title: Distinguishing phosphatidic acid pools from de novo synthesis, PLD, and DGK.
abstract: In plants, phosphatidic acid (PA) functions as a metabolic precursor in the biosynthesis of glycerolipids, but it also acts as a key signaling lipid in the response to environmental stress conditions (Testerink and Munnik, J Exp Bot 62:2349-2361, 2011). In vivo (32)P-radiolabeling assays have shown the level of PA to increase within seconds/minutes of exposure to a stimulus. This response can be due to the activity of diacylglycerol kinase (DGK) and/or phospholipase D (PLD). A method is described to investigate which of the pathways is responsible for PA accumulation under a particular stress condition. First, a differential (32)P-radiolabeling protocol is used to discriminate (32)P-PA pools that are rapidly labeled versus those requiring long prelabeling times, reflecting DGK and PLD activities, respectively. Second, to specifically monitor the contribution of PLD, a transphosphatidylation assay is applied, which makes use of the artificial lipid phosphatidylbutanol as an in vivo marker of PLD activity.
authors.affiliation: Swammerdam Institute for Life Sciences, Section Plant Physiology, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Steven A
authors.initials: SA
authors.lastname: Arisz
authors.forename: Teun
authors.initials: T
authors.lastname: Munnik
mesh_terms: D019852:Diacylglycerol Kinase / Q000378:metabolism*; D007553:Isotope Labeling; D010712:Phosphatidic Acids / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D013997:Time Factors
chemical_list: D010712:Phosphatidic Acids; D010761:Phosphorus Radioisotopes; D019852:Diacylglycerol Kinase; D010739:Phospholipase D