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Tracking synthesis and turnover of triacylglycerol in leaves.

ABSTRACT: Triacylglycerol (TAG), typically represents <1% of leaf glycerolipids but can accumulate under stress and other conditions or if leaves are supplied with fatty acids, or in plants transformed with regulators or enzymes of lipid metabolism. To better understand the metabolism of TAG in leaves, pulse-chase radiolabelling experiments were designed to probe its synthesis and turnover. When Arabidopsis leaves were incubated with [(14)C]lauric acid (12:0), a major initial product was [(14)C]TAG. Thus, despite low steady-state levels, leaves possess substantial TAG biosynthetic capacity. The contributions of diacylglycerol acyltransferase1 and phospholipid:diacylglycerol acyltransferase1 to leaf TAG synthesis were examined by labelling of dgat1 and pdat1 mutants. The dgat1 mutant displayed a major (76%) reduction in [(14)C]TAG accumulation whereas pdat1 TAG labelling was only slightly reduced. Thus, DGAT1 has a principal role in TAG biosynthesis in young leaves. During a 4h chase period, radioactivity in TAG declined 70%, whereas the turnover of [(14)C]acyl chains of phosphatidylcholine (PC) and other polar lipids was much lower. Sixty percent of [(14)C]12:0 was directly incorporated into glycerolipids without modification, whereas 40% was elongated and desaturated to 16:0 and 18:1 by plastids. The unmodified [(14)C]12:0 and the plastid products of [(14)C]12:0 metabolism entered different pathways. Although plastid-modified (14)C-labelled products accumulated in monogalactosyldiacylglycerol, PC, phosphatidylethanolamine, and diacylglcerol (DAG), there was almost no accumulation of [(14)C]16:0 and [(14)C]18:1 in TAG. Because DAG and acyl-CoA are direct precursors of TAG, the differential labelling of polar glycerolipids and TAG by [(14)C]12:0 and its plastid-modified products provides evidence for multiple subcellular pools of both acyl-CoA and DAG.

SUBMITTER: Tjellstrom H

PROVIDER: S-EPMC4339603 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Tracking synthesis and turnover of triacylglycerol in leaves.

Tjellström Henrik H Strawsine Merissa M Ohlrogge John B JB

Journal of experimental botany 20150121 5

Triacylglycerol (TAG), typically represents <1% of leaf glycerolipids but can accumulate under stress and other conditions or if leaves are supplied with fatty acids, or in plants transformed with regulators or enzymes of lipid metabolism. To better understand the metabolism of TAG in leaves, pulse-chase radiolabelling experiments were designed to probe its synthesis and turnover. When Arabidopsis leaves were incubated with [(14)C]lauric acid (12:0), a major initial product was [(14)C]TAG. Thus, ...[more]

PMID: 25609824

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Project description:Ca2+-tolerant myocytes were isolated with endogenous triacylglycerol (TAG) stores prelabelled with [3H]palmitate and subsequently incubated for a 1h chase period with [14C]palmitate, 2% albumin and 5mM glucose. Measurements were then made of [14C]palmitate conversion into TAG and phospholipids, of loss of [3H]TAG, of glycerol release and of change in the total TAG content. Rates of de novo synthesis of TAG were calculated by a balance method. With 0. 5mM palmitate present, 5 microM adrenaline increased de novo synthesis of TAG by 81% and incorporation of [14C]palmitate into phospholipids by 59%. Significant increases in these processes with adrenaline were also seen with 0.08, 0.14 and 0.26 mM palmitate. The beta-agonist isoprenaline had little effect on de novo synthesis of TAG and had no effect on [14C]palmitate conversion into phospholipids. The alpha1-agonist phenylephrine mimicked adrenaline in increasing [14C]palmitate conversion into phospholipids but had no effect on de novo synthesis of TAG. Adrenaline did not significantly alter the myocyte glycerol 3-phosphate content but caused a persistent 40% increase in the activity of the form of glycerolphosphate acyltransferase found predominantly in the sarcoplasmic reticulum. With 0.5 mM palmitate present, the value [14C]TAG formed -decrease in [3H]TAG consistently exceeded the enzymically measured change in cell TAG content. From this it was suggested that the specific radioactivity of [3H]TAG pool(s) mobilized during the chase period was lower than that of the overall cell TAG. In the basal state, complete mobilization of TAG measured as glycerol release was low, but cycling of TAG to diacylglycerol or monoacylglycerol and back to TAG appeared to be high. With adrenaline present, glycerol release was increased 5-6-fold but recycling of lower acylglycerols to TAG was abolished. Glycerol release was inhibited by increasing extracellular palmitate from 0.08 to 0.5 mM. Adrenaline partially over-rode this effect.

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Tracking synthesis and turnover of triacylglycerol in leaves.

Publications

Tracking synthesis and turnover of triacylglycerol in leaves.

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