Advertisement

Perspective for the Use of Genetic Transformants in Order to Enhance the Synthesis of the Desired Metabolites: Engineering Chloroplasts of Microalgae for the Production of Bioactive Compounds

  • Udo Johanningmeier
  • Dirk Fischer
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 698)

Abstract

Eukaryotic microalgae have recently gained particular interest as bioreactors because they provide attractive alternatives to bacterial, yeast, plant and other cell-based systems currently in use. Over the last years there has been considerable progress in genetic engineering technologies for algae. Biotechnology companies start to apply these techniques to alter metabolic pathways and express valuable compounds in different cell compartments. In particular, the eukaryotic unicellular alga Chlamydomonas reinhardtii appears to be a most promising cell factory since high amounts of foreign proteins have been expressed in its chloroplast compartment. For this alga the complete nuclear, plastidal and mitochondrial genome sequences have been determined and databases are available for any searching or cloning requirements. Apart from being easily transformable, stable transgenic strains and production volumes in full containment can be obtained within a relatively short time. Furthermore, C. reinhardtii is a green alga which belongs to the category of organisms generally recognized as safe (GRAS status). Thus, enhancing food with edible algae like Chlamydomonas engineered to (over)produce functional ingredients has the potential to become an important factor in food and feed technologies.

Keywords

Bioactive Peptide Foreign Protein Classical Swine Fever Virus Chloroplast Gene Euglena Gracilis 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Walker TL, Purton S, Becker DK et al. Microalgae as bioreactors. Plant Cell Rep 2005; 24:629–641.PubMedCrossRefGoogle Scholar
  2. 2.
    Debuchy R, Purton S, Rochaix JD. The argininosuccinate lyase gene of Chlamydomonas reinhardtii: an important tool for nuclear transformation and for correlating the genetic and molecular maps of the ARG7 locus. EMBO J 1989; 8:2803–2809.PubMedGoogle Scholar
  3. 3.
    Kindle KL, Schnell RA, Fernandez E et al. Stable nuclear transformation of Chlamydomonas using the Chlamydomonas gene for nitrate reductase. J Cell Biol 1989; 109:2589–2601.PubMedCrossRefGoogle Scholar
  4. 4.
    Ferris PJ. Localization of the nic-7, ac-29 and thi-10 genes within the mating-type locus of Chlamydomonas reinhardtii. Genetics 1995; 141:543–549.PubMedGoogle Scholar
  5. 5.
    Spolaore P, Joannis-Cassan C, Duran E et al. Commercial applications of microalgae. J Biosci Bioeng 2006; 101:87–96.PubMedCrossRefGoogle Scholar
  6. 6.
    Newell CA. Plant transformation technology: Developments and applications. Mol Biotechnol 2000; 16:53–65.PubMedCrossRefGoogle Scholar
  7. 7.
    Rochaix JD. Chlamydomonas reinhardtii as the photosynthetic yeast. Annu Rev Genet 1995; 29:209–230.PubMedCrossRefGoogle Scholar
  8. 8.
    Grossman AR, Harris EE, Hauser C et al. Chlamydomonas reinhardtii at the crossroads of genomics. Eukaryot Cell 2003; 2:1137–1150.PubMedCrossRefGoogle Scholar
  9. 9.
    Harris EH. Chlamydomonas as a Model Organism. Rev Plant Physiol Plant Mol Biol 2001; 52:363–406.CrossRefGoogle Scholar
  10. 10.
    Merchant SS, Prochnik SE, Vallon O et al. The Chlamydomonas genome reveals the evolution of key animal and plant functions. Science 2007; 318:245–250.PubMedCrossRefGoogle Scholar
  11. 11.
    Raven JA, Allen JF. Genomics and chloroplast evolution: what did cyanobacteria do for plants? Genome Biol 2003; 4:209–213.PubMedCrossRefGoogle Scholar
  12. 12.
    Mattoo A, Giardi MT, Raskind A et al. Dynamic metabolism of photosystem II reaction center proteins and pigments. A review. Physiol Plant 1999; 107:454–461.CrossRefGoogle Scholar
  13. 13.
    Maliga P. Plastid transformation in higher plants. Annu Rev Plant Biol 2004; 55:289–313.PubMedCrossRefGoogle Scholar
  14. 14.
    Daniell H et al. Containment of herbicide resistance through genetic engineering of the chloroplast genome. Nat Biotechnol 1998; 16:345–343.PubMedCrossRefGoogle Scholar
  15. 15.
    Lutz KA et al. Expression of bar in the plastid genome confers herbicide resistance. Plant Physiol 2001; 125:1585–1590.PubMedCrossRefGoogle Scholar
  16. 16.
    DeGray G et al. Expression of an antimicrobial peptide via the chloroplast genome to control phytopathogenic bacteria and fungi. Plant Physiol 2001; 127:852–862.PubMedCrossRefGoogle Scholar
  17. 17.
    Kota M et al. Overexpression of the Bacillus thuringiensis (Bt) Cry2Aa2 protein in chloroplasts confers resistance to plants against susceptible and Bt-resistant insects. Proc Natl Acad Sci USA 1999; 96:1840–1845.PubMedCrossRefGoogle Scholar
  18. 18.
    McBride KE et al. Amplification of a chimeric Bacillus gene in chloroplasts leads to an extraordinary level of an insecticidal protein in tobacco. Bio/Technology 1995; 13:362–365.PubMedCrossRefGoogle Scholar
  19. 19.
    Daniell H, Khan MS, Allison L. Milestones in chloroplast genetic engineering: an environmentally friendly era in biotechnology. TRENDS in Plant Science 2002; 7:84–91.PubMedCrossRefGoogle Scholar
  20. 20.
    Staub JM et al. High-yield production of a human therapeutic protein in tobacco chloroplasts. Nat Biotechnol 2000; 18:333–338.PubMedCrossRefGoogle Scholar
  21. 21.
    De Cosa B, Moar W, Lee S-B et al. Overexpression of the Bt cry2Aa2 operon in chloroplasts leads to formation of insecticidal crystals. Nat Biotechnol 2001; 19:71–74.CrossRefGoogle Scholar
  22. 22.
    Nakashita H, Arai Y, Shikanai T et al. Introduction of bacterial metabolism into higher plants by polycistronic transgene expression. Biosci Biotechnol Biochem 2001; 65:1688–1691.PubMedCrossRefGoogle Scholar
  23. 23.
    Lössl A, Eibl C, Harloff HJ et al. Polyester synthesis in transplastomic tobacco (Nicotiana tabacum L.): significant contents of polyhydroxybutyrate are associated with growth reduction. Plant Cell Rep 2003; 21:891–899.PubMedGoogle Scholar
  24. 24.
    Arai Y, Shikanai T, Doi Y et al. Production of polyhydroxybutyrate by polycistronic expression of bacterial genes in tobacco plastid. Plant Cell Physiol 2004; 45:1176–1184.PubMedCrossRefGoogle Scholar
  25. 25.
    Wurbs D, Ruf S, Bock R. Contained metabolic engineering in tomatoes by expression of carotenoid biosynthesis genes from the plastid genome. Plant J 2007; 49:276–288.PubMedCrossRefGoogle Scholar
  26. 26.
    Sidorov VA, Kasten D, Pang S-Z et al. Stable chloroplast transformation in potato: use of green fluorescent protein as a plastid marker. Plant J 1999; 19:209–216.PubMedCrossRefGoogle Scholar
  27. 27.
    Ruf S, Hermann M, Berger IJ et al. Stable genetic transformation of tomato plastids and expression of a foreign protein in fruit. Nat Biotechnol 2001; 19:870–875.PubMedCrossRefGoogle Scholar
  28. 28.
    Dufourmantel N, Pelissier B, Garcon F et al. Generation of fertile transplastomic soybean. Plant Mol Biol 2004; 55:479–489.PubMedCrossRefGoogle Scholar
  29. 29.
    Kumar S, Dhingra A, Daniell H. Plastid-expressed betaine aldehyde dehydrogenase gene in carrot cultured cells, roots and leaves confers enhanced salt tolerance. Plant Physiol 2004; 136:2843–2854.PubMedCrossRefGoogle Scholar
  30. 30.
    Lelivelt CLC, McCabe MS, Newell CA et al. Stable plastid transformation in lettuce (Lactuca sativa L.). Plant Mol Biol 2005; 58:763–774.PubMedCrossRefGoogle Scholar
  31. 31.
    Kanamoto H, Yamashita A, Asao H et al. Efficient and stable transformation of Lactuca sativa L. cv. Cisco (lettuce) plastids. Transgenic Res 2006; 15:205–217.PubMedCrossRefGoogle Scholar
  32. 32.
    Nugent GD, Coyne S, Nguyen TT et al. Nuclear and plastid transformation of Brassica oleracea var. botrytis (cauliflower) using PEG-mediated uptake into protolasts. Plant Sci 2006; 170:135–142.CrossRefGoogle Scholar
  33. 33.
    Ellstrand NC. When transgenes wander, should we worry? Plant Physiol 2001; 125:1543–1545.PubMedCrossRefGoogle Scholar
  34. 34.
    Ellstrand NC. Current knowledge of gene flow in plants: implications for transgene flow. Philos Trans R Soc Lond B Biol Sci 2003; 358:1163–1170.PubMedCrossRefGoogle Scholar
  35. 35.
    Ruf S, Karcher D, Bock R. Determining the transgene containment level provided by chloroplast transformation. Proc Natl Acad Sci USA 2007; 104:6998–7002.PubMedCrossRefGoogle Scholar
  36. 36.
    Boynton JE, Gillham NW, Harris EH et al. Chloroplast transformation in Chlamydomonas with high velocity microprojectiles. Science 1988; 240:1534–1538.PubMedCrossRefGoogle Scholar
  37. 37.
    Klein TM, Wolf ED, Wu R et al. High-velocity microprojectiles for delivering nucleic acids into living cells. Nature 1987; 327:70–73.CrossRefGoogle Scholar
  38. 38.
    Przibilla E, Heiss S, Johanningmeier U et al. Site-specific mutagenesis of the D1 subunit of Photosystem II in wildtype Chlamydomonas. Plant Cell 1991; 3:169–174.PubMedCrossRefGoogle Scholar
  39. 39.
    Newman SM, Gillham NW, Harris EH et al. Targeted disruption of chloroplast genes in Chlamydomonas reinhardtii. Mol Gen Genet. 1991; 230:65–74.PubMedCrossRefGoogle Scholar
  40. 40.
    Goldschmidt-Clermont M. Transgenic expression of aminoglycoside adenine transferase in the chloroplast: a selectable marker of site-directed transformation of Chlamydomonas. Nucleic Acids Res 1991; 19:4083–4089.PubMedCrossRefGoogle Scholar
  41. 41.
    Bateman JM, Purton S. Tools for chloroplast transformation in Chlamydomonas: expression vectors and a new dominant selectable marker. Mol Gen Genet 2000; 263:404–410.PubMedCrossRefGoogle Scholar
  42. 42.
    Lapidot M, Raveh D, Sivan A et al. Stable chloroplast transformation of the unicellular red alga Porphyridium species. Plant Physiol 2002; 129:7–12.PubMedCrossRefGoogle Scholar
  43. 43.
    Doetsch NA, Favreau MR, Kuscuoglu N et al. Chloroplast transformation in Euglena gracilis: splicing of a group III twintron transcribed from a transgenic psbK operon. Curr Genet 2001; 39:49–60.PubMedCrossRefGoogle Scholar
  44. 44.
    Blowers AD, Bogorad L, Shark KB et al. Studies on Chlamydomonas chloroplast transformation: foreign DNA can be stably maintained in the chromosome. Plant Cell 1989; 1:123–132.PubMedCrossRefGoogle Scholar
  45. 45.
    Blowers AD, Ellmore GS, Klein U et al. Transcriptional analysis of endogenous and foreign genes in chloroplast transformants of Chlamydomonas. Plant Cell 1990; 2:1059–1070.PubMedCrossRefGoogle Scholar
  46. 46.
    Ishikura K, Takaoka Y, Kato K et al. Expression of a foreign gene in Chlamydomonas reinhardtii chloroplast. J Biosci Bioeng 1999; 87:307–314.PubMedCrossRefGoogle Scholar
  47. 47.
    Minko I, Holloway SP, Nikaido S et al. Renilla luciferase as a vital reporter for chloroplast gene expression in Chlamydomonas. Mol Gen Genet 1999; 262:421–425.PubMedCrossRefGoogle Scholar
  48. 48.
    Franklin S, Ngo B, Efuet E et al. Development of a GFP reporter gene for Chlamydomonas reinhardtii chloroplast. Plant J 2002; 30:733–744.PubMedCrossRefGoogle Scholar
  49. 49.
    Mayfield SP, Franklin SE, Lerner RA. Expression and assembly of fully active antibody in algae. Proc Natl Acad Sci USA 2003; 100:438–442.PubMedCrossRefGoogle Scholar
  50. 50.
    Mayfield SP, Schultz J. Development of a luciferase reporter gene, luxCt, for Chlamydomonas reinhardtii chloroplast. Plant J 2004; 37:449–458.PubMedCrossRefGoogle Scholar
  51. 51.
    Salvador ML, Suay L, Anthonisen IL et al. Changes in the 5′-untranslated region of the rbcL gene accelerate transcript degradation more than 50-fold in the chloroplast of Chlamydomonas reinhardtii. Curr Genet 2004; 45:176–182.PubMedCrossRefGoogle Scholar
  52. 52.
    Klein U, Salvador ML, Bogorad L. Activity of the Chlamydomonas chloroplast rbcL gene promoter is enhanced by a remote sequence element. Proc Natl Acad Sci USA 1994; 91:10819–10823.PubMedCrossRefGoogle Scholar
  53. 53.
    Suay L, Salvador ML, Abesha E et al. Specific roles of 5′ RNA secondary structures in stabilizing transcripts in chloroplasts. Nucleic Acids Res 2005; 33:4754–4761.PubMedCrossRefGoogle Scholar
  54. 54.
    Klein U, De Camp JD, Bogorad L. Two types of chloroplast gene promoters in Chlamydomonas reinhardtii. Proc Natl Acad Sci USA 1992; 89:3453–3457.PubMedCrossRefGoogle Scholar
  55. 55.
    Eberhard S, Drapier D, Wollman FA. Searching limiting steps in the expression of chloroplast encoded proteins: relations between gene copy number, transcription, transcript abundance and translation rate in the chloroplast of Chlamydomonas reinhardtii. Plant J 2002; 31:149–160.PubMedCrossRefGoogle Scholar
  56. 56.
    Barnes D, Franklin S, Schultz J et al. Contribution of 5′-and 3′-untranslated regions of plastid mRNAs to the expression of Chlamydomonas reinhardtii chloroplast genes. Mol Genet Genomics 2005; 274:625–636.PubMedCrossRefGoogle Scholar
  57. 57.
    Mayfield SP, Manuell AL, Chen S et al. Chlamydomonas reinhardtii chloroplasts as protein factories. Curr Opin Biotechnol 2007; 18:126–133.PubMedCrossRefGoogle Scholar
  58. 58.
    Griesbeck C, Kobl I, Heitzer M. Chlamydomonas reinhardtii: a protein expression system for pharmaceutical and biotechnological proteins. Mol Biotechnol 2006; 34:213–223.PubMedCrossRefGoogle Scholar
  59. 59.
    Fukusaki EI, Nishikawa T, Kato K et al. Introduction of the Archaebacterial Geranylgeranyl Pyrophosphate Synthase Gene into Chlamydomonas reinhardtii chloroplast. J Biosci Bioeng 2003; 95:283–287.PubMedGoogle Scholar
  60. 60.
    Sun M, Qian K, Su N et al. Foot and mouth disease virus VP1 protein fused with cholera toxin B subunit expressed in Chlamydomonas reinhardtii chloroplast. Biotechnol Lett 2003; 25:1087–1092.PubMedCrossRefGoogle Scholar
  61. 61.
    Mayfield SP, Franklin SE. Expression of human antibodies in eukaryotic micro-algae. Vaccine 2005; 23:1828–1832.PubMedCrossRefGoogle Scholar
  62. 62.
    Su ZL, Qian KX, Tan CP et al. Recombination and heterologous expression of allophycocyanin gene in the chloroplast of Chlamydomonas reinhardtii. Acta Biochim Biophys Sin (Shanghai) 2005; 37:709–712.CrossRefGoogle Scholar
  63. 63.
    Zhang YK, Shen GF, Ru BG. Survival of human metallothioneine-2 transplastomic Chlamydomonas reinhardtii to ultraviolet B exposure. Acta Biochim Biophys Sin (Shanghai) 2006; 38:187–193.CrossRefGoogle Scholar
  64. 64.
    Matsuo T, Onai K, Okamoto K et al. Real-time monitoring of chloroplast gene expression by a luciferase reporter: evidence for nuclear regulation of chloroplast circadian period. Mol Cell Biol 2006; 26:863–870.PubMedCrossRefGoogle Scholar
  65. 65.
    Yang Z, Li Y, Chen F et al. Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast. Chin Sci Bull 2006; 51:1703–1709.CrossRefGoogle Scholar
  66. 66.
    Kato K, Marui T, Kasai S et al. Artificial control of transgene expression in Chlamydomonas reinhardtii chloroplast using the lac regulation system from Escherichia coli. J Biosci Bioeng 2007; 104:207–213.PubMedCrossRefGoogle Scholar
  67. 67.
    He DM, Qian KX, Shen GF et al. Recombination and expression of classical swine fever virus (CSFV) structural protein E2 gene in Chlamydomonas reinhardtii chloroplasts. Colloids Surf B Biointerfaces 2007; 55:26–30.PubMedCrossRefGoogle Scholar
  68. 68.
    Wang X, Brandsma M, Tremblay R et al. A novel expression platform for the production of diabetes-associated autoantigen human glutamic acid decarboxylase (hGAD65). BMC Biotechnol 2008; 8:87–89.PubMedCrossRefGoogle Scholar
  69. 69.
    Han S, Hu Z, Lei A. Expression and function analysis of the metallothionein-like (MT-like) gene from Festuca rubra in Chlamydomonas reinhardtii chloroplast. Sci China Ser C-Life Sci 2008; 51:1076–1081.CrossRefGoogle Scholar
  70. 70.
    Korhonen H, Pihlanto A. Food-derived bioactive peptides-opportunities for designing future foods. Curr Pharm 2003; 9:1297–1308.CrossRefGoogle Scholar
  71. 71.
    Korhonen H, Pihlanto A. Bioactive peptides: production and functionality. Int Dairy J 2006; 16:945–960.CrossRefGoogle Scholar
  72. 72.
    Hartmann R, Meisel H. Food-derived peptides with biological activity: from research to food applications. Curr Opin Biotechnol 2007; 18:163–169.PubMedCrossRefGoogle Scholar
  73. 73.
    DeGray G, Rajasekaran K, Smith F et al. Expression of an antimicrobial peptide via the chloroplast genome to control phytopathogenic bacteria and fungi. Plant Physiol 2001; 127:852–862.PubMedCrossRefGoogle Scholar
  74. 74.
    Zasloff M. Antimicrobial peptides of multicellular organisms. Nature 2002; 415:389–395.PubMedCrossRefGoogle Scholar
  75. 75.
    Jacob L, Zasloff M. Potential therapeutic applications of magainins and other antimicrobial agents of animal origin: antimicrobial Peptides. Ciba Found Symp 1994; 186:197–223.PubMedGoogle Scholar

Copyright information

© Landes Bioscience and Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Udo Johanningmeier
    • 1
  • Dirk Fischer
  1. 1.Institut für PflanzenphysiologieMartin-Luther Universität Halle-WittenbergHalle (Saale)Germany

Personalised recommendations