Testing models of fatty acid transfer and lipid synthesis in spinach leaf using in vivo oxygen-18 labeling.
Oxygen-18 labeling has been applied to the study of plant lipid biosynthesis for the first time. [(13)C(2)(18)O(2)]Acetate was incubated with spinach (Spinacia oleracea) leaves and the (18)O content in fatty acid methyl esters isolated from different lipid classes measured by gas chromatography-mass spectometry. Fatty acids isolated from lipids synthesized within the plastid, such as monogalactosyldiacylglycerol, show an (18)O content consistent with the exogenous acetate undergoing a single activation step and with the direct utilization of acyl-acyl carrier protein by the acyl transferases of the chloroplast. In contrast, fatty acids isolated from lipids assembled in the cytosol, such as phosphatidylcholine, show a 50% reduction in the (18)O content. This is indicative of export of the fatty acyl groups from the plastid via a free carboxylate anion, and is consistent with the acyl-acyl carrier protein thioesterase:acyl-coenzyme A (CoA) synthetase mediated export mechanism. If this were not the case and the acyl group was transferred directly from acyl-acyl carrier protein to an acyl acceptor on the cytosolic side, there would be either complete retention of (18)O or, less likely, complete loss of (18)O, but not a 50% loss of (18)O. Thus, existing models for fatty acid transfer from the plastid and for spatially separate synthesis of "prokaryotic" and "eukaryotic" lipids have both been confirmed.
Pollard
M
M
Department of Botany and Plant Pathology, Michigan State University, East Lansing, Michigan 48824, USA. pollard9@pilot.msu.edu
Ohlrogge
J
J
D000085:Acetates / Q000378:metabolism*; D000213:Acyl Carrier Protein / Q000378:metabolism; D000217:Acyltransferases / Q000378:metabolism; D002247:Carbon Isotopes; D002736:Chloroplasts / Q000378:metabolism; D003066:Coenzyme A Ligases / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D007700:Kinetics; D008055:Lipids / Q000096:biosynthesis*; D010103:Oxygen Isotopes; D010713:Phosphatidylcholines / Q000096:biosynthesis; D018515:Plant Leaves / Q000378:metabolism; D018087:Plastids / Q000378:metabolism; D018724:Spinacia oleracea / Q000378:metabolism*
D000085:Acetates; D000213:Acyl Carrier Protein; D002247:Carbon Isotopes; D005227:Fatty Acids; D008055:Lipids; D010103:Oxygen Isotopes; D010713:Phosphatidylcholines; D000217:Acyltransferases; D003066:Coenzyme A Ligases