Phosphorus supply is a major factor responsible for reduced crop yields. As a result, plants utilize various adaptive mechanisms against phosphorus depletion, including lipid remodelling. Here we report the involvement of a novel plant lipid, glucuronosyldiacylglycerol, against phosphorus depletion. Lipidomic analysis of Arabidopsis plants cultured in phosphorus-depleted conditions revealed inducible accumulation of glucuronosyldiacylglycerol. Investigation using a series of sulfolipid sulfoquinovosyldiacylglycerol synthesis-deficient mutants of Arabidopsis determined that the biosynthesis of glucuronosyldiacylglycerol shares the pathway of sulfoquinovosyldiacylglycerol synthesis in chloroplasts. Under phosphorus-depleted conditions, the Arabidopsis sqd2 mutant, which does not accumulate either sulfoquinovosyldiacylglycerol or glucuronosyldiacylglycerol, was the most severely damaged of three sulfoquinovosyldiacylglycerol-deficient mutants. As glucuronosyldiacylglycerol is still present in the other two mutants, this result indicates that glucuronosyldiacylglycerol has a role in the protection of plants against phosphorus limitation stress. Glucuronosyldiacylglycerol was also found in rice, and its concentration increased significantly following phosphorus limitation, suggesting a shared physiological significance of this novel lipid against phosphorus depletion in plants.

Summary 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGR) catalyzes the ®rst committed step in the cytosolic isoprenoid biosynthesis pathway in higher plants. To understand the contribution of HMGR to plant development, we isolated T-DNA insertion mutants for HMG1 and HMG2. The hmg1 and hmg2 mutants were both more sensitive than the wild type (WT) to lovastatin, an inhibitor of HMGR. The hmg2 mutant showed no visible phenotype under normal growth conditions. In contrast, the hmg1 mutant exhibited dwar®ng, early senescence, and sterility. Expression of senescence-associated genes 12 (SAG12 ), a marker gene for senescence, was induced in the hmg1 mutant at an earlier stage than in the WT. Levels of trans-cytokinins ± hormones known to inhibit senescence ± were not lower in hmg1. The mutant did not have the typical appearance of brassinosteroid (BR)-de®cient mutants, except for a dwarf phenotype, because of the suppression of cell elongation. The expression of several genes involved in cell elongation was suppressed in hmg1. WT plants treated exogenously with inhibitors of sterol biosynthesis had similar gene expression and sterility characteristics as the hmg1 mutants. Pleiotropic phenotypes were rescued by feeding with squalene, the precursor of sterols and triterpenoids. The sterol levels in hmg1 mutants were lower than in the WT. These ®ndings suggest that HMG1 plays a critical role in triterpene biosynthesis, and that sterols and/or triterpenoids contribute to cell elongation, senescence, and fertility.

Plants synthesize a wide range of hydrophobic compounds, generally known as lipids. Here, we report an application of liquid chromatography ion trap time-of-flight mass spectrometry (LC-IT-TOF-MS) for plant lipidomics. Using hydrophilic interaction chromatography (HILIC) for class separation, typical membrane lipids including glycerolipids, steryl glucosides and glucosylceramides, and hydrophobic plant secondary metabolites such as saponins were analyzed simultaneously. By this method, we annotated approximately 100 molecules from Arabidopsis thaliana. To demonstrate the application of this method to biological study, we analyzed Arabidopsis mutant trigalactosyldiacylglycerol3 (tgd3), which has a complex metabolic phenotype including the accumulation of unusual forms of galactolipids. Lipid profiling by LC-MS revealed that tgd3 accumulated an unusual form of digalactosyldiacylglycerol, annotated as Gal(β1 → 6)βGalDG. The compositional difference between normal and unusual forms of digalactosyldiacylglycerol was detected by this method. In addition, we analyzed well-known Arabidopsis mutants ats1-1, fad6-1, and fad7-2, which are also disrupted in lipid metabolic genes. Untargeted lipidome analysis coupled with multivariate analysis clearly discriminated the mutants and their distinctive metabolites. These results indicated that HILIC-MS is an efficient method for plant lipidomics.Electronic supplementary materialThe online version of this article (doi:10.1007/s11306-011-0318-z) contains supplementary material, which is available to authorized users.

Plants have two isoprenoid biosynthetic pathways: the cytosolic mevalonate (MVA) pathway and the plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway. Since the discovery of the MEP pathway, possible metabolic cross-talk between these pathways has prompted intense research. Although many studies have shown the existence of such cross-talk using feeding experiments, it remains to be determined if native cross-talk, rather than exogenously applied metabolites, can compensate for complete blockage of the MVA pathway. Previously, Arabidopsis mutants for HMG1 and HMG2 encoding HMG-CoA reductase (HMGR) were isolated. Although it was shown that HMGR1 is a functional HMGR, the enzyme activity of HMGR2 has not been confirmed. It is demonstrated here that HMG2 encodes a functional reductase with similar activity to HMGR1, using enzyme assays and complementation experiments. To estimate the contribution of native cross-talk, an attempt was made to block the MVA pathway by making double mutants lacking both HMG1 and HMG2, but no double homozygotes were detected in the progeny of self-pollinated HMG1/hmg1 hmg2/hmg2 plants. hmg1 hmg2 male gametophytes appeared to be lethal based on crossing experiments, and microscopy indicated that ∼50% of the microspores from the HMG1/hmg1 hmg2/hmg2 plant appeared shrunken and exhibited poorly defined endoplasmic reticulum membranes. In situ hybridization showed that HMG1 transcripts were expressed in both the tapetum and microspores, while HMG2 mRNA appeared only in microspores. It is concluded that native cross-talk from the plastid cannot compensate for complete blockage of the MVA pathway, at least during male gametophyte development, because either HMG1 or HMG2 is required for male gametophyte development.

While exogenous toxic compounds such as herbicides are thought to be sequestered into vacuoles in the form of glutathione (GSH) conjugates, little is understood about natural plant products conjugated with GSH. To identify natural products conjugated with GSH in plants, metabolites in the Arabidopsis g-glutamyl transpeptidase (ggt) 4 knockout mutants that are blocked in the degradation of GSH conjugates in the vacuole were compared with those in wild-type plants. Among the metabolites identified, one was confirmed to be the 12-oxo-phytodienoic acid (OPDA)-GSH conjugate, indicating that OPDA, a precursor of jasmonic acid (JA), is transported into the vacuole as a GSH conjugate.