Identification of a Pair of Phospholipid:Diacylglycerol Acyltransferases from Developing Flax (Linum usitatissimum L.) Seed Catalyzing the Selective Production of Trilinolenin

• Published in: The Journal of biological chemistry Vol. 288; no. 33; pp. 24173 - 24188
• Main Authors: Pan, Xue, Siloto, Rodrigo M.P., Wickramarathna, Aruna D., Mietkiewska, Elzbieta, Weselake, Randall J.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 16.08.2013; American Society for Biochemistry and Molecular Biology
• Subjects: Acyltransferases - genetics; Acyltransferases - metabolism; alpha-Linolenic Acid - pharmacology; Arabidopsis - drug effects; Arabidopsis - genetics; Biocatalysis - drug effects; Diacylglycerol O-Acyltransferase - metabolism; Esters - metabolism; Flax; Flax - drug effects; Flax - enzymology; Flax - genetics; Gas Chromatography-Mass Spectrometry; Gene Expression Profiling; Gene Expression Regulation, Plant - drug effects; Genes, Plant - genetics; Genetic Complementation Test; Lipid Metabolism; Lipids; Metabolic Engineering; Molecular Genetics; Mutation - genetics; Organ Specificity - drug effects; Organ Specificity - genetics; Phenotype; Phospholipid:diacylglycerol Acyltransferases; Plant Molecular Biology; Plant Oils - metabolism; Plants, Genetically Modified; Real-Time Polymerase Chain Reaction; Recombination, Genetic - drug effects; Recombination, Genetic - genetics; Saccharomyces cerevisiae - drug effects; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Seeds - drug effects; Seeds - enzymology; Seeds - genetics; Substrate Specificity - drug effects; Triacylglycerol; Triglycerides - biosynthesis; α-Linolenic Acid; Metabolic Engineering; Molecular Genetics; Phospholipid:diacylglycerol Acyltransferases; Triacylglycerol; Lipid Metabolism; Plant Molecular Biology; Flax; α-Linolenic Acid
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.475699

The oil from flax (Linum usitatissimum L.) has high amounts of α-linolenic acid (ALA; 18:3cisΔ9,12,15) and is one of the richest sources of omega-3 polyunsaturated fatty acids (ω-3-PUFAs). To produce ∼57% ALA in triacylglycerol (TAG), it is likely that flax contains enzymes that can efficiently transfer ALA to TAG. To test this hypothesis, we conducted a systematic characterization of TAG-synthesizing enzymes from flax. We identified several genes encoding acyl-CoA:diacylglycerol acyltransferases (DGATs) and phospholipid:diacylglycerol acyltransferases (PDATs) from the flax genome database. Due to recent genome duplication, duplicated gene pairs have been identified for all genes except DGAT2-2. Analysis of gene expression indicated that two DGAT1, two DGAT2, and four PDAT genes were preferentially expressed in flax embryos. Yeast functional analysis showed that DGAT1, DGAT2, and two PDAT enzymes restored TAG synthesis when produced recombinantly in yeast H1246 strain. The activity of particular PDAT enzymes (LuPDAT1 and LuPDAT2) was stimulated by the presence of ALA. Further seed-specific expression of flax genes in Arabidopsis thaliana indicated that DGAT1, PDAT1, and PDAT2 had significant effects on seed oil phenotype. Overall, this study indicated the existence of unique PDAT enzymes from flax that are able to preferentially catalyze the synthesis of TAG containing ALA acyl moieties. The identified LuPDATs may have practical applications for increasing the accumulation of ALA and other polyunsaturated fatty acids in oilseeds for food and industrial applications. Background: Triacylglycerol (TAG) can be formed via an acyl-CoA-dependent or acyl-CoA-independent pathway. Results: Overexpressing particular flax phospholipid:diacylglycerol acyltransferase (PDAT) genes in yeast and Arabidopsis resulted in an enhanced proportion of α-linolenic acid (ALA) in TAG. Conclusion: Certain PDATs have the unique ability to efficiently channel ALA into TAG. Significance: The identified PDATs will benefit future projects aimed at producing oils with enhanced polyunsaturated fatty acid content.