Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches
Abstract
:1. Introduction
2. Discussion
2.1. Role of Metabolism in NUE
2.2. Effect of Transgene Expression on Nitrogen Metabolism
2.3. Metabolomics Technology
2.4. Metabolic Flux Analysis
2.5. Modeling Fluxes in NUE
3. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Species | Synechocystis sp. PC 6803 | Chlamydomos reinhardtii | Chlamydomonas reinhardtii | Arabidopsis thaliana | Zea mays | Zea mays | |||
---|---|---|---|---|---|---|---|---|---|
N depletion condition: length of time and concentration shift | 4 h; 5 to 0 mM NH4+ | 24 h; 7 to 0 mM NH4+ | 1, 2, 6 days; 7.48 to 0 mM NH4+ | 0, 2, 10 days; 6 to 0 mM NO3−) | Inbred lines A188, B73; 0.15 or 15 mM NO3− | Inbred line B73 0.1 or 10 mM NO3− | |||
Technique | CE-MS, LC-MS/MS | GC-TOF-MS | GC-MS | GC-MS, anion HPLC | GC-MS | GC-MS | |||
Reference | [32] | [33] | [34] | [35] | [26] | [11] | |||
Metabolite | 1–2–6d | Shoot | Root | A188 | B73 | Veg. | Mat. | ||
Amino Acids (percentages, %) | |||||||||
Alanine | 200 | 6 | 44–87–64 | 66–27 | 86–84 | 11–15 | 28–40 | 18 | 18 |
Arginine | 48 | 129–97–65 | 14–51 | 97–82 | 26 | ||||
Asparagine | 55 | 18 | 64–20 | 163–24 | 11–85 | 32–78 | 1 | ||
Aspartate | 21 | 63 | 21–17–14 | 44–27 | 48–32 | 28–52 | 45–163 | 4 | |
Cysteine | 135 | 58–57 | 72–83 | ||||||
Glutamate | 88 | 18 | 21–72–57 | 68–54 | 82–29 | 27–50 | 57–62 | 12 | |
Glutamine | 43 | 10 | 58–50–56 | 25–96 | 3 | ||||
Glycine | 262 | 35 | 76–161–86 | 56–22 | 178–706 | 8–38 | 21–131 | 3 | |
Histidine | 136 | 55 | |||||||
Isoleucine | 576 | 250 | 38–42–29 | 116–67 | 103–90 | 48–40 | 61–49 | 14 | |
Leucine | 688 | 59 | 25–21–14 | 139–70 | 97–78 | 56–38 | 84–34 | 40 | |
Lysine | 124 | 42 | 28–62–58 | 84–37 | 106–72 | 77–84 | 198–91 | 26 | 54 |
Methionine | 241 | 18 | 146–201–215 | 32 | |||||
Phenylalanine | 433 | 44 | 16–25–11 | 129–66 | 73–44 | 49–42 | 64–40 | 19 | |
Proline | 198 | 34 | 71–49–95 | 72–23 | 73–38 | 40–57 | 70–65 | 8 | |
Serine | 472 | 14 | 21–61–42 | 134–75 | 86–57 | 11–31 | 25–86 | 3 | |
Threonine | 349 | 246 | 38–187–95 | 81–46 | 97–46 | 25–28 | 47–74 | 6 | 26 |
Tryptophan | 83 | 495 | 8–24–6 | 88–53 | 74–52 | 62 | |||
Tyrosine | 1284 | 111 | 24–51–26 | 171–92 | 103–90 | 40–54 | 71–61 | 21 | 40 |
Valine | 235 | 33 | 40–57–50 | 91–66 | 103–90 | 44–47 | 58–61 | 10 | |
Organic Acids (percentages, %) | |||||||||
Aconitate | 115 | 55–30 | 102–65 | 45 | 45 | ||||
Benzoate | 73 | 43–114–102 | 92–85 | 64–90 | |||||
Citrate | 59 | 56 | 133–1478–1134 | 59–25 | 156–45 | 886 | |||
Erythonate | 134–242 | 113–120 | 50 | 48 | |||||
Fumarate | 1015 | 16 | 33–125–95 | 442–397 | 94–67 | 41–78 | 51–68 | 52 | 135 |
Glycerate | 141 | 58 | 40–48–46 | 172–67 | 1209–5301 | ||||
2-oxoglutarate | 360 | 46 | 95–45–114 | 105–86 | 152–97 | 80–95 | 108–45 | ||
Lactate | 39–63–54 | 109–130 | 22–52 | ||||||
Malate | 876 | 26 | 34–87–82 | 113–97 | 993–461 | 34–20 | 23–23 | 61 | |
Maleate | 28–56–148 | 157–182 | 84–85 | 900 | |||||
Oxaloacetate | 94 | 1–36–1 | 160–83 | 125–192 | |||||
Pyruvate | 334 | 25 | 21–27–100 | 81–75 | 71–106 | 75–50 | 81–34 | 11 | |
Shikimate | 168 | 40 | 103–77 | 160–53 | 170–216 | 131–312 | 26 | ||
Succinate | 398 | 81 | 97–216–167 | 178–346 | 114–67 | ||||
Threonate | 39 | 96–131–111 | 99–132 | 267–238 | |||||
Alcohols and Sugars (percentages, %) | |||||||||
Glycerol | 77 | 1.6–0.9–0.5 | 100–93 | 64–70 | |||||
Inositol | 13 | 30–52–84 | 97–74 | 177–258 | 67 | ||||
Fructose | 312–202–97 | 462–218 | 687–277 | 44–34 | 29–31 | 11 | |||
Galactose | 17–7–7 | 343–487 | 208–225 | 30 | 15 | ||||
Glucose | 27–10–3 | 405–545 | 413–515 | 30–21 | 29–24 | 6 | |||
Maltose | 62 | 95–93 | 79–86 | 121 | |||||
Mannose | 223–184 | 132–362 | 16 | 24 | |||||
Raffinose | 183 | 973–7981 | 198–313 | 275–268 | 270–159 | 333 | |||
Sucrose | 89–89 | 99–110 | 71 | ||||||
Xylose | 67–195–231 | 119–149 | 271–357 | ||||||
Phosphorylated Compounds (percentages, %) | |||||||||
6-phosphogluconic acid | 136 | 6 | 120–177–171 | ||||||
Fructose-6P: Fru-6P | 148 | 64 | 20–44–73 | 65–55 | 76–65 | 72–296 | 137–526 | 21 | |
Fructose-1,6-bisP | 82 | 67 | |||||||
Glucose-1-P | 119 | 61 | |||||||
Glucose-6-P | 148 | 89 | 53–99–44 | 70–57 | 88–84 | 77–356 | 131–559 | 14 | |
Glycerate-3P | 139 | 161 | 52–230–83 | 11 | 150 | ||||
myo-inositol-P | 108–83–71 | 148–77 | 66–70 | ||||||
Phosphoenol-pyruvate | 104 | 19 | |||||||
Ribulose-5P | 127 | 99 | 69–174 | 100–333 | |||||
Nitrogenous Compounds (percentages, %) | |||||||||
γ-aminobutyric acid | 536 | 167–114–43 | 204–138 | 217–96 | 29–25 | 38–39 | 8 | ||
Adenine | 100 | 9 | 24–52–56 | ||||||
Citrulline | 23 | 23 | 11–73 | 23–138 | |||||
Hydroxylamine | 139 | 114–81–72 | 59–8 | 19–34 | |||||
Ornithine | 21 | 6 | 127–87–59 | 10–72 | 48–94 | ||||
Putrescine | 9 | 11–13–8 | 12 | 9 | |||||
Uracil | 10 | 13–17–18 |
Genetic Construct | Conditions | Technique | Core Metabolomic Results (Compared to WT) | References |
---|---|---|---|---|
N metabolism | ||||
Oryza sativa GS1;1 and GS1;2 overexpressed in Oryza sativa cv. Zhonghua 11 under the control of the CaMV 35S promoter | Metabolic analysis done on tillering stage roots and shoots of plants growth with Low N and Moderate N | GC-TOF-MS | Low N: GS1;1 and GS1; 2 increased sugars, organic acids, free amino acids in shoots and decreased in roots. Moderate N: same results for both lines in shoots as for low N, in roots GS1;1 increased sugars, organic acids and free amino acids GS1;2 roots had decreased metabolites. | [41] |
Pisum sativum AS1 overexpressed in Nicotiana tabacum under the control of the CaMV 35S promoter | 16 h light/8 h dark, 21 day old plants grown in sand, fertilized with Hoagland solution with 10 mM NO3− | HPLC | 10–100 fold increased Asn. Decreased Gln, Asp. No change in Glu. | [46] |
Hordeum vulgare AlaAT overexpressed in Oryza sativa under the control of the root-specific OsANT1 promoter | 14 h light/10 h dark, 45 day old plants grown hydroponically in 0.5, 2.0, and 5.0 mM NH4+ | HPLC | Increased Gln, Glu, Asn, Asp, and Arg in roots and shoots. | [12] |
N recycling/protein degradation/C:N balance | ||||
Mus musculus ODC overexpressed in Populus nigra under the control of a 2X CaMV 35S promoter | Cell cultures grown in MS media | HPLC | Increased Ala, Thr, Val, Ile, and GABA. Decreased Gln, Glu, Orn, Arg, His, Ser, Gly, Cys, Phe, Trp, Asp, Lys, Leu, Met. | [53] |
Arabidopsis FUM2 overexpressed in Arabidopsis under the control of a 2X CaMV 35S promoter | 8 h light/16 h dark, plants grown for 42 days with 1.25 mg (low) or 31.5 mg (high) inorganic nitrogen | GC-MS | Increased starch, FUM2 knockouts reduced fumarate levels, varied amino acid levels according to light cycle. | [54] |
Regulatory transgenes | ||||
Zea mays Dof1 expressed in Arabidopsis under the control of the CaMV 35S promoter; also expressed in potato | Constant light, plants grown on modified MS medium; low N = 1 mM NH4NO3/1 mM KNO3; high N = 10 mM NH4NO3/10 mM KNO3 | Hitachi amino acid analyzer; enzymatic assay | Increased total [amino acid], NH4+ Decreased glucose, malate No change in sucrose, citrate, or 2-OG Similar to transgenic potato | [48] |
Zea mays Dof1 expressed in Oryza sativa under the control of the CaMV 35S promoter | 14 h day/10 h night, hydroponic growth at 360 (high) or 90 µM (low) NH4+ | CE-MS/MS | Increased concentration of some amino acids under high and low [N] | [49] |
N-responsive transgenes | ||||
Oryza sativa ENOD93 expressed in Oryza sativa under the control of the 35S C4PDK promoter | 16 h day/8 h night for 4 weeks then 10 h day/14 h night for 1 week for flowering, soil growth at 1 mM (low), 5 mM (median) or 10 mM (high) nitrate | Biochemical assays | Increased total amino acids in OsENOD93-ox line roots in all N levels but more so under N stress. No increase in amino acid levels in shoots. Higher biomass in OsENOD93-ox. | [52] |
Co-expressed N metabolism and Regulatory transgenes | ||||
Arabidopsis Dof1, GS1, GS2 expressed in tobacco under the control of the leaf specific rbcS promoter from tomato | Growth in perlite and low N nutrient solution for 60 and 90 days | RP-HPLC and biochemical assays | Transgenic tobacco co-expressing Dof1, GS1, GS2 had increased amino acids, glucose, sucrose and decreased nitrate, malic acid, citric acid and showed growth advantages | [43] |
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Beatty, P.H.; Klein, M.S.; Fischer, J.J.; Lewis, I.A.; Muench, D.G.; Good, A.G. Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches. Plants 2016, 5, 39. https://doi.org/10.3390/plants5040039
Beatty PH, Klein MS, Fischer JJ, Lewis IA, Muench DG, Good AG. Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches. Plants. 2016; 5(4):39. https://doi.org/10.3390/plants5040039
Chicago/Turabian StyleBeatty, Perrin H., Matthias S. Klein, Jeffrey J. Fischer, Ian A. Lewis, Douglas G. Muench, and Allen G. Good. 2016. "Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches" Plants 5, no. 4: 39. https://doi.org/10.3390/plants5040039