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Expression of mercuric reductase from Bacillus megaterium MB1 in eukaryotic microalga Chlorella sp. DT: an approach for mercury phytoremediation

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Abstract

A eukaryotic microalga, Chlorella sp. DT, was transformed with the Bacillus megaterium strain MB1 merA gene, encoding mercuric reductase (MerA), which mediates the reduction of Hg2+ to volatile elemental Hg0. The transformed Chlorella cells were selected first by hygromycin B and then by HgCl2. The existence of merA gene in the genomic DNA of transgenic strains was shown by polymerase chain reaction amplification, while the stable integration of merA into genomic DNA of transgenic strains was confirmed by Southern blot analysis. The ability to remove Hg2+ in merA transgenic strains was higher than that in the wild type. The merA transgenic strains showed higher growth rate and photosynthetic activity than the wild type did in the presence of a toxic concentration of Hg2+. Cultured with Hg2+, the expression level of superoxide dismutase in transgenic strains was lower than that in the wild type, suggesting that the transgenic strains faced a lower level of oxidative stress. All the results indicated that merA gene was successfully integrated into the genome of transgenic strains and functionally expressed to promote the removal of Hg2+.

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References

  • Arnon DI (1949) Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. Plant Physiol 24:1–15

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Barkay T, Miller SM, Summers AO (2003) Bacterial mercury resistance from atoms to ecosystems. FEMS Microbiol Rev 27:355–384

    Article  CAS  PubMed  Google Scholar 

  • Baroli I, Gutman BL, Ledford HK, Shin JW, Chin BL, Havaux M, Niyogi KK (2004) Photo-oxidative stress in a xanthophyll-deficient mutant of Chlamydomonas. J Biol Chem 279:6337–6344

    Article  CAS  PubMed  Google Scholar 

  • Bizily SP, Rugh CL, Meagher RB (2000) Phytodetoxification of hazardous organomercurials by genetically engineered plants. Nat Biotechnol 18:213–217

    Article  CAS  PubMed  Google Scholar 

  • Chen PC, Lorezen H (1986) Changes in the productivity and nuclear divisions in synchronous Chlorella and circadian rhythm. Plant Cell Physiol 27:1423–1427

    CAS  Google Scholar 

  • Chen PC, Lai CL (1996) Physiological adaptation during cell dehydration and rewetting of a newly-isolated Chlorella species. Physiol Plant 96:453–457

    Article  CAS  Google Scholar 

  • Chen Y, Wang Y, Sun Y, Zhang L, Li W (2001) Highly efficient expression of rabbit neutrophil peptide-1 gene in Chlorella ellipsodea cells. Curr Genet 39:365–370

    Article  CAS  PubMed  Google Scholar 

  • Chow KC, Tung WL (1999) Electroporation of Chlorella vulgaris. Plant Cell Rep 18:778–780

    Article  CAS  Google Scholar 

  • Clare DA, Rabinowitch HD, Fridovich I (1984) Superoxide dismutase and chilling injury in Chlorella ellipsoidea. Arch Biochem Biophys 231:158–163

    Article  CAS  PubMed  Google Scholar 

  • Dawson HN, Burlingame R, Cannons AC (1997) Stable transformation of Chlorella: rescue of nitrate reductase-deficient mutants with the nitrate reductase gene. Curr Microbiol 35:356–362

    Article  CAS  PubMed  Google Scholar 

  • Devars S, Aviles C, Cervantes C, Moreno-Sanchez R (2000) Mercury uptake and removal by Euglena gracilis. Arch Microbiol 174:175–180

    Article  CAS  PubMed  Google Scholar 

  • Harada M (1995) Minamata disease: methylmercury poisoning in Japan caused by environmental pollution. Crit Rev Toxicol 25:1–24

    Article  CAS  PubMed  Google Scholar 

  • Hassett-Sipple B, Swartout J, Schoeny R, Mahaffey KR, Rice GE (1997) Mercury study report to congress, vol 5. Health effects of mercury and mercury compounds. US Environmental Protection Agency, Washington, DC, pp 1–349

    Google Scholar 

  • Hawkins RL, Nakamura M (1999) Expression of human growth hormone by eukaryotic alga, Chlorella. Curr Microbiol 38:335–341

    Article  CAS  PubMed  Google Scholar 

  • He YK, Sun JG, Feng XZ, Czako M, Marton L (2001) Differential mercury volatilization by tobacco organs expressing a modified bacterial merA gene. Cell Res 11:231–236

    Article  CAS  PubMed  Google Scholar 

  • Heaton AC, Rugh CL, Kim T, Wang NJ, Meagher RB (2003) Toward detoxifying mercury-polluted aquatic sediments with rice genetically engineered for mercury resistance. Environ Toxicol Chem 22:2940–2947

    Article  CAS  PubMed  Google Scholar 

  • Hoffman P, Werner D (1966) Spectrophotometric chlorophyll determination having special regard to various types of equipment. Jena Rev 7:300–303

    Google Scholar 

  • Huang CC, Narita M, Yamagata T, Itoh Y, Endo G (1999) Structure analysis of a class II transposon encoding the mercury resistance of the Gram-positive bacterium Bacillus megaterium MB1, a strain isolated from Minamata Bay, Japan. Gene 234:361–369

    Article  CAS  PubMed  Google Scholar 

  • Inthorn D, Sidtitoon N, Silapanuntakul S, Incharoensakdi A (2002) Sorption of mercury, cadmium and lead by microalgae. Sci Asia 28:253–261

    Article  CAS  Google Scholar 

  • Jarvis EE, Brown LM (1991) Transient expression of firefly luciferase in protoplasts of the green alga Chlorella ellipsoidea. Curr Genet 19:317–321

    Article  CAS  Google Scholar 

  • Juneau P, Dewez D, Matsui S, Kim SG, Popovic R (2001) Evaluation of different algal species sensitivity to mercury and metolachlor by PAM-fluorometry. Chomosphere 45:589–598

    Article  CAS  PubMed  Google Scholar 

  • Kim DH, Kim YT, Cho JJ, Bae JH, Hur SB, Hwang I, Choi TJ (2002) Stable integration and functional expression of flounder growth hormone gene in transformed microalga, Chlorella ellipsoidea. Mar Biotechnol 4:63–73

    Article  CAS  Google Scholar 

  • Krause GH, Weis E (1984) Chlorophyll fluorescence as a tool in plant physiology. II. Interpretation of fluorescence signals. Photosynth Res 5:139–157

    Article  CAS  PubMed  Google Scholar 

  • Lee DH, Lee CB (2000) Chilling stress-induced changes of antioxidant enzymes in the leaves of cucumber: in gel enzyme activity assays. Plant Sci 159:75–85

    Article  CAS  PubMed  Google Scholar 

  • Leon-Banares R, Gonzalex-Ballester D, Galvan A, Fernandez E (2004) Transgenic microalgae as green cell-factories. Trends Biotechnol 22:45–52

    Article  CAS  PubMed  Google Scholar 

  • Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    CAS  PubMed  Google Scholar 

  • McElroy D, Zhang W, Cao J, Wu R (1990) Isolation of an efficient actin promoter for use in rice transformation. Plant Cell 2:163–171

    CAS  PubMed  PubMed Central  Google Scholar 

  • Richier S, Merle PL, Furla P, Pigozzi D, Sola F, Allemand D (2003) Characterization of superoxide dismutases in anoxia- and hyperoxia-tolerant symbiotic cnidarians. Biochim Biophys Acta 1621:84–91

    Article  CAS  PubMed  Google Scholar 

  • Rugh CL, Wilde HD, Stack NM, Thompson DM, Summers AO, Meagher RB (1996) Mercuric ion reduction and resistance in transgenic Arabidopsis thaliana plants expressing a modified merA gene. Proc Natl Acad Sci U S A 93:3182–3187

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rugh CL, Senecoff JF, Meagher RB, Merkle SA (1998) Development of transgenic yellow poplar for mercury phytoremediation. Nat Biotechnol 16:925–928

    Article  CAS  PubMed  Google Scholar 

  • Schiering N, Wkasch W, Moore MJ, Distefano MD, Walsh CT, Pai EE (1991) Structure of the detoxification catalyst mercuric ion reductase from Bacillus sp. strain RC607. Nature 352:168–172

    Article  CAS  PubMed  Google Scholar 

  • Sigaud-Kutner TCS, Leitão MAS, Okamoto OK (2003) Heavy metal-induced oxidative stress in algae. J Phycol 39:1008–1018

    Article  Google Scholar 

  • Siripornadulsil S, Traina S, Verma DPS, Sayre RT (2002) Molecular mechanisms of proline-mediated tolerance to toxic heavy metals in transgenic microalgae. Plant Cell 14:2837–2847

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Twyman RM, Stoger E, Kohli A, Capell T, Christou P (2002) Selectable and screenable markers for rice transformation. In: Jackson JF, Linskens HF (eds) Molecular methods of plant analysis, vol 22. Testing for genetic manipulation in plants. Springer, Berlin Heidelberg New York, pp 1–14

    Google Scholar 

  • Wilde EW, Benemann JR (1993) Bioremoval of heavy metals by the use of microalgae. Biotechnol Adv 11:781–812

    Article  CAS  PubMed  Google Scholar 

  • Wilkinson SC, Goulding KH, Robinson PK (1990) Mercury removal by immobilized algae in batch culture systems. J Appl Phycol 2:223–230

    Article  Google Scholar 

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Acknowledgements

This work was partially supported by a grant (NSC93-2311-B-005-017) from National Science Council, Taiwan, to L.F.C. We thank Professor Hungchen E. Yen for the help in manuscript revision.

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Authors and Affiliations

  1. Department of Life Sciences, National Chung-Hsing University, Tai-Chung 40227, Taiwan

    Chieh-Chen Huang, Meng-Wei Chen, Ju-Liang Hsieh, Wen-Hao Lin, Pei-Chung Chen & Lee-Feng Chien

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  1. Chieh-Chen Huang
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  2. Meng-Wei Chen
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  3. Ju-Liang Hsieh
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  4. Wen-Hao Lin
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Correspondence to Lee-Feng Chien.

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Huang, CC., Chen, MW., Hsieh, JL. et al. Expression of mercuric reductase from Bacillus megaterium MB1 in eukaryotic microalga Chlorella sp. DT: an approach for mercury phytoremediation. Appl Microbiol Biotechnol 72, 197–205 (2006). https://doi.org/10.1007/s00253-005-0250-0

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  • DOI: https://doi.org/10.1007/s00253-005-0250-0

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