Abstract
Superoxide dismutase (SOD) is an important antioxidant protein that protects organisms against various oxidative stresses by eliminating hydrogen peroxide. In the present study, a full-length cDNA sequence of manganese superoxide dismutase (MnSOD) from the Ulva prolifera (denoted as UpMnSOD) was cloned by a reverse-transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end method. The UpMnSOD cDNA sequence contained 955 bp with an open reading frame of 699 bp encoding 232 amino acid residues. The cDNA contained a 5′-untranslated region (UTR) of 29 bp nucleotides and a long 3′-UTR of 227 bp nucleotides. The calculated molecular mass was 25.7 kDa, and the estimated isoelectronic point (pI) of this protein was 6.83. Histidine (His) 59, His103, His194, and aspartate (Asp) 190 were found to be activated sites in UpMnSOD. Multiple alignment analysis revealed that the deduced amino acid sequence of MnSOD shared high identity (86 %) with Ulva fasciata. A phylogenetic tree construct indicated that UpMnSOD clustered into one subgroup with the MnSOD from U. fasciata. Quantitative real-time analysis revealed that UpMnSOD expression was induced at both 35 and 5 °C. The result indicated that UpMnSOD expression levels reached a maximum point of 2.63-fold (P < 0.05) and 1.28-fold compared to the control at 25 °C after 6-h treatment at 35 and 5 °C, respectively, and that long-time heat shock and cold treatment decreased the mRNA expression levels of MnSOD. Furthermore, UpMnSOD expression increased after 15 °C treatment, a 2.4-fold increase after 12-h treatment. Treatment with 1 mmol·L−1 salicylic acid (SA) at 35 °C also increased the gene expression of UpMnSOD and reached a maximum of 1.48-fold of the control after 6-h treatment, followed by a gradual decrease. However, the gene expression level of UpMnSOD increased rapidly within 3 h then decreased quickly under SA 5 mmol·L−1 + 35 °C. These data indicate that MnSOD was perhaps involved in the acute response against temperature stress, and salicylic acid could alleviate the high temperature stress effect on U. prolifera.
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Abele D, Tesch C, Wencke P, Portner HO (2001) How do oxidative stress relate to thermal tolerance in the Antarctic bivalve Yoldia eightsi. Antarct Sci 13:111–118
Ansaldo M, Polo J, Evelson P, Luqet C, Llesuy S (2000) Antioxidant levels from different antarctic fish caught around South Georgia island and Shag Rocks. Polar Biol 23:160–165
Arora A, Sairam RK, Srivastava GC (2002) Oxidative stress and antioxidative system in plants. Curr Sci India 82:1227–1238
Boo SY, Wong CM, Rodrigues KF, Najimudin N, Murad AMA, Mahadi NM (2013) Thermal stress responses in antarctic yeast, Glaciozyma antarctica PI12, characterized by real-time quantitative PCR. Polar Biol 36:381–389
Borsani O, Valpuesta V, Botella MA (2001) Evidence for a role of salicylic acid in the oxidative damage generated by NaCl and osmotic stress in Arabidopsis seedlings. Plant Physiol 126:1024–1030
Chiang CM, Kuo WS, Lin KH (2014) Cloning and gene expression analysis of sponge gourd ascorbate peroxidase gene and winter squash superoxide dismutase gene under respective flooding and chilling stresses. Hort Environ Biotechnol 55:129–137
Collén J, Davison IR (2001) Seasonality and thermal acclimation of reactive oxygen metabolism in Fucus vesiculosus (Phaeophyceae). J Phycol 37:474–481
Boussiba S (2000) Carotenogenesis in the green alga Haematococcus pluvialis: cellular physiology and stress response. J Plant Physiol 108:111–117
Fan MH, Sun X, Wang RX (2014) Gene cloning and expression analysis of antioxidant gene CAT from Ulva prolifera. J Fish China 38:1976–1983 (Chinese)
Fan WS, Huang HZ, Xu HF (2011) Effects of additive exogenous salicylic acid on cold resistance of Gracilaria. Mar Sci 35:38–43 (Chinese)
Fan YM, Zhang L, Qin S (2012) Haematococcus pluvialis iron superoxide gene cloning and sequence analysis. Mar Sci 36:79–85 (Chinese)
Hayat Q, Hayat S, Irfan M (2010) Effect of exogenous salicylic acid under changing environment: a review. Environ Exp Bot 68:14–25
Heise K, Puntarulo S, Nikinmaa M, Lucassen M, Portner HO, Abele D (2006) Oxidative stress and HIF-1 DNA binding during stressful cold exposure and recovery in the North Sea eelpout (Zoarces viviparous). Comp Biochem Physiol 143:494–503
Jia SG, Wang XM, Liu GM (2011) Gene expression analysis of“green tide”alga Ulva prolifera (Chlorophyta) in China. Genes Genomics 33:173–178
Kwon SJ, Choi EY, Choi YJ (2006) Proteomics studies of posttranslational modifications in plants. J Exp Bot 57:1547–1551
Lewis A, Blokhina O, Virolainen E (2003) Antioxidants, oxidative damage and oxygen deprivation stress: a review. Ann Bot 91:179–194
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
Li YX, Zhang XW, Dong X (2012) Differential gene expression in Ulva prolifera under low light and low temperature conditions. Curr Genet 58:235–244
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25:402–408
Mohankumar K, Ramasamy P (2006) White spot syndrome virus infection decreases the activity of antioxidant enzymes in Fenneropenaeus indicus. Virus Res 115:69–75
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
Nemeth M, Janda T, Horvath E, Paldi E, Szalai G (2002) Exogenous salicylic acid increases polyamine content but may decrease drought tolerance in maize. Plant Sci 162:569–574
Priya B, Premanandh J, Dhanalakshmi TR, Uma L, Prabaharan D, Subramanian G (2007) Comparative analysis of cyanobacterial superoxide dismutases to discriminate canonical forms. BMC Genomics 8:435–444
Provasoli L (1968) Media and prospect for the cultivation of marine algae. In: Watanabe A, Hattori A (eds) Cultures and collection of algae. Proc USA Japan Conf, Hakone, Sept. 1966. Jap Soc Plant Physiol, pp 63–75
Rahat N, Noushina I, Shabina S (2011) Salicylic acid alleviates decreases in photosynthesis under salt stress by enhancing nitrogen and sulfur assimilation and antioxidant metabolism differentially in two mungbean cultivars. J Plant Physiol 168:807–815
Salveit ME (2000) Chilling injury is reduced in cucumber and rice seedlings and in tomato pericarp discs by heat-shocks applied after chilling. Posthar Biol Technol 21:169–177
Seshadri G, Che PL, Boopathy AV, Davis ME (2012) Characterization of superoxide dismutases in cardiac progenitor cells demonstrates a critical role for manganese superoxide dismutase. Stem Cells Dev 21:3136–3146
Sung MS, Hsu YT, Wu TM, Lee TM (2009) Hyper salinity and hydrogen peroxide up-regulation of gene expression of antioxidant enzymes in Ulva fasciata against oxidative stress. Mar Biotechnol 11:199–209
Wahid AS, Gelani M, Ashraf MRF (2007) Heat tolerance in plants: an overview. Environ Exp Bot 61:199–223
Wang JX, Shi ZX, Xu XD (2004) Residual plastids of bleached mutants of Euglena gracilis and their effects on the expression of nucleus-encoded genes. Prog Nat Sci 14:213–217
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
Wang JX, Milton S, Hu Q (2011) Cloning and expression of isoenzymes of superoxide dismutase in Haematococcus pluvialis (Chlorophyceae) under oxidative stress. J Appl Phycol 23:995–1003
Wang Y, Wang Y, Zhu L (2012) Comparative studies on the ecophysiological differences of two green tide macroalgae under controlled laboratory conditions. PLoS One 7(8):e38245
Wang F, Wu QH, Zhang ZD, Chen SF, Zhou RC (2013) Cloning, expression, and characterization of iron superoxide dismutase in Sonneratia alba, a highly salt tolerant mangrove tree. Protein J 32:259–265
Wu TM, Hsu YT, Sung MS (2009) Expression of genes involved in redox homeostasis and antioxidant defense in a marine macroalgae Ulva fasciata by excess copper. Aquat Toxicol 94:275–285
Yang JH, Dong SM, Zhu HX, Jiang QC, Yang JX (2013) Molecular and expression analysis of manganese superoxide dismutase (MnSOD) gene under temperature and starvation stress in rotifer Brachionus calyciflorus. Mol Biol Rep 40:2927–2937
Zhang BL, Yan XH, Huang LB (2011) Evaluation of an improved strain of Porphyra yezoensis Ueda (Bangiales, Rhodophyta) with high-temperature tolerance. J Appl Phycol 23:841–847
Zhu ZB, Sun X, Xu NJ (2012) Effects of salicylic acid on the resistance of Gracilariopsis lemaneiformis to high temperature. J Fish China 36:1304–1309 (Chinese)
Acknowledgments
This study was supported by the National Natural Science Foundation of China (No. 40876073 and No. 41276122) and Research Fund for the Doctoral Program of Higher Education of China (20123305110002). This research was also sponsored by K.C. Wong Magna Fund of Ningbo University, Zhejiang, China.
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Fan, M., Sun, X., Xu, N. et al. cDNA cloning, characterization and expression analysis of manganese superoxide dismutase in Ulva prolifera . J Appl Phycol 28, 1391–1401 (2016). https://doi.org/10.1007/s10811-015-0632-3
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DOI: https://doi.org/10.1007/s10811-015-0632-3