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Cloning and expression of isoenzymes of superoxide dismutase in Haematococcus pluvialis (Chlorophyceae) under oxidative stress

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Abstract

Increasing evidence indicates that the biosynthesis and accumulation of the keto-carotenoid astaxanthin represents a chronic (in days or weeks) molecular defense mechanism in the green alga, Haematococcus pluvialis, to protect cells from abiotic stress. However, information with regard to the acute (in minutes or hours) response of the cells to stress is scarce. In this study, two cDNAs encoding manganese superoxide dismutase (MnSOD) were cloned and their full length nucleotide sequences were obtained. Quantitative real-time RT-PCR analysis revealed a general increase in MnSOD transcriptions in response to various abiotic stressors, e.g., high light, excessive amounts of iron or sodium acetate, either singly or in combination. Meanwhile, four isoforms of MnSOD were detected by a nondenaturing polyacrylamide gel electrophoresis approach. In addition, an iron-containing superoxide dismutase (FeSOD) was detected in the cells using the same technique. Differential expressions of SOD isoenzymes under different abiotic stressors were also investigated and discussed, at both mRNA and protein levels.

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References

  • Alscher RG, Erturk N, Lenwood SH (2002) Role of superoxide dismutases (SODs) in controlling oxidative stress in plants. J Exp Bot 53:1331–1341

    Article  CAS  PubMed  Google Scholar 

  • Armbrust E, Berges J, Bowler C, Green B, Martinez D, Putnam H, Zhou S, Allen E, Apt E, Bechner M, Brzezinski M, Chaal B, Chiovitti A, Davis A, Demarest M, Detter J, Glavina T, Goodstein D, Hadi M, Hellsten U, Hildebrand M, Jenkins B, Jurka J, Kapitonov V, Kröger N, Lau W, Lane T, Larimer F, Lippmeier J, Lucas S, Medina M, Montsant A, Obornik M, Parker M, Palenik B, Pazour G, Richardson P, Rynearson T, Saito M, Schwartz D, Thamatrakoln K, Valentin K, Vardi A, Wilkerson F, Rokhsa D (2004) The genome of the diatom Thalassiosira pseudonana: ecology, evolution, and metabolism. Science 306:79–86

    Article  CAS  PubMed  Google Scholar 

  • Asada K (1994) Production and action of active oxygen species in photosynthetic tissues. In: Foyer C, Mullineaux P (eds) Photooxidative Stresses in Plants: Causes and Amelioration. CRC Press, Boca Raton, pp 77–104

    Google Scholar 

  • Asada K, Yoshikawa K, Takahashi M, Maeda Y, Enmanji K (1975) Superoxide dismutases from a blue-green alga, Plectonema boryanum. J Biol Chem 250:2801–7

    CAS  PubMed  Google Scholar 

  • Asada K, Kanematsu S, Uchida K (1977) Superoxide dismutases in photosynthetic organisms: absence of the cuprozinc enzyme in eukaryotic algae. Arch Biochem Biophys 179:243–256

    Article  CAS  PubMed  Google Scholar 

  • Beauchamp C, Fridovich I (1971) Superoxide dismutase: Improved assays and an assay applicable to acrylamide gels. Anal Biochem 44:276–287

    Article  CAS  PubMed  Google Scholar 

  • Blokhina O, Virolainen E, Fagerstedt KV (2003) Antioxidants, oxidative damage and oxygen deprivation stress: a review. Ann Bot 91:179–94

    Article  CAS  PubMed  Google Scholar 

  • Boussiba S (2000) Carotenogenesis in the green alga Haematococcus pluvialis: cellular physiology and stress response. Physiol Plant 108:111–117

    Article  CAS  Google Scholar 

  • Boussiba S, Wang B, Yuan JP, Zarka A, Chen F (1999) Changes in pigments profile in the green alga Haematococcus pluvialis exposed to environmental stresses. Biotechnol Lett 21:601–604

    Article  CAS  Google Scholar 

  • Bowler C, Camp W, Vanmontagu M, Inze D (1994) Superoxide-dismutase in plants. CRC Crit Rev Plant Sci 13:199–218

    CAS  Google Scholar 

  • Buchanan BB, Luan S (2005) Redox regulation in the chloroplast thylakoid lumen: a new frontier in photosynthesis research. J Exp Bot 56:1439–1447

    Article  CAS  PubMed  Google Scholar 

  • Church S (1990) Manganese superoxide dismutase: nucleotide and deduced amino acid sequence of a cDNA encoding a new human transcript. Biochim Biophys Acta 1087:250–252

    CAS  PubMed  Google Scholar 

  • Elstner EF, Heupel A (1976) Inhibition of nitrite formation from hydroxylammonium chloride: a simple assay for superoxide dismutase. Anal Biochem 70:616–620

    Article  CAS  PubMed  Google Scholar 

  • Erturk HN (1999) Responses of superoxide dismutases to oxidative stress in Arabidopsis thaliana. In: Biology. Virginia Polytechnic Institute and State University, Blacksburg, VA, p 179

    Google Scholar 

  • Fridovich I (1975) Superoxide dismutases. Annu Rev Biochem 44:147–159

    Article  CAS  PubMed  Google Scholar 

  • Fridovich I (1986) Biological effects of the superoxide radical. Arch Biochem Biophys 247:1–11

    Article  CAS  PubMed  Google Scholar 

  • Gabriel O (1971) Locating enzymes on gels. In: Colowick SP, Kaplan NO (eds) Methods in Enzymology, vol 22. Academic, New York, p 578

    Google Scholar 

  • Geigenberger P, Kolbe A, Tiessn A (2005) Redox regulation of carbon storage and partitioning in response to light and sugars. J Exp Bot 56:1469–1479

    Article  CAS  PubMed  Google Scholar 

  • Grun S, Lindermayr C, Sell S (2006) Nitric oxide and gene regulation in plants. J Exp Bot 57:507–516

    Article  CAS  PubMed  Google Scholar 

  • Hagen C, Braune W, Vogel K, Hader DP (1993) Functional aspects of secondary carotenoids in Haematococcus lacustris (Girod) Rostafinski (Volvocales) V. Influences on photomovement. Plant Cell Environ 16:991–995

    Article  CAS  Google Scholar 

  • Ho Y, Howard A, Crapo J (1991) Molecular structure of a functional rat gene for manganese-containing superoxide dismutase. Am J Respir Cell Mol Biol 4:278–286

    CAS  PubMed  Google Scholar 

  • Kaminaka H, Morita S, Tokumoto M, Yokoyama H, Masumura T, Tanaka K (1999) Molecular cloning and characterization of a cDNA for an iron-superoxide dismutase in rice (Oryza sativa L.). Biosci Biotech Biochem 63:302–308

    Article  CAS  Google Scholar 

  • Kanematsu S, Asada K (1990) Characteristic amino acid sequences of chloroplast and cytosol isozymes of CuZn-superoxide dismutase in spinach, rice and horsetail. Plant Cell Physiol 31:99–112

    CAS  Google Scholar 

  • Karpinski S, Escobar C, Karpinska B, Creissen G, Mullineaux PM (1997) Photosynthetic electron transport regulates the expression of cytosolic ascorbate peroxidase genes in Arabidopsis during excess light stress. Plant Cell 9:627–640

    Article  CAS  PubMed  Google Scholar 

  • Kitayama K (1994) Molecular analysis of photooxidative stress responses in Chlamydomonas reinhardtii. Thesis, Indiana University, Bloomington

  • Kliebenstein DJ, Monde RA, Last RL (1998) Superoxide dismutase in Arabidopsis: an eclectic enzyme family with disparate regulation and protein localization. Plant Physiol 118:637–650

    Article  CAS  PubMed  Google Scholar 

  • Kobayashi M, Kakizono T, Nagai S (1991) Astaxanthin production by a green-alga, Haematococcus pluvialis accompanied with morphological-changes in acetate media. J Ferment Bioeng 71:335–339

    Article  CAS  Google Scholar 

  • Kobayashi M, Kakizono T, Nishio N, Nagai S, Kurimura Y, Tsuji Y (1997) Antioxidant role of astaxanthin in the green alga Haematococcus pluvialis. Appl Microbiol Biotechnol 48:351–356

    Article  CAS  Google Scholar 

  • Kwon SJ, Choi EY, Choi YJ (2006) Proteomics studies of post-translational modifications in plants. J Exp Bot 57:1547–1551

    Article  CAS  PubMed  Google Scholar 

  • Li YT, Sommerfeld M, Chen F, Hu Q (2008) Consumption of oxygen by astaxanthin biosynthesis: a protective mechanism against oxidative stress in Haematococcus pluvialis (Chlorophyceae). J Plant Physiol 165:1783–1797

    Article  CAS  PubMed  Google Scholar 

  • Lu F, Vonshak A, Zarka A, Boussiba S (1998) Does astaxanthin protect Haematococcus against light damage? Z Naturforsch 53c:93–100

    Google Scholar 

  • Matsuzaki M, Misumi O, Shin-I T, Maruyama S, Takahara M, Miyagishima SY, Mori T, Nishida K, Yagisawa F, Nishida K, Yoshida Y, Nishimura Y, Nakao S, Kobayashi T, Momoyama Y, Higashiyama T, Minoda A, Sano M, Nomoto H, Oishi K, Hayashi H, Ohta F, Nishizaka S, Haga S, Miura S, Morishita T, Kabeya Y, Terasawa K, Suzuki Y, Ishii Y, Asakawa S, Takano H, Ohta N, Kuroiwa H, Tanaka K, Shimizu N, Sugano S, Sato N, Nozaki H, Ogasawara N, Kohara Y, Kuroiwa T (2004) Genome sequence of the ultrasmall unicellular red alga Cyanidioschyzon merolae 10D. Nature 428:653–657

    Article  CAS  PubMed  Google Scholar 

  • McCord JM, Fridovich I (1969) Superoxide dismutase. An enzymic function for erythrocuprein (hemocuprein). J Biol Chem 244:6049–6055

    CAS  PubMed  Google Scholar 

  • Mur LAJ, Tim L, Carver W (2006) No way to live: The various roles of nitric oxide in plant-pathogen interactions. J Exp Bot 57:489–505

    Article  CAS  PubMed  Google Scholar 

  • Parker M, Blake C, Barra D, Bossa F, Schinina M, Bannister W, Bannister J (1987) Structural identity between the iron- and manganese-containing superoxide dismutases. Protein Eng 1:393–397

    Article  CAS  PubMed  Google Scholar 

  • Perl-Treves R, Perl A (2002) Oxidative stress: an introduction. In: Van Montagu M, Inzé D (eds) Oxidative stress in plants. Taylor & Francis, London, pp 1–32

    Google Scholar 

  • Sakamoto A, Nosaka Y, Tanaka K (1993) Cloning and sequencing analysis of a complementary DNA for manganese-superoxide dismutase from rice (Oryza sativa L.). Plant Physiol 103:1477–1478

    Article  CAS  PubMed  Google Scholar 

  • Sakurai H, Kusumoto N, Kitayama K, Robert TK (1993) Isozymes of superoxide dismutase in Chlamydomonas and purification of one of the major isozymes containing Fe. Plant Cell Physiol 34:1133–1137

    CAS  Google Scholar 

  • Shaish A, Avron M, Pick U, Ben-Amotz A (1993) Are active oxygen species involved in induction of β-carotene in Dunaliella bardawil? Planta 190:363–8

    Article  CAS  Google Scholar 

  • Steinbrenner J, Linden H (2001) Regulation of two carotenoid biosynthesis genes coding for phytoene synthase and carotenoid hydroxylase during stress-induced astaxanthin formation in the green alga Haematococcus pluvialis. Plant Physiol 125:810–817

    Article  CAS  PubMed  Google Scholar 

  • Steinbrenner J, Linden H (2003) Light induction of carotenoid biosynthesis genes in the green alga Haematococcus pluvialis: regulation by photosynthetic redox control. Plant Mol Biol 52:343–56

    Article  CAS  PubMed  Google Scholar 

  • Tsang EW, Bowler C, Hérouart D, Van Camp W, Villarroel R, Genetello C, Van Montagu M, Inzé D (1991) Differential regulation of superoxide dismutases in plants exposed to environmental stress. Plant Cell 3:783–92

    Article  CAS  PubMed  Google Scholar 

  • Ukeda H, Sarker AK, Kawana D, Sawamura M (1999) Flow-injection assay of superoxide dismutase based on the reduction of highly water-soluble tetrazolium. Anal Sci 15:353–357

    Article  CAS  Google Scholar 

  • Wang JX, Shi ZX, Xu XD (2004a) Residual plastids of bleached mutants of Euglena gracilis and their effects on the expression of nucleus-encoded genes. Prog Nat Sci 14:213–217

    Article  CAS  Google Scholar 

  • Wang JX, Sommerfeld M, Hu Q (2009) Occurrence and environmental stress responses of two plastid terminal oxidases in Haematococcus pluvialis (Chlorophyceae). Planta 230:191–203

    Article  CAS  PubMed  Google Scholar 

  • Wang JX, Zhang XZ, Chen YS, Sommerfeld M, Hu Q (2008) Toxicity assessment of manufactured nanomaterials using the unicellular green alga Chlamydomonas reinhardtii. Chemosphere 73:1121–1128

    Article  CAS  PubMed  Google Scholar 

  • Wang S, Chen F, Sommerfeld M, Hu Q (2004b) Proteomic analysis of molecular response to oxidative stress by the green alga Haematococcus pluvialis (Chlorophyceae). Planta 220:17–29

    Article  CAS  PubMed  Google Scholar 

  • Wuerges J, Lee J, Yim Y, Yim H, Kang S, Carugo K (2004) Crystal structure of nickel-containing superoxide dismutase reveals another type of active site. Proc Natl Acad Sci USA 101:8569–8574

    Article  CAS  PubMed  Google Scholar 

  • Youn HD, Kim E, Roe J, Hah Y, Kang S (1996) A novel nickel-containing superoxide dismutase from Streptomyces spp. Biochem J 318:889–896

    CAS  PubMed  Google Scholar 

  • Zhekisheva M, Boussiba S, Khozin-Goldberg I, Zarka A, Cohen Z (2002) Accumulation of oleic acid in Haematococcus pluvialis (Chlorophyceae) under nitrogen starvation or high light is correlated with that of astaxanthin esters. J Phycol 38:325–331

    Article  CAS  Google Scholar 

  • Zhu D, Scandalios J (1993) Maize mitochondrial manganese superoxide dismutases are encoded by a differentially expressed multigene family. Proc Natl Acad Sci USA 90:9310–9314

    Article  CAS  PubMed  Google Scholar 

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Correspondence to Qiang Hu.

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Wang, J., Sommerfeld, M. & Hu, Q. Cloning and expression of isoenzymes of superoxide dismutase in Haematococcus pluvialis (Chlorophyceae) under oxidative stress. J Appl Phycol 23, 995–1003 (2011). https://doi.org/10.1007/s10811-010-9631-6

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  • DOI: https://doi.org/10.1007/s10811-010-9631-6

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