Intracellular Ca2+-dependent formation of N-acyl-phosphatidylethanolamines by human cytosolic phospholipase A2ε

Involvement of the γ Isoform of cPLA2 in the Biosynthesis of Bioactive N-Acylethanolamines

“…The differences in the expression levels between ε and other isoforms were presumably attributed to the differences in the expression methods. Metabolic labeling of cPLA 2 /Tet-on cells with [ 14 C]ethanolamine, followed by the ionomycin treatment, exhibited the production of large amounts of radioactive NAPE, NAE, and lysoNAPE due to the activation of cPLA 2 (Figure 3B) as reported previously [18]. Interestingly, as compared with the sole expression of , the co-expression of with γ, but not with α, β, δ, or ζ, showed a lower level of NAPE (Figure 3C) and higher levels of NAE (Figure 3D) and lysoNAPE (Figure 3E).…”

“…The extra bands were presumed to be degradative or modified proteins of each isoform. We recently reported that cPLA 2 -expressing cells pretreated with [ 14 C]ethanolamine produce a large amount of [ 14 C]NAPE in response to the Ca 2+ ionophore ionomycin [18]. Thus, in the present study, we used this metabolic labeling as a simple method to detect Ca 2+ -dependent N-acyltransferase activity in living cells.…”

“…Since all the cPLA 2 isoforms were previously reported to exhibit PLA 1 /A 2 activity for glycerophospholipids such as PC [17], it was likely that these isoforms also showed PLA 1 /A 2 activity for NAPE in living cells. For this purpose, we used cPLA 2 /Tet-on cells, which stably expressed FLAG-tagged cPLA 2 in the presence of doxycycline (DOX) and produced a large amount of NAPE when stimulated by a Ca 2+ ionophore [18]. We transiently expressed one of α, β, γ, δ, and ζ isoforms with a FLAG tag in cPLA 2 /Teton cells in the presence of DOX.…”

“…A Tet-on cell line (FL-cPLA 2 ε/Tet-on), which DOX-dependently expresses FLAGtagged cPLA 2 ε, was established by the transfection of HEK293 cells with pcDNA5/TO vector harboring FLAG-tagged cPLA 2 ε as reported previously [18]. The cells were maintained for at least four days in the presence of 1 µg/mL DOX.…”

Updating Phospholipase A2 Biology

“…Recently, it has been shown that cPLA 2 ε (group IVE PLA 2 ; encoded by PLA2G4E ) functions as a Ca 2+ -dependent N -acyltransferase [ 102 ]. In response to Ca 2+ ionophore stimulation, HEK293 cells overexpressing cPLA 2 ε produce various NAPE and NAE species, accompanied by concomitant decreases in PE and PC and increases in LPE and LPC, indicating that cPLA 2 ε produces NAPEs by utilizing the diacyl- and plasmalogen types of PE as acyl acceptors and the diacyl types of PC and PE as acyl donors [ 103 ] ( Figure 4 A). The activity of cPLA 2 ε is markedly enhanced by the presence of phosphatidylserine (PS) or other anionic phospholipids, and cPLA 2 ε largely co-localizes with PS in the plasma membrane and organelles involved in the endocytic pathway [ 104 , 105 ].…”