Skip to main content
Log in

Overexpression of Yeast Arabinono-1,4-Lactone Oxidase Gene (ALO) Increases Tolerance to Oxidative Stress and Al Toxicity in Transgenic Tobacco Plants

  • Original Paper
  • Published:
Plant Molecular Biology Reporter Aims and scope Submit manuscript

Abstract

l-Ascorbic acid (AsA) is the most abundant antioxidant and a major redox buffer that regulates plant responses to environmental stresses. AsA is also a precursor of oxalate in plants, while oxalate is associated with aluminum (Al) tolerance in some plant species. To enhance AsA in plants and increase tolerance against oxidative stress and Al toxicity, the yeast d-arabinono-1,4-lactone oxidase (ALO) gene was overexpressed in transgenic tobacco plants. Overexpression of ALO promoted synthesis of AsA and oxalate in leaves, but did not affect oxalate level in roots. Compared to the wild type, transgenic plants maintained higher levels of AsA, glutathione, maximal photochemical efficiency (F v/F m) and the actual PSII efficiency (ΦPSII) of photosystem II (PSII), and non-photochemical quenching (NPQ), and accumulated less H2O2 and malondialdehyde (MDA) in leaves in response to methyl viologen- and high-light-induced oxidative stress. In addition, transgenic plants showed higher AsA level and fresh weight of shoot and roots and lower levels of Al, H2O2, and MDA in root tips after Al treatment; however, there is no significant difference in organic acid (citrate and oxalate) exudation between the wild-type and transgenic plants in response to Al treatment. The results suggest that AsA and oxalate could be up-regulated by overexpressing ALO. The elevated AsA increased oxidative tolerance due to scavenging of reactive oxygen species (ROS) and induction of NPQ in leaves and enhanced Al tolerance as an antioxidant for avoiding Al-triggered oxidative damages in roots.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10

Similar content being viewed by others

References

  • Agius F, González-Lamothe R, Caballero JL, Muñosz-Blanco J, Botella MA, Valpuesta V (2003) Engineering increased vitamin C levels in plants by overexpression of a D-galacturonic acid reductase. Nat Biotech 21:177–181

    Article  CAS  Google Scholar 

  • Anoop VM, Basu U, McCammon MT, McAlister-Henn L, Taylor GJ (2003) Modulation of citrate metabolism alters aluminum tolerance in yeast and transgenic canola overexpressing a mitochondrial citrate synthase. Plant Physiol 132:205–2217

    Article  Google Scholar 

  • Bilger W, Björkman O (1990) Role of the xanthophylls cycle in photoprotection elucidated by measurements of light-induced absorbance changes. Photosynth Res 25:173–185

    Article  CAS  PubMed  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantification of microgram quantities of proteins utilizing the principle of protein-dye binding. Anal Biochem 72:48–254

    Article  Google Scholar 

  • Bulley SM, Rassam M, Hoser D, Otto W, Schünemann N, Wright M, MacRae E, Gleave A, Laing W (2009) Gene expression studies in kiwifruit and gene over-expression in Arabidopsis indicates that GDP-L-galactose guanyltransferase is a major control point of vitamin C biosynthesis. J Exp Bot 60:765–778

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Chen Z, Gallie DR (2005) Increasing tolerance to ozone by elevating foliar ascorbic acid confers greater protection against ozone than increasing avoidance. Plant Physiol 138:1673–1689

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Conklin PL, Barth C (2004) Ascorbic acid, a familiar small molecule intertwined in the response of plants to ozone, pathogens, and the onset of senescence. Plant, Cell Environ 27:959–970

    Article  CAS  Google Scholar 

  • Conklin PL, Norris SR, Wheeler GL, Williams EH, Smirnoff N, Last RL (1999) Genetic evidence for the role of GDP-mannose in plant ascorbic acid (vitamin C) biosynthesis. Proc Natl Acad Sci U S A 96:4198–4203

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Conklin PL, Gatzek S, Wheeler GL, Dowdle J, Raymond MJ, Rolinski S, Isupov M, Littlechild JA, Smirnoff N (2006) Arabidopsis thaliana VTC4 encodes L-galactose-1-P phosphatase, a plant ascorbic acid biosynthetic enzyme. J Biol Chem 281:15662–15670

    Article  CAS  PubMed  Google Scholar 

  • de la Fuente JM, Ramirez-Rodrigez V, Cabrera-Ponce JS, Herrera-Esterella L (1997) Aluminum tolerance in transgenic plants by alteration in citrate synthesis. Science 276:1566–1568

    Article  PubMed  Google Scholar 

  • Debolt S, Melino V, Ford C (2007) Ascorbate as a biosynthetic precursor in plants. Ann Bot 99:3–8

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Delhaize E, Craig S, Beaton CD, Bennet RJ, Jagadish VC, Randall PJ (1993) Aluminum tolerance in wheat (Triticum aestivum L.): I. Uptake and distribution of aluminum in root apices. Plant Physiol 103:685–693

    CAS  PubMed Central  PubMed  Google Scholar 

  • Delhaize E, Hebb DM, Ryan PR (2001) Expression of a Pseudomonas aeruginosa citrate synthase gene in tobacco is not associated with either enhanced citrate accumulation or efflux. Plant Physiol 125:2059–2067

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Deng W, Luo K, Li Z, Yang Y, Hu N, Wu Y (2009) Overexpression of Citrus junos mitochondrial citrate synthase gene in Nicotiana benthamiana confers aluminum tolerance. Planta 230:355–365

    Article  CAS  PubMed  Google Scholar 

  • Devi SR, Yamamoto Y, Matsumoto H (2003) An intracellular mechanism of aluminum associated with high antioxidant status in cultured tobacco cells. J Inorg Biochem 97:59–68

    Article  CAS  PubMed  Google Scholar 

  • Dong D, Peng X, Yan X (2004) Organic acid exudation induced by phosphorus deficiency and/or aluminium toxicity in two contrasting soybean genotypes. Physiol Plant 122:190–199

    Article  CAS  Google Scholar 

  • Dowdle J, Ishikawa T, Gatzek S, Rolinski S, Smirnoff N (2007) Two genes in Arabidopsis thaliana encoding GDP-L-galactose phosphorylase are required for ascorbate biosynthesis and seedling viability. Plant J 52:673–689

    Article  CAS  PubMed  Google Scholar 

  • Eltayeb AE, Kawano N, Badawi GH, Kaminaka H, Sanekata T, Morishima I, Shibahara T, Inanaga S, Tanaka K (2006) Enhanced tolerance to ozone and drought stresses in transgenic tobacco overexpressing dehydroascorbate reductase in cytosol. Physiol Plant 127:57–65

    Article  CAS  Google Scholar 

  • Foyer CH, Noctor G (1999) Leaves in the dark see in the light. Science 284:599–601

    Article  CAS  PubMed  Google Scholar 

  • Foyer CH, Noctor G (2005) Oxidant and antioxidant signaling in plants: a re-evaluation of the concept of oxidative stress in a physiological context. Plant, Cell Environ 28:1056–1071

    Article  CAS  Google Scholar 

  • Gallie DR (2013) L-Ascorbic acid: a multifunctional molecule supporting plant growth and development. Scientifica 2013:1–24

    Article  Google Scholar 

  • Gatzek S, Wheeler GL, Smirnoff N (2002) Antisense suppression of L-galactose dehydrogenase in Arabidopsis thaliana provides evidence for its role in ascorbate synthesis and reveals light modulated L-galactose synthesis. Plant J 30:541–553

    Article  CAS  PubMed  Google Scholar 

  • Genty B, Briantais JM, Baker N (1989) The relationship between the quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence. Biochim Biophys Acta 990:87–92

    Article  CAS  Google Scholar 

  • Green MA, Fry SC (2005) Vitamin C degradation in plant cells via enzymatic hydrolysis of 4-O-oxalyl-l-threonate. Nature 433:83–87

    Article  CAS  PubMed  Google Scholar 

  • Guo Z, Tan H, Zhu Z, Lu S, Zhou B (2005) Effect of intermediates on ascorbic acid and oxalate biosynthesis of rice and in relation to its stress resistance. Plant Physiol Biochem 43:955–962

    Article  CAS  PubMed  Google Scholar 

  • Hemavathi U, Upadhyaya CP, Akula N, Young KE, Chun SC, Kim DH, Park SW (2010) Enhanced ascorbic acid accumulation in transgenic potato confers tolerance to various abiotic stresses. Biotechnol Lett 32:321–330

    Article  CAS  PubMed  Google Scholar 

  • Huh W-K, Kim S-T, Yang K-S, Seok Y-J, Hah YC, Kang S-O (1994) Characterization of D-arabinono-1,4-lactone oxidase from Candida albicans ATCC 10231. Eur J Biochem 225:1073–1079

    Article  CAS  PubMed  Google Scholar 

  • Huh W-K, Lee B-H, Kim S-T, Kim Y-R, Rhie G-E, Baek YW, Hwang C-S, Lee J-S, Kang S-O (1998) D-Erythroascorbic acid is an important antioxidant molecule in Saccharomyces cerevisiae. Mol Microbiol 30:895–903

    Article  CAS  PubMed  Google Scholar 

  • Ivanov B, Edwards G (2000) Influence of ascorbate and the Mehler peroxidase reaction on non-photochemical quenching of chlorophyll fluorescence in maize mesophyll chloroplasts. Planta 210:765–774

    Article  CAS  PubMed  Google Scholar 

  • Jones DL, Blancaflor EB, Kochian LV, Gilroy S (2006) Spatial coordination of aluminium uptake, production of reactive oxygen species, callose production and wall rigidication in maize roots. Plant, Cell Environ 29:1309–1318

    Article  CAS  Google Scholar 

  • Kochian LV, Hoekenga OA, Pineros MA (2004) How do crop plants tolerate acid soils? Mechanisms of aluminum tolerance and phosphorous efficiency. Annu Rev Plant Biol 55:459–493

    Article  CAS  PubMed  Google Scholar 

  • Kostman TA, Tarlyn NM, Loewus FA, Franceschi VR (2001) Biosynthesis of L-ascorbic acid and conversion of carbons 1 and 2 of L-ascorbic acid to oxalic acid occurs within individual calcium oxalate crystal idioblasts. Plant Physiol 125:634–640

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Kuo MC, Kao CH (2003) Aluminum effects on lipid peroxidation and antioxidative enzyme activities in rice leaves. Biol Plant 46:149–152

    Article  CAS  Google Scholar 

  • Lee B-H, Hu W-K, Kim S-T, Lee J-S, Kang S-O (1999) Bacterial production of D-erythroascorbic acid and L-ascorbic acid through functional expression of Saccharomyces cerevisiae D-arabinono-1,4-lactone oxidase in Escherichia coli. Appl Environ Microbiol 65:4685–4687

    CAS  PubMed Central  PubMed  Google Scholar 

  • Linster CL, Gomez TA, Christensen KC, Adler LN, Young BD, Brenner C, Clarke SG (2007) Arabidopsis VTC2 encodes a GDP-L-galactose phosphorylase, the last unknown enzyme in the Smirnoff-Wheeler pathway to ascorbic acid in plants. J Biol Chem 282:18879–18885

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Liu W, An H-M, Yang M (2013) Overexpression of Rosa roxburghii L-galactono-1,4-lactone dehydrogenase in tobacco plant enhances ascorbate accumulation and abiotic stress tolerance. Acta Physiol Plant 35:1617–1624

    Article  CAS  Google Scholar 

  • Loewus FA (1999) Biosynthesis and metabolism of ascorbic acid in plants and of analogs of ascorbic acid in fungi. Phytochemistry 52:193–210

    Article  CAS  Google Scholar 

  • Ma JF, Zheng SJ, Hiradate S, Matsumoto H (1997) Detoxifying aluminum with buckwheat. Nature 390:569–570

    Article  Google Scholar 

  • Melino VJ, Soole KL, Ford CM (2009) Ascorbate metabolism and the developmental demand for tartaric and oxalic acids in ripening grape berries. BMC Plant Biol 9:145. doi:10.1186/1471-2229-9-145

    Article  PubMed Central  PubMed  Google Scholar 

  • Müller-Moulé P, Conklin PL, Niyogi KK (2002) Ascorbate deficiency can limit violaxanthin de-epoxidase activity in vivo. Plant Physiol 128:193–210

    Article  Google Scholar 

  • Müller-Moulé P, Havaux M, Niyogi KK (2003) Zeaxanthin deficiency enhances the high light sensitivity of an ascorbate-deficient mutant of Arabidopsis. Plant Physiol 133:748–760

    Article  PubMed Central  PubMed  Google Scholar 

  • Murray MG, Thompson WF (1980) Rapid isolation of high molecular weight DNA. Nucleic Acids Res 8:4321–4325

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Navascués J, Pérez-Rontomé C, Sánchez DH, Staudinger C, Wienkoop S, Rellán-Á lvarez R, Becana M (2011) Oxidative stress is a consequence, not a cause, of aluminum toxicity in the forage legume Lotus corniculatus. New Phytol 193:625–636

    Article  PubMed  Google Scholar 

  • Nishikimi M, Noguchi E, Yagi K (1978) Occurrence in yeast of L-galactonolactone oxidase which is similar to a key enzyme for ascorbic acid biosynthesis in animals, L-gulonolactone oxidase. Arch Biochem Biophys 191:479–486

    Article  CAS  PubMed  Google Scholar 

  • Orozco-Cárdenas ML, Ryan C (1999) Hydrogen peroxide is generated systemically in plant leaves by wounding and systemin via the octadecanoid pathway. Proc Natl Acad Sci U S A 96:6553–6557

    Article  PubMed Central  PubMed  Google Scholar 

  • Running JA, Burlingame RP, Berry A (2003) The pathway of L-ascorbic acid biosynthesis in the colourless microalga Prototheca moriformis. J Exp Bot 54:1841–1849

    Article  CAS  PubMed  Google Scholar 

  • Ryan PR, Delhaize E, Jones DL (2001) Function and mechanism of organic anion exudation from plant roots. Annu Rev Plant Physiol Plant Mol Biol 52:527–560

    Article  CAS  PubMed  Google Scholar 

  • Schreiber U, Schliwa U, Bilger W (1986) Continuous recording of photochemical and non-photochemical chlorophyll fluorescence quenching with a new type of modulation fluorometer. Photosynth Res 10:51–62

    Article  CAS  PubMed  Google Scholar 

  • Sharma P, Dubey RS (2007) Involvement of oxidative stress and role of antioxidative defense system in growing rice seedlings exposed to toxic concentrations of aluminum. Plant Cell Rep 26:2027–2038

    Article  CAS  PubMed  Google Scholar 

  • Smirnoff N, Wheeler GL (2000) Ascorbic acid in plants: biosynthesis and function. CRC Cri Rev in Plant Sci 19:267–290

  • Smith IK (1985) Stimulation of glutathione synthesis in photorespiring plants by catalase inhibitors. Plant Physiol 79:1044–1047

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Tahara K, Yamanoshita T, Norisada M, Hasegawa I, Kashima H, Sasaki S, Kojima K (2008) Aluminum distribution and reactive oxygen species accumulation in root tips of two Melaleuca trees differing in aluminum resistance. Plant Soil 307:167–178

    Article  CAS  Google Scholar 

  • Takahama U, Oniki T (1992) Regulation of peroxidase-dependent oxidation of phenolics in the apoplast of spinach leaves by ascorbate. Plant Cell Physiol 33:379–384

    CAS  Google Scholar 

  • Tokuna T, Miyahara K, Tabata K, Esaka M (2005) Generation and properties of ascorbic acid-over-producing transgenic tobacco cells expressing sense RNA for L-galactono-1,4-lactone dehydrogenase. Planta 220:854–863

    Article  Google Scholar 

  • Valpuesta V, Botella MA (2004) Biosynthesis of L-ascorbic acid in plants: new pathways for an old antioxidant. Trends Plant Sci 9:573–577

    Article  CAS  PubMed  Google Scholar 

  • Wan X, Tan J, Lu S, Lin C, Hu Y, Guo Z (2009) The increased tolerance to oxidative stress in transgenic tobaccos expressing a wheat oxalate oxidase gene via induction of antioxidant enzymes is mediated by H2O2. Physiol Plant 136:30–44

    Article  CAS  PubMed  Google Scholar 

  • Wang J-W, Kao CH (2007) Protective effect of ascorbic acid and glutathione on AlCl3-inhibited growth of rice roots. Biol Plant 51:493–500

    Article  CAS  Google Scholar 

  • Wang Q, Yi Q, Hu Q, Zhao Y, Nian H, Li K, Yu Y, Izui K, Chen L (2012) Simultaneous overexpression of citrate synthase and phosphoenolpyruvate carboxylase in leaves augments citrate exclusion and Al resistance in transgenic tobacco. Plant Mol Biol Rep 30:992–1005

    Article  CAS  Google Scholar 

  • Wheeler GL, Hibes MA, Smirnoff N (1998) The biosynthetic pathway of vitamin C in higher plants. Nature 393:365–369

  • Yamamoto Y, Kobayashi Y, Matsumoto H (2001) Lipid peroxidation is an early symptom triggered by aluminum, but not the primary cause of elongation inhibition in pea roots. Plant Physiol 125:199–208

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Yamamoto Y, Kobayashi Y, Devi SR, Rikiishi S, Matsumoto H (2002) Aluminum toxicity is associated with mitochondrial dysfunction and the production of reactive oxygen species in plant cells. Plant Physiol 128:63–72

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Yin L, Wang S, Eltayeb AE, Uddin MI, Yamamoto Y, Tsuji W, Takeuchi Y, Tanaka K (2010) Overexpression of dehydroascorbate reductase, but not monodehydroascorbate reductase, confers tolerance to aluminum stress in transgenic tobacco. Planta 231:609–621

    Article  CAS  PubMed  Google Scholar 

  • Zhang Z, Wang J, Zhang R, Huang R (2012) The ethylene response factor AtERF98 enhances tolerance to salt through the transcriptional activation of ascorbic acid synthesis in Arabidopsis. Plant J 71:273–287

    Article  CAS  PubMed  Google Scholar 

  • Zhuo C, Wang T, Lu S, Zhao Y, Li X, Guo Z (2013) A cold responsive galactinol synthase gene from Medicago falcata (MfGolS1) is induced by myo-inositol and confers multiple tolerances to abiotic stresses. Physiol Plant 149:67–78

Download references

Acknowledgments

This work was supported financially by the National Natural Science Foundation of China (30270128) and Guangdong Provincial Natural Science Foundation (010594).

Author information

Authors and Affiliations

Authors

Corresponding authors

Correspondence to Shaoyun Lu or Zhenfei Guo.

Additional information

Zhen Chen, Chao Qin, and Ling Lin equally contributed to this work.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chen, Z., Qin, C., Lin, L. et al. Overexpression of Yeast Arabinono-1,4-Lactone Oxidase Gene (ALO) Increases Tolerance to Oxidative Stress and Al Toxicity in Transgenic Tobacco Plants. Plant Mol Biol Rep 33, 806–818 (2015). https://doi.org/10.1007/s11105-014-0794-1

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11105-014-0794-1

Keywords

Navigation