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Cloning a glutathione peroxidase gene from Nelumbo nucifera and enhanced salt tolerance by overexpressing in rice

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Abstract

A full-length cDNA clone encoding an 866 bp-length glutathione peroxidase protein (NnGPX) was isolated from lotus (Nelumbo nucifera L.). The deduced amino acid sequence of the NnGPX gene had significant homology with ATGPX6. A 3D structural model of the NnGPX was constructed by homology modeling. The cloned NnGPX gene was expressed in Escherichia coli, and a fusion protein of about 40 kDa was detected after isopropyl thiogalactoside induction. Under different concentrations of Na2SeO3 treatments, NnGPX was found to be an enzyme that does not contain selenium. Real-time PCR analysis showed that the NnGPX gene was expressed in all organs of lotus, and its high expression mainly occurred in organs with active metabolisms. NnGPX transcript increased remarkably in response to cold, heat, mechanical damage, and salt treatment. Subsequently, the NnGPX gene was introduced in Oryza sativa cv. Yuetai B. PCR results verified the integration of this gene into the genome of rice and reverse transcription-PCR verified that this gene had been expressed in transgenic rice. The transgenic plants were significantly more tolerant to salt stress compared with the wild-type.

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References

  1. Bowler C, Montagu MV, Inze D (1992) Superoxide dismutase and stress tolerance. Annu Rev Plant Physiol Plant Mol Biol 4:83–116

    Article  Google Scholar 

  2. Fink RC, Scandalios JG (2002) Molecular evolution and structure–function relationships of the superoxide dismutase gene families in angiosperms and their relationship to other eukaryotic and prokaryotic superoxide dismutases. Arch Biochem Biophys 399:19–36

    Article  CAS  PubMed  Google Scholar 

  3. Rotruck JT, Pope AL, Ganther HE, Swanson AB, Hafeman DG, Hoekstra WG (1973) Selenium: biochemical role as a component of glutathione peroxidases. Science 179:588–590

    Article  CAS  PubMed  Google Scholar 

  4. Stadman TC (1996) Selenocysteine. Annu Rev Biochem 65:83–100

    Article  Google Scholar 

  5. David AP, Maarten AP (1982) Extreme longevity of lotus seeds from Pulantien. Nature 299:148–149

    Article  Google Scholar 

  6. Ding YF, Cheng HY, Song SQ (2008) Changes in extreme high-temperature tolerance and activities of antioxidant enzymes of sacred lotus seeds. Sci China C Life Sci 51:842–853

    Article  CAS  PubMed  Google Scholar 

  7. Huang SZ, Tang XJ, Zhang L, Fu JR (2003) Thermotolerance and activity of antioxidative enzymes in lotus seeds. J Plant Physiol Mol Biol 29:421–424

    CAS  Google Scholar 

  8. Sujay R, Atul W, Kakali M, Bishnu PS, Pulok KM (2006) Antioxidant activity of Nelumbo nucifera (sacred lotus) seeds. J Ethnopharmacol 104:322–327

    Article  Google Scholar 

  9. Li Y, Zhang Y, Wei S, Liu L, Chen Y (2012) Recovery of antioxidant gene expression in sacred lotus (Nelumbo nucifera Gaertn) embryonic axes enhances tolerance to extreme high temperature. Afr J Biotechnol 11:12011–12019

    CAS  Google Scholar 

  10. Dong C, Li GL, Li ZQ, Zhu HL, Zhou MQ, Hu ZL (2009) Molecular cloning and expression analysis of an Mn-SOD gene from Nelumbo nucifera. Appl Biochem Biotech 158:605–614

    Article  CAS  Google Scholar 

  11. Dong C, Zheng XF, Li GL, Zhu HL, Zhou MQ, Hu ZL (2011) Molecular cloning and expression of two cytosolic copper-zinc superoxide dismutases genes from Nelumbo nucifera. Appl Biochem Biotechnol 163:679–691

    Article  CAS  PubMed  Google Scholar 

  12. Zheng ZL, Zuo ZY, Liu ZG, Tsai KC, Liu AF, Zou GL (2005) Construction of a 3D model of nattokinase, a novel fibrinolytic enzyme from Bacillus natto: a novel nucleophilic catalytic mechanism for nattokinase. J Mol Graph Model 23:373–380

    Article  CAS  PubMed  Google Scholar 

  13. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  14. Shen G, Pang Y, Wu W, Deng Z, Liu X, Lin J, Zhao L, Sun X, Tang K (2005) Molecular cloning, characterization and expression of a novel Asr gene from Ginkgo biloba. Plant Physiol Biochem 43:836–843

    Article  CAS  PubMed  Google Scholar 

  15. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(–Delta Delta C(T)) method). Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  16. Liu Y, Teng Z, Wang S, He G (2008) Effects of high temperature stress on soluble sugar and membrane protective enzyme of rice. J Southwest Uni 30:59–63

    Google Scholar 

  17. Zhang DZ, Wang PH, Zhao HX (1990) Determination of the content of free proline in wheat leaves. Plant Physiol Commun 94:462–465

    Google Scholar 

  18. Rogerio M, Christophe D, Felipe KT, Marcia MP (2008) Glutathione peroxidase family—an evolutionary overview. FEBS J 275:3959–3970

    Article  Google Scholar 

  19. Lee PY, Kho CW, Lee H, Kang S, Lee SC, Park BC, Cho S, Bae KH, Park SG (2007) Glutathione peroxidase 3 of Saccharomyces cerevisiae suppresses non-enzymatic proteolysis of glutamine synthetase in an activity-independent manner. Biochem Biophys Res Comm 362:405–409

    Article  CAS  PubMed  Google Scholar 

  20. Utomo A, Jiang XZ, Furuta S, Yun J, Levin DS, Wang YCJ, Desai KV, Green JE, Chen PL, Lee WH (2004) Identification of a novel putative non-selenocysteine containing phospholipid hydroperoxide glutathione peroxidase (NPGPx) essential for alleviating oxidative stress generated from polyunsaturated fatty acids in breast cancer cells. J Biol Chem 279:43522–43529

    Article  CAS  PubMed  Google Scholar 

  21. Criqui MC, Jamet E, Parmentier Y, Marbach J, Durr A, Fleck J (1992) Isolation and characterization of a plant cDNA showing homology to animal glutathione peroxidases. Plant Mol Biol 18:623–627

    Article  CAS  PubMed  Google Scholar 

  22. Holland D, Ben-Hayyim G, Faltin Z, Camoin L, Strosberg AD, Eshdat Y (1993) Molecular characterization of salt-stress-associated protein in Citrus: protein and cDNA sequence homology to mammalian glutathione peroxidase. Plant Mol Biol 21:923–927

    Article  CAS  PubMed  Google Scholar 

  23. Sugimoto M, Sakamoto W (1997) Putative phospholipid hydroperoxide glutathione peroxidase gene from Arabidopsis thaliana induced by oxidative stress. Genes Genet Syst 72:306–311

    Article  Google Scholar 

  24. Li WJ, Feng H, Fan JH, Zhang RQ, Zhao NM, Liu JY (2000) Molecular cloning and expression of a phospholipid hydroperoxide glutathione peroxidase homolog in Oryza sativa. Biochim Biophys Acta Gene Struct Expr 1493:225–230

    Article  CAS  Google Scholar 

  25. Jung BG, Lee KO, Lee SS, Chi YH, Jang HH, Kang SS, Lee K, Lim D, Yoon SC, Yun DJ, Inoue Y, Cho MJ, Lee SY (2002) A Chinese cabbage cDNA with high sequence identity to phospholipid hydroperoxide glutathione peroxidases encodes a novel isoform of thioredoxin-dependent peroxidase. J Biol Chem 277:12572–12578

    Article  CAS  PubMed  Google Scholar 

  26. Nicolas N, Vale´rie C, Jose´ G, Eric G, Masakazu H, Pascal R, David BK, Emmanuelle Issakidis, Jean-Pierre Jacquot, Nicolas Rouhier (2006) Plant glutathione peroxidases are functional peroxiredoxins distributed in several subcellular compartments and regulated during biotic and abiotic stresses. Plant Physiol 142:1364–1379

    Article  Google Scholar 

  27. Rocher C, Lalanne JL, Chaudiere J (1992) Purification and properties of a recombinant sulfur analog of murine selenium-glutathione peroxidase. Eur J Biochem 205:955–960

    Article  CAS  PubMed  Google Scholar 

  28. Roxas VP, Lodhi SA, Garrett DK, Mahan JR, Allen RD (2000) Stress tolerance in transgenic tobacco seedlings that overexpress glutathione S-transferase/glutathione peroxidase. Plant Cell Physiol 41:1229–1234

    Article  CAS  PubMed  Google Scholar 

  29. Milla M, Maurer A, Huete A, Gustafson J (2003) Glutathione peroxidase genes in Arabidopsis are ubiquitous and regulated by abiotic stresses through diverse signaling pathways. Plant J 36:602–615

    Article  Google Scholar 

  30. Miao Y, Lv D, Wang P, Wang X, Chen J, Miao C, Song C (2006) An Arabidopsis glutathione peroxidase functions as both a redox transducer and a scavenger in abscisic acid and drought stress responses. Plant Cell 18:2749–2766

    Article  PubMed Central  CAS  PubMed  Google Scholar 

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Acknowledgments

This research was supported by National Key Technologies R&D Program (No. 2012BAD27B01). The authors thank the anonymous referees for their critical comments that contributed toward improving this manuscript.

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Authors and Affiliations

  1. State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, Wuhan, 430072, People’s Republic of China

    Ying Diao, Huaxue Xu, Guolin Li, Aiqing Yu, Xia Yu, Xingfei Zheng, Shaoqing Li & Zhongli Hu

  2. School of Pharmaceutical Sciences, Wuhan University, Wuhan, 430072, People’s Republic of China

    Ying Diao & Youwei Wang

  3. Institute of WTO, Wuhan University, Wuhan, 430072, People’s Republic of China

    Wanling Hu

Authors
  1. Ying Diao
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  2. Huaxue Xu
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  3. Guolin Li
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  5. Xia Yu
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  6. Wanling Hu
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  10. Zhongli Hu
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Corresponding authors

Correspondence to Shaoqing Li or Zhongli Hu.

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Ying Diao and Huaxue Xu contributed equally to this work.

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Diao, Y., Xu, H., Li, G. et al. Cloning a glutathione peroxidase gene from Nelumbo nucifera and enhanced salt tolerance by overexpressing in rice. Mol Biol Rep 41, 4919–4927 (2014). https://doi.org/10.1007/s11033-014-3358-4

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  • DOI: https://doi.org/10.1007/s11033-014-3358-4

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