Many plants deposit TAG in seeds and fruits as the major form of storage lipid. TAG production is of tremendous socioeconomic value in food, nutraceutical, and industrial applications, and thus numerous conventional and molecular genetic strategies have been explored in attempts to increase TAG content and modify the FA composition of plant seed oils. Much research has focused on the acyl-CoA-dependent reaction catalyzed by diacylglycerol acyltransferase (DGAT), which is an integral endoplasmic reticulum protein and has also been shown to be present in oil bodies and plastids. DGAT enzymes exhibit diverse biochemical properties among different plant species, many of which are summarized here. In addition to catalyzing a critical step in TAG biosynthesis, there is evidence that DGAT has roles in lipid metabolism associated with germination and leaf senescence. TAG can also be formed in plants via two different acyl-CoA-independent pathways, catalyzed by phospholipid: diacylglycerol acyltransferase and diacylglycerol transacylase. The current understanding of the terminal step in TAG formation in plants and the development of molecular genetic approaches aimed at altering TAG yield and FA composition of TAG are discussed.

Fatty acids (FAs) and sterols are primary metabolites that exert interrelated functions as structural and signaling lipids. Despite their common syntheses from acetyl-coenzyme A, homeostatic cross talk remains enigmatic. Six Arabidopsis (Arabidopsis thaliana) acyl-coenzyme A-binding proteins (ACBPs) are involved in FA metabolism. ACBP1 interacts with PHOSPHOLIPASE Da1 and regulates phospholipid composition. Here, its specific role in the negative modulation of sterol synthesis during embryogenesis is reported. ACBP1, likely in a liganded state, interacts with STEROL C4-METHYL OXIDASE1-1 (SMO1-1), a rate-limiting enzyme in the sterol pathway. Proembryo abortion in the double mutant indicated that the ACBP1-SMO1-1 interaction is synthetic lethal, corroborating with their strong promoter activities in developing ovules. Gas chromatography-mass spectrometry revealed quantitative and compositional changes in FAs and sterols upon overexpression or mutation of ACBP1 and/or SMO1-1. Aberrant levels of these metabolites may account for the downstream defect in lipid signaling. GLABRA2 (GL2), encoding a phospholipid/sterol-binding homeodomain transcription factor, was up-regulated in developing seeds of acbp1, smo1-1, and ACBP1+/2smo1-1 in comparison with the wild type. Consistent with the corresponding transcriptional alteration of GL2 targets, high-oil, low-mucilage phenotypes of gl2 were phenocopied in ACBP1+/2smo1-1. Thus, ACBP1 appears to modulate the metabolism of two important lipid classes (FAs and sterols) influencing cellular signaling.

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SummaryThe binding of recombinant AtACBP1 to very-long-chain acyl-CoA esters is related to AtACBP1 function in Arabidopsis stem cuticle metabolism. Loss-of-function mutation adversely affected stem cuticle composition and structure.

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Phytate, the major organic phosphorus in soil, is not readily available to plants as a source of phosphorus (P). It is either complexed with cations or adsorbed to various soil components. The present study was carried out to investigate the extracellular phytase activities of tobacco (Nicotiana tabacum variety GeXin No.1) and its ability to assimilate external phytate-P. Whereas phytase activities in roots, shoots and growth media of P i -fed 14-day-old seedlings were only 1.3-4.9% of total acid phosphatase (APase) activities, P starvation triggered an increase in phytase secretion up to 914.9 mU mg )1 protein, equivalent to 18.2% of total APase activities. Much of the extracellular phytase activities were found to be rootassociated than root-released. The plants were not able to utilize phytate adsorbed to sand, except when insoluble phytate salts were preformed with Mg 2+ and Ca 2+ ions for supplementation. Tobacco grew better in sand supplemented with Mg-phytate salts (31.9 mg dry weight plant )1 ; 0.68% w/w P concentration) than that with Ca-phytate salts (9.5 mg plant )1 ; 0.42%), presumably due to its higher solubility. We conclude that insolubility of soil phytate is the major constrain for its assimilation. Improving solubility of soil phytate, for example, by enhancement of citrate secretion, may be a feasible approach to improve soil phytate assimilation.

Diacylglycerol acyltransferase (DGAT) catalyzes the acyl-CoA-dependent acylation of sn-1,2-diacylglycerol to produce triacylglycerol (TAG). This enzyme, which is critical to numerous facets of oilseed development, has been highlighted as a genetic engineering target to increase storage lipid production in microorganisms designed for biofuel applications. Here, four transcriptionally active DGAT1 genes were identified and characterized from the oil crop Brassica napus. Overexpression of each BnaDGAT1 in Saccharomyces cerevisiae increased TAG biosynthesis. Further studies showed that adding an N-terminal tag could mask the deleterious influence of the DGATs' native N-terminal sequences, resulting in increased in vivo accumulation of the polypeptides and an increase of up to about 150-fold in in vitro enzyme activity. The levels of TAG and total lipid fatty acids in S. cerevisiae producing the N-terminally tagged BnaDGAT1.b at 72 h were 53 and 28 % higher than those in cultures producing untagged BnaA.DGAT1.b, respectively. These modified DGATs catalyzed the synthesis of up to 453 mg fatty acid/L by this time point. The results will be of benefit in the biochemical analysis of recombinant DGAT1 produced through heterologous expression in yeast and offer a new approach to increase storage lipid content in yeast for industrial applications.

Oxylipins are crucial components in plant wound responses that are mobilised via the plant vasculature. Previous studies have shown that the overexpression of an Arabidopsis acyl-CoA-binding protein, AtACBP3, led to an accumulation of oxylipin-containing galactolipids, and AtACBP3pro::BETA-GLUCURONIDASE (GUS) was expressed in the phloem of transgenic Arabidopsis. To investigate the role of AtACBP3 in the phloem, reverse transcription-polymerase chain reaction and western blot analysis of phloem exudates from the acbp3 mutant and wild type revealed that the AtACBP3 protein, but not its mRNA, was detected in the phloem sap. Furthermore, micrografting demonstrated that AtACBP3 expressed from the 35S promoter was translocated from shoot to root. Subsequently, AtACBP3 was localised to the companion cells, sieve elements and the apoplastic space of phloem tissue by immunogold electron microscopy using anti-AtACBP3 antibodies. AtACBP3pro::GUS was induced locally in Arabidopsis leaves upon wounding, and the expression of wound-responsive jasmonic acid marker genes (JASMONATE ZIM-DOMAIN10, VEGETATIVE STORAGE PROTEIN2, and LIPOXYGENASE2) increased more significantly in both locally wounded and systemic leaves of the wild type in comparison to acbp3 and AtACBP3-RNAi. Oxylipin-related fatty acid (FA) (C18:2-FA, C18:3-FA and methyl jasmonate) content was observed to be lower in acbp3 and AtACBP3-RNAi than wild-type phloem exudates using gas chromatography-mass spectrometry. Experiments using recombinant AtACBP3 in isothermal titration calorimetry analysis showed that medium- and long-chain acyl-CoA esters bind (His)6-AtACBP3 with KD values in the micromolar range. Taken together, these results suggest that AtACBP3 is likely to be a phloem-mobile protein that affects the FA pool and jasmonate content in the phloem, possibly by its binding to acyl-CoA esters.