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

Pan, Xue; Siloto, Rodrigo M.P.; Wickramarathna, Aruna D.; Mietkiewska, Elzbieta; Weselake, Randall J.

doi:10.1074/jbc.m113.475699

Cited by 98 publications

(124 citation statements)

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“…sativa PDAT overexpression affects the fatty acid profile differently than the overexpression of Arabidopsis and flax PDAT in Arabidopsis (Ståhl et al, 2004;Pan et al, 2013), suggesting different specificities of the enzymes in different species. The increase in 18:2n-6 at the expense of 18:3n-3 suggests that the usage of 18:2n-6 by PDAT supersedes that of FAD3, and 18:2n-6 is to a high degree removed from PC by PDAT before it is desaturated to 18:3n-3.…”

Section: Pdat Overexpressionmentioning

confidence: 99%

“…For Arabidopsis, neither PDAT overexpression nor knockout leads to any changes in total lipid content or fatty acid composition of the seed oil (Ståhl et al, 2004;Mhaske et al, 2005;Banas et al, 2014). Overexpression of flax PDAT in Arabidopsis, on the one hand, gives an increase in 18:3n-3 as well as, to a lesser degree, 18:2n-6 (Pan et al, 2013). PDAT overexpression in C. sativa, on the other hand, leads to a large increase in 18:2n-6 content both alone and in combination with amiDGAT1 constructs (Fig.…”

Section: Pdat Overexpressionmentioning

confidence: 99%

“…DGAT1 has been shown in Arabidopsis to be the major, but not the sole, enzyme to contribute to the accumulation of seed TAGs, working together mainly with PDAT (Zhang et al, 2009). While overexpression of intrinsic Arabidopsis PDAT influenced neither total lipid content nor the fatty acid profile (Ståhl et al, 2004), PDAT from flax (Linum usitatissimum), another species with even higher 18:3n-3 content than C. sativa, was shown to increase the 18:3n-3 content when overexpressed in Arabidopsis (Pan et al, 2013). Therefore, it has become apparent that the same enzyme homolog from different species can have quite different substrate specificities, which may affect the fatty acid profile of the seed oil of specific species.…”

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Two Acyltransferases Contribute Differently to Linolenic Acid Levels in Seed Oil

et al. 2017

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ORCID IDs: 0000-0001-6929-7859 (S.M.); 0000-0003-3152-0394 (D.S.); 0000-0001-8255-3255 (C.H.); 0000-0003-0489-3072 (K.C.); 0000-0002-9888-7003 (I.F.).Acyltransferases are key contributors to triacylglycerol (TAG) synthesis and, thus, are of great importance for seed oil quality. The effects of increased or decreased expression of ACYL-COENZYME A:DIACYLGLYCEROL ACYLTRANSFERASE1 (DGAT1) or PHOSPHOLIPID:DIACYLGLYCEROL ACYLTRANSFERASE (PDAT) on seed lipid composition were assessed in several Camelina sativa lines. Furthermore, in vitro assays of acyltransferases in microsomal fractions prepared from developing seeds of some of these lines were performed. Decreased expression of DGAT1 led to an increased percentage of 18:3n-3 without any change in total lipid content of the seed. The tri-18:3 TAG increase occurred predominantly in the cotyledon, as determined with matrix-assisted laser desorption/ionization-mass spectrometry, whereas species with two 18:3n-3 acyl groups were elevated in both cotyledon and embryonal axis. PDAT overexpression led to a relative increase of 18:2n-6 at the expense of 18:3n-3, also without affecting the total lipid content. Differential distributions of TAG species also were observed in different parts of the seed. The microsomal assays revealed that C. sativa seeds have very high activity of diacylglycerol-phosphatidylcholine interconversion. The combination of analytical and biochemical data suggests that the higher 18:2n-6 content in the seed oil of the PDAT overexpressors is due to the channeling of fatty acids from phosphatidylcholine into TAG before being desaturated to 18:3n-3, caused by the high activity of PDAT in general and by PDAT specificity for 18:2n-6. The higher levels of 18:3n-3 in DGAT1-silencing lines are likely due to the compensatory activity of a TAG-synthesizing enzyme with specificity for this acyl group and more desaturation of acyl groups occurring on phosphatidylcholine.

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Section: Pdat Overexpressionmentioning

confidence: 99%

Section: Pdat Overexpressionmentioning

confidence: 99%

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confidence: 99%

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Two Acyltransferases Contribute Differently to Linolenic Acid Levels in Seed Oil

et al. 2017

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“…In vivo substrates for DCR were not identified; however, it was proposed that DCR might be involved in intracellular concatenation of the cutin polyester (Panikashvili et al, 2009;Molina and Kosma, 2015). Although DCR has diacylglycerol acyltransferase activity in vitro (Rani et al, 2010), flax DCR could not complement the Arabidopsis dgat1 knockout mutant (Pan et al, 2013) and DAG or TAG has not been identified as an intermediate in cutin production. That DCR is the most highly expressed of all enzyme transcripts in Bayberry knobs and is temporally correlated with wax production implies that the enzyme has a key role in surface wax production in Bayberry.…”

Section: Genes Related To Biosynthesis and Secretion Of Surface Lipidmentioning

confidence: 99%

A Novel Pathway for Triacylglycerol Biosynthesis Is Responsible for the Accumulation of Massive Quantities of Glycerolipids in the Surface Wax of Bayberry (Myrica pensylvanica) Fruit

Simpson

Ohlrogge

2016

Plant Cell

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ORCID ID: 0000-0003-4019-0681 (J.P.S.)Bayberry (Myrica pensylvanica) fruits synthesize an extremely thick and unusual layer of crystalline surface wax that accumulates to 32% of fruit dry weight, the highest reported surface lipid accumulation in plants. The composition is also striking, consisting of completely saturated triacylglycerol, diacylglycerol, and monoacylglycerol with palmitate and myristate acyl chains. To gain insight into the unique properties of Bayberry wax synthesis, we examined the chemical and morphological development of the wax layer, monitored wax biosynthesis through The most highly expressed lipid-related genes were associated with production of cutin, whereas transcripts for conventional TAG synthesis were >50-fold less abundant. The biochemical and expression data together indicate that Bayberry surface glycerolipids are synthesized by a pathway for TAG synthesis that is related to cutin biosynthesis. The combination of a unique surface wax and massive accumulation may aid understanding of how plants produce and secrete non-membrane glycerolipids and also how to engineer alternative pathways for lipid production in non-seeds.

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“…fatty acids including glycerolipids and fatty acyl-CoA were transmethylated by base-catalyzed transesterifi cation using 5% (w/v) sodium methoxide (CH 3 ONa) (Sigma-Aldrich) at room temperature for 30 min ( 21 ). For the third method, diazomethane was used as to react with unesterifi ed fatty acids (free fatty acids) in the presence of a little methanol, which catalyzes the reaction to form FAMEs ( 22 ).…”

Section: Downloaded Frommentioning

confidence: 99%

Metabolic engineering of Pichia pastoris to produce ricinoleic acid, a hydroxy fatty acid of industrial importance

Meesapyodsuk

Chen

et al. 2015

Journal of Lipid Research

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This article is available online at http://www.jlr.org However, due to the presence of a highly potent toxin (ricin), this native oilseed plant is not considered an ideal source for hydroxy fatty acid production. Therefore, tremendous effort has recently been made in identifying genes involved in the biosynthesis of this fatty acid and using the genes to engineer oilseed crops for producing ricinoleic acid ( 3-7 ). However, to date these attempts have met with only partial success. When a hydroxylase gene from a native plant species was introduced into oilseed crops, the hydroxy fatty acid content in transgenic seeds rarely exceeded 20% of the total fatty acids ( 8-10 ). In 2008, a new hydroxylase for the biosynthesis of ricinoleic acid was identifi ed from a nonplant origin, and expression of this gene in Arabidopsis resulted in accumulation of a slightly higher level of hydroxy fatty acids in transgenic seeds ( 11 ). Very recently, coexpression of genes encoding factors involved in the networks of the hydroxylation or acyl traffi cking process along with a hydroxylase has also been attempted ( 4-7 ). Although the improved production of hydroxy fatty acids is observed, the amount in transgenic seeds still rarely exceeds 25% of the total fatty acids. It appears that selection of genes from diverse sources and addition of cofactors can make a difference for transgenic production of hydroxyl fatty acids, but production of a single hydroxy fatty acid at the commercially viable level in plants still remains as a challenging task.Microorganisms have recently emerged as promising systems for producing biofuel for transportation and platform chemicals for biopolymers because some microbes offer high output of biomass with good oil content in a short period of time and are easily used for genetic manipulation to implement a metabolic engineering strategy. Exploitation of metabolic engineering of lipid pathways in Abstract Ricinoleic acid (12-hydroxyoctadec-cis -9-enoic acid) has many specialized uses in bioproduct industries, while castor bean is currently the only commercial source for the fatty acid. This report describes metabolic engineering of a microbial system ( Pichia pastoris ) to produce ricinoleic acid using a "push" (synthesis) and "pull" (assembly) strategy. CpFAH, a fatty acid hydroxylase from Claviceps purpurea , was used for synthesis of ricinoleic acid, and CpDGAT1, a diacylglycerol acyl transferase for the triacylglycerol synthesis from the same species, was used for assembly of the fatty acid. Coexpression of CpFAH and CpDGAT1 produced higher lipid contents and ricinoleic acid levels than expression of CpFAH alone. Coexpression in a mutant haploid strain defective in the ⌬ 12 desaturase activity resulted in a higher level of ricinoleic acid than that in the diploid strain. Intriguingly, the ricinoleic acid produced was mainly distributed in the neutral lipid fractions, particularly the free fatty acid form, but with little in the polar lipids. This work demonstrates the effectiveness of the metabolic ...

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

Cited by 98 publications

References 59 publications

Two Acyltransferases Contribute Differently to Linolenic Acid Levels in Seed Oil

Two Acyltransferases Contribute Differently to Linolenic Acid Levels in Seed Oil

A Novel Pathway for Triacylglycerol Biosynthesis Is Responsible for the Accumulation of Massive Quantities of Glycerolipids in the Surface Wax of Bayberry (Myrica pensylvanica) Fruit

Metabolic engineering of Pichia pastoris to produce ricinoleic acid, a hydroxy fatty acid of industrial importance

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