Planta

, Volume 225, Issue 1, pp 153–164 | Cite as

Genetic and transgenic perturbations of carbon reserve production in Arabidopsis seeds reveal metabolic interactions of biochemical pathways

  • Yun Lin
  • Alexander V. Ulanov
  • Vera Lozovaya
  • Jack Widholm
  • Guirong Zhang
  • Jinhua Guo
  • Howard M. Goodman
Original Article

Abstract

The biosynthesis of seed oil and starch both depend on the supply of carbon from the maternal plant. The biochemical interactions between these two pathways are not fully understood. In the Arabidopsis mutant shrunken seed 1 (sse1)/pex16, a reduced rate of fatty acid synthesis leads to starch accumulation. To further understand the metabolic impact of the decrease in oil synthesis, we compared soluble metabolites in sse1 and wild type (WT) seeds. Sugars, sugar phosphates, alcohols, pyruvate, and many other organic acids accumulated in sse1 seeds as a likely consequence of the reduced carbon demand for lipid synthesis. The enlarged pool size of hexose-P, the metabolites at the crossroad of sugar metabolism, glycolysis, and starch synthesis, was likely a direct cause of the increased flow into starch. Downstream of glycolysis, more carbon entered the TCA cycle as an alternative to the fatty acid pathway, causing the total amount of TCA cycle intermediates to rise while moving the steady state of the cycle away from fumarate. To convert the excess carbon metabolites into starch, we introduced the Escherichia coli starch synthetic enzyme ADP-glucose pyrophosphorylase (AGPase) into sse1 seeds. Expression of AGPase enhanced net starch biosynthesis in the mutant, resulting in starch levels that reached 37% of seed weight. However, further increases above this level were not achieved and most of the carbon intermediates remained high in comparison with the WT, indicating that additional mechanisms limit starch deposition in Arabidopsis seeds.

Keywords

Seeds Storage Metabolite profiling Carbon metabolism Oil Starch Arabidopsis thaliana 

Notes

Acknowledgments

We thank Dr. Nicki Engeseth for critical reading of the manuscript. This work was supported by start-up funds from the Department of Crop Sciences, University of Illinois to Y.L., The Illinois Agricultural Experimental Station, and a grant from The Department of Molecular Biology, Massachusetts General Hospital to H.M.G.

Supplementary material

425_2006_337_MOESM1_ESM.doc (38 kb)
Supplementary material

References

  1. Ballicora MA, Iglesias AA, Preiss J (2004) ADP-glucose pyrophosphorylase: a regulatory enzyme for plant starch synthesis. Photosynth Res 79:1–24PubMedCrossRefGoogle Scholar
  2. Cernac A, Benning C (2004) Wrinkled1 encodes an AP2/EREB domain protein involved in the control of storage compound biosynthesis in Arabidopsis. Plant J 40:575–585PubMedCrossRefGoogle Scholar
  3. Cross JM, Clancy M, Shaw JR, Greene TW, Schmidt RR, Okita TW, Hannah LC (2004) Both subunits of ADP-glucose pyrophosphorylase are regulatory. Plant Physiol 135:137–144PubMedCrossRefGoogle Scholar
  4. Dickinson DB, Preiss J (1969) ADP-glucose pyrophosphorylase from maize endosperm. Arch Biochem Biophys 130:119–128PubMedCrossRefGoogle Scholar
  5. Downing WE, Mauxion F, Fauvarque M-O, Reviron M-P, de Vienne D, Vartanian N, Giraudat J (1992) A Brassica napus transcript encoding a protein related to the Kunitz protease inhibitor family accumulates upon water stress in leaves, not seeds. Plant J 2:685–693PubMedCrossRefGoogle Scholar
  6. Emes MJ, Bowsher CG, Hedley C, Burrell MM, Scrase-Field ES, Tetlow IJ (2003) Starch synthesis and carbon partitioning in developing endosperm. J Exp Bot 54:569–575PubMedCrossRefGoogle Scholar
  7. Fiehn O, Kopka J, Trethewey RN, Willmitzer L (2000a) Identification of uncommon plant metabolites based on calculation of elemental compositions using gas chromatography and quadropole mass spectrometry. Anal Chem 72:3573–3580CrossRefGoogle Scholar
  8. Fiehn O, Kopka J, Dormann P, Altmann T, Trethewey RN, Willmitzer L (2000b) Metabolite profiling for plant functional genomics. Nat Biotechnol 18:1157–1161CrossRefGoogle Scholar
  9. Focks N, Benning C (1998) Wrinkled1: a novel, low-seed-oil mutant of Arabidopsis with a deficiency in the seed-specific regulation of carbohydrate metabolism. Plant Physiol 118:91–101PubMedCrossRefGoogle Scholar
  10. Geigenberger P, Kolbe A, Tiessen A (2005) Redox regulation of carbon storage and partitioning in response to light and sugars. J Exp Bot 56:1469–1479PubMedCrossRefGoogle Scholar
  11. Gibon Y, Vigeolas H, Tiessen A, Geigenberger P, Stitt M (2002) Sensitive and high throughput metabolite assays for inorganic pyrophosphate, ADPGlc, nucleotide phosphates, and glycolytic intermediates based on a novel enzymic cycling system. Plant J 30:221–235PubMedCrossRefGoogle Scholar
  12. Gibon Y, Blaesing OE, Hannemann J, Carillo P, Hohne M, Hendriks JH, Palacios N, Cross J, Selbig J, Stitt M (2004) A robot-based platform to measure multiple enzyme activities in Arabidopsis using a set of cycling assays: comparison of changes of enzyme activities and transcript levels during diurnal cycles and in prolonged darkness. Plant Cell 16:3304–3325PubMedCrossRefGoogle Scholar
  13. Ghosh HP, Preiss J (1966) Adenosine diphosphate glucose pyrophosphorylase. A regulatory enzyme in the biosynthesis of starch in spinach leaf chloroplasts. J Biol Chem 241:4491–4504PubMedGoogle Scholar
  14. Ghosh HP, Meyer C, Remy E, Peterson D, Preiss J (1992) Cloning, expression, and nucleotide sequence of glgC gene from an allosteric mutant of Escherichia coli B. Arch Biochem Biophys 296: 122–128PubMedCrossRefGoogle Scholar
  15. Giroux MJ, Shaw J, Barry G, Cobb BG, Greene T, Okita T, Hannah LC (1996) A single mutation that increases maize seed weight. Proc Natl Acad Sci USA 93:5824–5829PubMedCrossRefGoogle Scholar
  16. James MG, Denyer K, Myers AM (2003) Starch synthesis in the cereal endosperm. Curr Opin Plant Biol 6:215–222PubMedCrossRefGoogle Scholar
  17. Kleczkowski LA, Villand P, Preiss J, Olsen OA (1993a) Kinetic mechanism and regulation of ADP-glucose pyrophosphorylase from barley (Hordeum vulgare) leaves. J Biol Chem 268:6228–6233Google Scholar
  18. Kleczkowski LA, Villand P, Luthi E, Olsen OA, Preiss J (1993b) Insensitivity of barley endosperm ADP-glucose pyrophosphorylase to 3-phosphoglycerate and orthophosphate regulation. Plant Physiol 101:179–186CrossRefGoogle Scholar
  19. Kleczkowski LA (1999) A phosphoglycerate to inorganic phosphate ratio is the major factor in controlling starch levels in chloroplasts via ADP-glucose pyrophosphorylase regulation. FEBS Lett 448:153–156PubMedCrossRefGoogle Scholar
  20. Krebbers E, Seurinck J, Cashmore AR, Timko MP (1988) Four genes in two diverged subfamilies encode the ribulose-1,5-biphosphate carboxylase small subunit polypeptides of Arabidopsis thaliana. Plant Mol Biol 11:745–759CrossRefGoogle Scholar
  21. Lee YM, Kumar A, Preiss J (1987) Amino acid sequence of an Escherichia coli ADP-glucose synthetase allosteric mutant as deduced from the DNA sequence of the glgC gene. Nucleic Acids Res 15:10603PubMedGoogle Scholar
  22. Lin Y, Sun L, Nguyen L, Rachubinski RA, Goodman HM (1999) The Pex16p homolog SSE1 and storage organelle formation in Arabidopsis seeds. Science 284:328–330PubMedCrossRefGoogle Scholar
  23. Lin Y, Cluette-Brown JE, Goodman HM (2004) The peroxisome deficient Arabidopsis mutant sse1 exhibits impaired fatty acid synthesis. Plant Physiol 135:814–827PubMedCrossRefGoogle Scholar
  24. Mansfield SG, Briarty LG (1992) Cotyledon cell development in Arabidopsis thaliana during reserve deposition. Can J Bot 70:151–164Google Scholar
  25. Norton G, Harris JF (1975) Compositional changes in developing rape seed (Brassica napas L.). Planta 123:163–174CrossRefGoogle Scholar
  26. Parcy F, Valon C, Raynal M, Gaubier-Comella P, Delseny M, Giraudat J (1994) Regulation of gene expression programs during Arabidopsis seed development: Role of the ABI3 locus and of endogenous abscisic acid. Plant Cell 6:1567–1582PubMedCrossRefGoogle Scholar
  27. Plant AL, van Rooijen GJ, Anderson CP, Moloney MM (1994) Regulation of an Arabidopsis oleosin gene promoter in transgenic Brassica napus. Plant Mol Biol 25:193–205PubMedCrossRefGoogle Scholar
  28. Periappuram C, Steinhauer L, Barton DL, Taylor DC, Chatson B, Zou J (2000) The plastidic phosphoglucomutase from Arabidopsis. A reversible enzyme reaction with an important role in metabolic control. Plant Physiol 122:1193–1199PubMedCrossRefGoogle Scholar
  29. Preiss J, Shen L, Greenberg E, Genter N (1966) Biosynthesis of bacterial glycogen. IV. Activation and inhibition of the adenosine diphosphate glucose pyrophosphorylase of Escherichia coli B. Biochemistry 5:1833–1845PubMedCrossRefGoogle Scholar
  30. Preiss J (1984) Bacterial glycogen synthesis and its regulation. Annu Rev Microbiol 38:419–458PubMedCrossRefGoogle Scholar
  31. Roessner U, Wagner C, Kopka J, Trethewey RN, Willmitzer L (2000) Simultaneous analysis of metabolites in potato tuber by gas chromatography-mass spectrometry. Plant J 23:131–142PubMedCrossRefGoogle Scholar
  32. Salamone PR, Kavakli IH, Slattery CJ, Okita TW (2002) Directed molecular evolution of ADP-glucose pyrophosphorilase. Proc Natl Acad Sci USA 99:1070–1075PubMedCrossRefGoogle Scholar
  33. Sikka VK, Choi S, Kavakli IH, Sakulsingharoj C, Gupta S, Ito H, Okita TW (2001) Subcellular compartmentation and allosteric regulation of the rice endosperm ADP-glucose pyrophosphorylase. Plant Sci 161:461–468CrossRefGoogle Scholar
  34. Smidansky ED, Clancy M, Meyer FD, Lanning SP, Blake NK, Talbert LE, Giroux MJ (2002) Enhanced ADP-glucose pyrophosphorylase activity in wheat endosperm increases seed yield. Proc Natl Acad Sci USA 99:1724–1729PubMedCrossRefGoogle Scholar
  35. Smidansky ED, Martin JM, Hannah LC, Fischer AM, Giroux MJ (2003) Seed yield and plant biomass increases in rice are conferred by deregulation of endosperm ADP-glucose pyrophosphorylase. Planta 216:656–664PubMedGoogle Scholar
  36. Sokolov LN, Dejardin A, Kleczkowski LA (1998) Sugars and light/dark exposure trigger differential regulation of ADP-glucose pyrophosphorylase genes in Arabidopsis thaliana (thale cress). Biochem J 336:681–687PubMedGoogle Scholar
  37. Sowokinos J, Preiss J (1982) Pyrophosphorylases in Solanum tuberosum. III. Purification, physical, and catalytic properties of ADP-glucose pyrophosphorylase in potatoes. Plant Physiol 69:1459–1466PubMedCrossRefGoogle Scholar
  38. Stark DM, Timmerman KP, Barry GF, Preiss J, Kishore GM (1992) Regulation of the amount of starch in plant tissues by ADP-glucose pyrophosphoryalse. Science 258:287–292CrossRefPubMedGoogle Scholar
  39. Tetlow IJ, Morell MK, Emes MJ (2004) Recent developments in understanding the regulation of starch metabolism in higher plants. J Exp Bot 55:2131–2145PubMedCrossRefGoogle Scholar
  40. Vigeolas H, Mohlmann T, Martini N, Neuhaus HE, Geigenberger P (2004) Embryo-specific reduction of ADP-glc pyrophosphorylase leads to an inhibition of starch synthesis and a delay in oil accumulation in developing seeds of oilseed rape. Plant Physiol 136:2676–2686PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Yun Lin
    • 1
  • Alexander V. Ulanov
    • 1
  • Vera Lozovaya
    • 1
  • Jack Widholm
    • 1
  • Guirong Zhang
    • 1
  • Jinhua Guo
    • 1
  • Howard M. Goodman
    • 2
    • 3
  1. 1.Department of Crop SciencesUniversity of IllinoisUrbanaUSA
  2. 2.Department of Molecular BiologyMassachusetts General HospitalBostonUSA
  3. 3.Department of GeneticsHarvard Medical SchoolBostonUSA

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