Lipidomic analysis of N-acylphosphatidylethanolamine molecular species in Arabidopsis suggests feedback regulation by N-acylethanolamines

Abstract

N-Acylphosphatidylethanolamine (NAPE) and its hydrolysis product, N-acylethanolamine (NAE), are minor but ubiquitous lipids in multicellular eukaryotes. Various physiological processes are severely affected by altering the expression of fatty acid amide hydrolase (FAAH), an NAE-hydrolyzing enzyme. To determine the effect of altered FAAH activity on NAPE molecular species composition, NAE metabolism, and general membrane lipid metabolism, quantitative profiles of NAPEs, NAEs, galactolipids, and major and minor phospholipids for FAAH mutants of Arabidopsis were determined. The NAPE molecular species content was dramatically affected by reduced FAAH activity and elevated NAE content in faah knockouts, increasing by as much as 36-fold, far more than the NAE content, suggesting negative feedback regulation of phospholipase D-mediated NAPE hydrolysis by NAE. The N-acyl composition of NAPE remained similar to that of NAE, suggesting that the NAPE precursor pool largely determines NAE composition. Exogenous NAE 12:0 treatment elevated endogenous polyunsaturated NAE and NAPE levels in seedlings; NAE levels were increased more in faah knockouts than in wild-type or FAAH overexpressors. Treated seedlings with elevated NAE and NAPE levels showed impaired growth and reduced galactolipid synthesis by the “prokaryotic”, but not the “eukaryotic”, pathway. Overall, our data provide new insights into the regulation of NAPE-NAE metabolism and coordination of membrane lipid metabolism and seedling development.

N‐Acylethanolamines: lipid metabolites with functions in plant growth and development

“…Other types, such as NAE 12:0, NAE 14:0, NAE 18:0 and NAE 18:3, are also present but at lower amounts, and may vary from species to species (Chapman et al ., ; Venables et al ., ). The NAE composition in seeds reflects the acyl content of their metabolic precursor, N ‐acylphosphatidylethanolamines (NAPE; Coulon et al ., ; Kilaru et al ., ), rather than the composition of the acyl moieties of the triacylglycerols stored in seeds. Most importantly, NAE levels in seeds decrease markedly with germination and seedling establishment (Chapman et al ., ; Kilaru et al ., ).…”

“…The NAE composition in seeds reflects the acyl content of their metabolic precursor, N ‐acylphosphatidylethanolamines (NAPE; Coulon et al ., ; Kilaru et al ., ), rather than the composition of the acyl moieties of the triacylglycerols stored in seeds. Most importantly, NAE levels in seeds decrease markedly with germination and seedling establishment (Chapman et al ., ; Kilaru et al ., ). As discussed below, there is accumulating evidence that this decrease in NAE levels is physiologically relevant for the embryo‐to‐seedling transition through interaction with abscisic acid (ABA) signaling pathways (Figure ; Teaster et al ., ; Keereetaweep et al ., ).…”

N-Acylated phospholipid metabolism and seedling growth

“…6,7 An MS-based lipidomics approach was taken to quantify NAE metabolites, including NAPEs in rodents. 8,9 In plants, Kilaru et al, 10 now report for the first time, a comprehensive profile of the NAPE species in Arabidopsis seeds and seedlings, and have discovered that the levels of this unusual phospholipid class are increased by more than 30-fold in conditions that inhibit seedling establishment. This appears to be most profound in seedlings with perturbations in the metabolite pools of N-acylethanolamines (NAEs), prompting Kilaru et al 10 to speculate that levels of NAEs may regulate this lipid mediator pathway.…”

“…In summary, lipidomic analysis suggests that PLD-mediated hydrolysis of NAPE is important to seedling establishment, and it may be an unappreciated feature of the NAE regulatory pathway in seedling growth, a pathway that intersects other phytohormone signaling pathways. 10,16 Furthermore, it also suggests that the formation of NAPE, by NAPE synthase, is an unregulated step, and that this proceeds in an unimpeded manner to result in the over accumulation of NAPE in seedlings with arrested growth.…”

“…10 It was already known that this treatment negatively regulates seedling growth via ABA signaling pathways, 24 but it was not previously appreciated that there was such an impact on the metabolism of NAPEs.…”