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Acta Biologica Hungarica

, Volume 65, Issue 2, pp 189–204 | Cite as

Biomass Derived from Transgenic Tobacco Expressing the Arabidopsis CESA3ixr1—2 Gene Exhibits Improved Saccharification

  • Dipak Kumar SahooEmail author
  • Indu B. Maiti
Article

Abstract

Studies in Arabidopsis thaliana and Nicotiana tabacum L. variety Samsun NN demonstrated that expression of the CESA3 cellulose synthase gene that contains a point mutation, named ixr1–2, results in greater conversion of plant-derived cellulose to fermentable sugars. The present study was designed to examine the improved enzymatic saccharification efficiency of lignocellulosic biomass of tobacco plants expressing AtCESA3ixr1–2. Three-month-old AtCESA3ixr1–2 transgenic and wild-type tobacco plants (Nicotiana tabacum L. variety Samsun NN) were grown in the presence and absence of isoxaben. Biomass obtained from leaf, stem, and root tissues were analyzed for enzymatic saccharification rates. During enzymatic saccharification, 45% and 25% more sugar was released from transgenic leaf and stem samples, respectively, when compared to the wild-type samples. This gain in saccharification efficiency was achieved without chemical or heat pretreatment. Additionally, leaf and stem biomass from transgenic AtCESA3ixr1–2 requires a reduced amount of enzyme for saccharification compared to biomass from wild-type plants. From a practical standpoint, a similar strategy could be employed to introduce the mutated CESA into energy crops like poplar and switchgrass to improve the efficiency of biomass conversion.

Keywords

Nicotiana tabacum biofuel biomass AtCESA3ixr1—2 

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References

  1. 1.
    Ambavaram, M. M., Krishnan, A., Trijatmiko, K. R., Pereira, A. (2011) Coordinated activation of cellulose and repression of lignin biosynthesis pathways in rice. Plant Physiol. 155, 916–931.CrossRefGoogle Scholar
  2. 2.
    Andrianov, V., Borisjuk, N., Pogrebnyak, N., Brinker, A., Dixon, J., Spitsin, S., Flynn, J., Matyszczuk, P., Andryszak, K., Laurelli, M., Golovkin, M., Koprowski, H. (2009) Tobacco as a pro bduction platform for biofuel: overexpression of Arabidopsis DGAT and LEC2 genes increases accumulation and shifts the composition of lipids in green biomass. Plant Biotechnol. J. 8, 1–11.Google Scholar
  3. 3.
    Banerjee, J., Sahoo, D. K., Dey, N., Houtz, R. L., Maiti, I. B. (2013) An intergenic region shared by At4g35985 and At4g35987 in Arabidopsis thaliana is a tissue specific and stress inducible bidirectional promoter analyzed in transgenic Arabidopsis and tobacco plants. PLoS One 8(11), e79622. doi:10.1371/journal.pone.0079622.CrossRefGoogle Scholar
  4. 4.
    Brunecky, R., Selig, M. J., Vinzant, T. B., Himmel, M. E., Lee, D., Blaylock, M. J., Decker, S. R. (2011) In planta expression of A. cellulolyticus Cel5A endocellulase reduces cell wall recalcitrance in tobacco and maize. Biotechnol. Biofuels 4, 1.CrossRefGoogle Scholar
  5. 5.
    Chapple, C., Ladisch, M., Meilan, R. (2007) Loosening lignin’s grip on biofuel production. Nat. Biotechnol. 25(7), 746–748.CrossRefGoogle Scholar
  6. 6.
    Chen, F., Dixon, R. A. (2007) Lignin modification improves fermentable sugar yields for biofuel production. Nat. Biotechnol. 25(7), 759–761.CrossRefGoogle Scholar
  7. 7.
    Chuck, G. S., Tobias, C., Sun, L., Kraemer, F., Li, C., Dibble, D., Arora, R., Bragg, J. N., Vogel, J. P., Singh, S., Simmons, B. A., Pauly, M., Hake, S. (2011) Overexpression of the maize Corngrass1 microRNA prevents flowering, improves digestibility, and increases starch content of switchgrass. Proc. Natl. Acad. Sci. USA 108(42), 17550–17555.CrossRefGoogle Scholar
  8. 8.
    Desprez, T., Juraniec, M., Crowell, E. F., Jouy, H., Pochylova, Z., Parcy, F., Hofte, H., Gonneau, M., Vernhettes, S. (2007) Organization of cellulose synthase complexes involved in primary cell wall synthesis in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA, 104, 15572–15577.CrossRefGoogle Scholar
  9. 9.
    Harris, D. M., Corbin, K., Wang, T., Gutierrez, R., Bertolo, A. L., Petti, C., Smilgies, D. M., Estevez, J. M., Bonetta, D., Urbanowicz, B. R., Ehrhardt, D., Somerville, C. R., Rose, J. K. C., Hong, M., DeBolt, S. (2012) Cellulose microfibril crystallinity is reduced by mutating C-terminal transmem brane region residues CESA1A903V and CESA3T942I of cellulose synthase. Proc. Natl. Acad. Sci. USA 109, 4098–4103.CrossRefGoogle Scholar
  10. 10.
    Harris, D., DeBolt, S. (2010) Synthesis, regulation and utilization of lignocellulosic biomass. Plant Biotechnol. J. 8, 244–262.CrossRefGoogle Scholar
  11. 11.
    Harris, D., Stork, J., DeBolt, S. (2009) Genetic modification in cellulose-synthase reduces crystallinity and improves biochemical conversion to fermentable sugar. GCB Bioenergy 1, 51–61.CrossRefGoogle Scholar
  12. 12.
    Heinzelman, P., Snow, C. D., Wu, I., Nguyen, C., Villalobos, A., Govindarajan, S., Minshull, J., Arnold, F. H. (2009) A family of thermostable fungal cellulases created by structure-guided recombination. Proc. Natl. Acad. Sci. USA 106(14), 5610–5615.CrossRefGoogle Scholar
  13. 13.
    Hématy, K., Höfte, H. (2006) Cellulose and cell elongation. Plant Cell Monograph 5, 33–56.Google Scholar
  14. 14.
    Himmel, M. E., Ding, S. Y., Johnson, D. K., Adney, W. S., Nimlos, M. R., Brady, J. W., Foust, T. D. (2007) Biomass recalcitrance: engineering plants and enzymes for biofuels production. Science 315, 804–807.CrossRefGoogle Scholar
  15. 15.
    Kroumova, A. B., Sahoo, D. K., Raha, S., Goodin, M., Maiti, I. B., Wagner, G. J. (2013) Expression of an apoplast-directed, T-phylloplanin-GFP fusion gene confers resistance against Peronospora tabacina disease in a susceptible tobacco. Plant Cell Rep. 32, 1771–1782.CrossRefGoogle Scholar
  16. 16.
    Kumar, D., Patro, S., Ranjan, R., Sahoo, D. K., Maiti, I. B., Dey, N. (2011) Development of useful recombinant promoter and its expression analysis in different plant cells using confocal laser scanning microscopy. PLoS One 6(9), e24627. doi:10.1371/journal.pone.0024627.CrossRefGoogle Scholar
  17. 17.
    Lee, D., Yu, A. H. C., Wong, K. K. Y., Saddler, J. N. (1994) Evaluation of the enzymatic susceptibility of cellulosic substrates using specific hydrolysis rates and enzyme adsorption. Appl. Biochem. Biotechnol. 45/46, 407–415.CrossRefGoogle Scholar
  18. 18.
    Liu, W., Hong, J., Bevan, D. R., Zhang, Y. H. P. (2009) Fast identification of thermostable beta-glucosidase mutants on cellobiose by a novel combinatorial selection/screening approach. Biotechnol. Bioeng. 103, 1087–1094.CrossRefGoogle Scholar
  19. 19.
    Mosier, N., Wyman, C. E., Dale, B. E., Elander, R. T., Lee, Y. Y., Holtzapple, M., Ladisch, M. (2005) Features of promising technologies for pretreatment of lignocellulosic biomass. Bioresour. Technol. 96, 673–686.CrossRefGoogle Scholar
  20. 20.
    Ralph, J., Akiyama, T., Kim, H., Lu, F., Schatz, P. F., Marita, J. M., Ralph, S. A., Reddy, M. S. S., Chen, F., Dixon, R. A. (2006) Effects of coumarate 3-hydroxylase down-regulation on lignin structure. J. Biol. Chem. 281, 8843–8853.CrossRefGoogle Scholar
  21. 21.
    Reiter, W. D., Chapple, C. C. S., Somerville, C. R. (1993) Altered growth and cell walls in a fucosedeficient mutant of Arabidopsis. Science 261, 1032–1035.PubMedGoogle Scholar
  22. 22.
    Rollin, J. A., Zhu, Z., Sathitsuksanoh, N., Zhang, Y. H. P. (2011) Increasing cellulose accessibility is more important than removing lignin: A comparison of cellulose solvent-based lignocellulose fractionation and soaking in aqueous ammonia. Biotechnol. Bioeng. 108, 22–30.CrossRefGoogle Scholar
  23. 23.
    Sahoo, D. K., Ranjan, R., Kumar, D., Kumar, A., Sahoo, B. S., Raha, S., Maiti, I. B., Dey, N. (2009) An alternative method of promoter assessment by confocal laser scanning microscopy. J. Virol. Methods 161, 114–121.CrossRefGoogle Scholar
  24. 24.
    Sahoo, D. K., Stork, J., Debolt, S., Maiti, I. B. (2013) Manipulating cellulose biosynthesis by expression of mutant Arabidopsis proM24::CESA3(ixr1–2) gene in transgenic tobacco. Plant Biotechnol. J. 11, 362–372.CrossRefGoogle Scholar
  25. 25.
    Sathitsuksanoh, N., Zhu, Z., Ho, T. J., Bai, M. D., Zhang, Y. H. P. (2010) Bamboo saccharification through cellulose solvent-based biomass pretreatment followed by enzymatic hydrolysis at ultra-low cellulase loadings. Biores. Technology 101, 4926–4929.CrossRefGoogle Scholar
  26. 26.
    Sathitsuksanoh, N., Zhu, Z., Templeton, N., Rollin, J., Harvey, S., Zhang, Y. H. P. (2009) Saccharification of a potential bioenergy crop, Phragmites australis (common reed), by lignocellulose fractionation followed by enzymatic hydrolysis at decreased cellulase loadings. Ind. Eng. Chem. Res. 48, 6441–6447.CrossRefGoogle Scholar
  27. 27.
    Scheller, H. V., Ulvskov, P. (2010) Hemicelluloses. Annu. Rev. Plant Biol. 61, 263–289.CrossRefGoogle Scholar
  28. 28.
    Schillberg, S., Fischer, R., Emans, N. (2003) Molecular farming of antibodies in plants. Naturwissenschaften 90, 145–155.PubMedGoogle Scholar
  29. 29.
    Scott, T. A., Melvin, E. H. (1953) The determination of dextran with anthrone. Anal. Chem. 25, 1656–1661.CrossRefGoogle Scholar
  30. 30.
    Somerville, C. (2006) Cellulose synthesis in higher plants. Annu. Rev. Cell. Dev. Biol. 22, 53–78.CrossRefGoogle Scholar
  31. 31.
    Sticklen, M. (2006) Plant genetic engineering to improve biomass characteristics for biofuels. Curr. Opin. Biotechnol. 17(3), 315–319.CrossRefGoogle Scholar
  32. 32.
    Tu, M., Chandra, R. P., Saddler, J. N. (2007) Evaluating the distribution of cellulases and the recycling of free cellulases during the hydrolysis of lignocellulosic substrates. Biotechnol. Prog. 23(2), 398–406.CrossRefGoogle Scholar
  33. 33.
    Updegraff, D. M. (1969) Semimicro determination of cellulose in biological materials. Anal. Biochem. 32(3), 420–424.CrossRefGoogle Scholar
  34. 34.
    Wyman, C. E. (2007) What is (and is not) vital to advancing cellulosic ethanol. Trends Biotechnol. 25(4), 153–157.CrossRefGoogle Scholar
  35. 35.
    Zhang, Y. H., Cui, J., Lynd, L. R., Kuang, L. R. (2006) A transition from cellulose swelling to cellulose dissolution by o-phosphoric acid: evidence from enzymatic hydrolysis and supramolecular structure. Biomacromolecules 7(2), 644–648.CrossRefGoogle Scholar
  36. 36.
    Zhu, Z., Sathitsuksanoh, N., Zhang, Y. H. P. (2009) Direct quantitative determination of adsorbed cellulase on lignocellulosic biomass with its application to study cellulase desorption for potential recycling. Analyst 134 (11), 2267–2272.CrossRefGoogle Scholar

Copyright information

© Akadémiai Kiadó Zrt. 2014

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

Authors and Affiliations

  1. 1.KTRDC, College of Agriculture, Food and EnvironmentUniversity of KentuckyLexingtonUSA

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