Application of a metabolomic method combining one-dimensional and two-dimensional gas chromatography-time-of-flight/mass spectrometry to metabolic phenotyping of natural variants in rice

Kusano, Miyako; Fukushima, Atsushi; Kobayashi, Makoto; Hayashi, Naomi; Jonsson, Pär; Moritz, Thomas; Ebana, Kaworu; Saito, Kazuki

doi:10.1016/j.jchromb.2007.05.002

Cited by 160 publications

(112 citation statements)

References 45 publications

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“…Samples of nodules, leaves, and roots (25 mg fresh weight of each sample and four biological replicates per tissue type) were extracted in 1 mL extraction mixture comprised of chloroform:MeOH:H 2 O (2:6:2 vol/vol/vol) and 10 stable isotope reference compounds (65). After extraction, 200 μL supernatant was evaporated to dryness and then, derivatized as previously described (66).…”

Section: Methodsmentioning

confidence: 99%

Adaptation of the symbiotic Mesorhizobium –chickpea relationship to phosphate deficiency relies on reprogramming of whole-plant metabolism

Esfahani

Kusano

Nguyen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Low inorganic phosphate (Pi) availability is a major constraint for efficient nitrogen fixation in legumes, including chickpea. To elucidate the mechanisms involved in nodule acclimation to low Pi availability, two Mesorhizobium-chickpea associations exhibiting differential symbiotic performances, Mesorhizobium ciceri CP-31 (McCP-31)-chickpea and Mesorhizobium mediterranum SWRI9 (MmSWRI9)-chickpea, were comprehensively studied under both control and low Pi conditions. MmSWRI9-chickpea showed a lower symbiotic efficiency under low Pi availability than McCP-31-chickpea as evidenced by reduced growth parameters and down-regulation of nifD and nifK. These differences can be attributed to decline in Pi level in MmSWRI9-induced nodules under low Pi stress, which coincided with up-regulation of several key Pi starvation-responsive genes, and accumulation of asparagine in nodules and the levels of identified amino acids in Pi-deficient leaves of MmSWRI9-inoculated plants exceeding the shoot nitrogen requirement during Pi starvation, indicative of nitrogen feedback inhibition. Conversely, Pi levels increased in nodules of Pi-stressed McCP-31-inoculated plants, because these plants evolved various metabolic and biochemical strategies to maintain nodular Pi homeostasis under Pi deficiency. These adaptations involve the activation of alternative pathways of carbon metabolism, enhanced production and exudation of organic acids from roots into the rhizosphere, and the ability to protect nodule metabolism against Pi deficiency-induced oxidative stress. Collectively, the adaptation of symbiotic efficiency under Pi deficiency resulted from highly coordinated processes with an extensive reprogramming of whole-plant metabolism. The findings of this study will enable us to design effective breeding and genetic engineering strategies to enhance symbiotic efficiency in legume crops.phosphate deficiency | nitrogen fixation | metabolomics | carbon and nitrogen metabolism | phosphate homeostasis P hosphorus plays a critical role in numerous plant metabolic processes and contributes to the biosynthesis of cellular macromolecules, such as ATP, nucleic acids, phospholipids, and phosphorylated sugars (1). Thus, phosphorus has been established as one of the most important elements required for normal plant growth and development (1). Unfortunately, limited availability of inorganic phosphate (Pi), which is the only absorbable form of phosphorus for plants, in soils is nearly universal, because Pi readily forms various insoluble compounds with metals, such as calcium and iron in alkaline and acidic soils, respectively (1, 2). Pi deficiency can be overcome by the application of Pi fertilizers; however, the excessive use of chemical fertilizers can have serious environmental consequences, including the contamination of soil and water resources (1). Additionally, the global demand for and use of Pi fertilizers are projected to increase significantly with the explosive growth of the global population.Thus, it has been predicted that global Pi res...

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Section: Methodsmentioning

confidence: 99%

Adaptation of the symbiotic Mesorhizobium –chickpea relationship to phosphate deficiency relies on reprogramming of whole-plant metabolism

Esfahani

Kusano

Nguyen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Segments of ∼3 cm (∼30 mg fresh weight) were harvested from the midsection of the uppermost fully expanded leaf (10th leaf) at noon on a sunny day and were immediately frozen in liquid nitrogen. Metabolite extraction, sample preparation, and gas chromatography-time-of-flight/mass spectrometry (GC-TOF/MS) were performed as described previously (36). Samples corresponding to 5 mg fresh weight were subjected to the analysis…”

Section: (Si Text)mentioning

confidence: 99%

Phosphoenolpyruvate carboxylase intrinsically located in the chloroplast of rice plays a crucial role in ammonium assimilation

Masumoto

Miyazawa

Ohkawa

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Phosphoenolpyruvate carboxylase (PEPC) is a key enzyme of primary metabolism in bacteria, algae, and vascular plants, and is believed to be cytosolic. Here we show that rice (Oryza sativa L.) has a plant-type PEPC, Osppc4, that is targeted to the chloroplast. Osppc4 was expressed in all organs tested and showed high expression in the leaves. Its expression in the leaves was confined to mesophyll cells, and Osppc4 accounted for approximately one-third of total PEPC protein in the leaf blade. Recombinant Osppc4 was active in the PEPC reaction, showing V max comparable to cytosolic isozymes. Knockdown of Osppc4 expression by the RNAi technique resulted in stunting at the vegetative stage, which was much more marked when rice plants were grown with ammonium than with nitrate as the nitrogen source. Comparison of leaf metabolomes of ammonium-grown plants suggested that the knockdown suppressed ammonium assimilation and subsequent amino acid synthesis by reducing levels of organic acids, which are carbon skeleton donors for these processes. We also identified the chloroplastic PEPC gene in other Oryza species, all of which are adapted to waterlogged soil where the major nitrogen source is ammonium. This suggests that, in addition to glycolysis, the genus Oryza has a unique route to provide organic acids for ammonium assimilation that involves a chloroplastic PEPC, and that this route is crucial for growth with ammonium. This work provides evidence for diversity of primary ammonium assimilation in the leaves of vascular plants.amino acid synthesis | glycolysis | nitrogen assimilation | organic acid synthesis | Oryza

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“…Therefore, various separation and detection techniques, such as GC-MS [31,65], liquid chromatography (LC)-MS [62,70], CE-MS [83,92], Fourier transform ion cyclotron resonance (FT-ICR) MS [1,73,75] and NMR [61,85] have been applied to metabolomics. Among them, GC-MS is the most popular method in current metabolomic studies [31].…”

Section: Analytical Technologiesmentioning

confidence: 99%

“…There have been many reports on the application of PCA to metabolomic data [7,12,19,54,65,71,75,93,94,95]. In addition, several statistical analytical methods have been used for analysis of metabolomic datasets; for example, hierarchical cluster analysis (HCA) [39,78], partial least squares discriminant analysis (PLS-DA) [52,65], and batch-learning self-organizing map (BL-SOM) [43,44,54]. Depending on the objective of each study, the most appropriate statistical analytical method should be exploited to analyze the metabolomic data.…”

Section: Informatics Techniquesmentioning

confidence: 99%

“…Kusano et al [65] used a metabolomic method combining one-dimensional (1D) and two-dimensional (2D) GC-TOF MS for metabolic phenotyping of the 68 rice varieties in the world rice core collection (WRC) and two other varieties. The WRC is a representative set of the cultivated rice (Oriza sativa L.) and covers approximately 90% of the DNA polymorphisms detected in the original population [57].…”

Section: Metabolomics For Rice Biologymentioning

confidence: 99%

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Rice Metabolomics

Oikawa

Mizutani

Kusano

et al. 2008

Rice

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Metabolomics is a recently developed technology for comprehensive analysis of metabolites in organisms. Plant metabolites that are produced for the growth, development, and chemical defense of plants against climatic alterations or natural predators are also useful to us as nutrients or medicines; hence, it is important to comprehend the amounts and varieties of plant metabolites. Besides providing an understanding of the metabolic state in plants under various circumstances, metabolomic techniques are applicable to the clarification of the functions of unknown genes by using natural variants or mutants of the target plants. Furthermore, a metabolomic approach might be useful in the breeding of crops, since valuable plant traits such as taste and yield are closely related to metabolic conditions. Here, we describe the methodology of metabolomics including sample extraction and preparation, metabolite detection, and data processing and analysis, and introduce the application of metabolomic studies to rice.

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Application of a metabolomic method combining one-dimensional and two-dimensional gas chromatography-time-of-flight/mass spectrometry to metabolic phenotyping of natural variants in rice

Cited by 160 publications

References 45 publications

Adaptation of the symbiotic Mesorhizobium –chickpea relationship to phosphate deficiency relies on reprogramming of whole-plant metabolism

Adaptation of the symbiotic Mesorhizobium –chickpea relationship to phosphate deficiency relies on reprogramming of whole-plant metabolism

Phosphoenolpyruvate carboxylase intrinsically located in the chloroplast of rice plays a crucial role in ammonium assimilation

Rice Metabolomics

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