase D was additionally expressed in PLA/AT-2-expressing cells, accumulating NAPE was efficiently converted to NAE. We also found that PLA/AT-2 was partly responsible for NAPE formation in HeLa cells that endogenously express PLA/AT-2. These results suggest that PLA/AT family proteins may produce NAPEs serving as precursors of bioactive NAEs in vivo.
authors: Department of Biochemistry, Kagawa University School of Medicine, 1750-1 Ikenobe, Miki, Kagawa 761-0793, Japan.
authors: Toru
authors: T
authors: Uyama
authors: Natsuki
authors: N
authors: Ikematsu
authors: Manami
authors: M
authors: Inoue
authors: Naoki
authors: N
authors: Shinohara
authors: Xing-Hua
authors: XH
authors: Jin
authors: Kazuhito
authors: K
authors: Tsuboi
authors: Takeharu
authors: T
authors: Tonai
authors: Akira
authors: A
authors: Tokumura
authors: Natsuo
authors: N
authors: Ueda
mesh_terms: D000217:Acyltransferases / Q000737:chemistry*; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002522:Chlorocebus aethiops; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D005810:Multigene Family; D020675:Peroxisomes / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000737:chemistry*; D010739:Phospholipase D / Q000378:metabolism; D010741:Phospholipases A / Q000737:chemistry*; D010743:Phospholipids / Q000737:chemistry; D034622:RNA Interference; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D010714:Phosphatidylethanolamines; D010743:Phospholipids; D000217:Acyltransferases; D010741:Phospholipases A; D010739:Phospholipase D