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Repression of lignin biosynthesis promotes cellulose accumulation and growth in transgenic trees

Abstract

Because lignin limits the use of wood for fiber, chemical, and energy production, strategies for its downregulation are of considerable interest. We have produced transgenic aspen (Populus tremuloides Michx.) trees in which expression of a lignin biosynthetic pathway gene Pt4CL1 encoding 4-coumarate:coenzyme A ligase (4CL) has been downregulated by antisense inhibition. Trees with suppressed Pt4CL1 expression exhibited up to a 45% reduction of lignin, but this was compensated for by a 15% increase in cellulose. As a result, the total lignin–cellulose mass remained essentially unchanged. Leaf, root, and stem growth were substantially enhanced, and structural integrity was maintained both at the cellular and whole-plant levels in the transgenic lines. Our results indicate that lignin and cellulose deposition could be regulated in a compensatory fashion, which may contribute to metabolic flexibility and a growth advantage to sustain the long-term structural integrity of woody perennials.

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Figure 1: Biosynthetic pathways to guaiacyl and syringyl monolignols for the formation of guaiacyl:syringyl lignin in woody angiosperms.
Figure 2: The effects of downregulation of Pt4CL1 expression on Pt4CL1 activity and lignin accumulation in transgenic aspen.
Figure 3: HSQC spectra of isolated milled wood lignins from (A) control and (B) transgenic line A6 indicate the similarity of major lignin structural units in both samples.
Figure 4: Enhanced growth in transgenic aspen.

References

  1. Freudenberg, K. Lignin: its constitution and formation from p-hydroxycinnamyl alcohols. Science 148, 595–600 (1965).

    Article  CAS  Google Scholar 

  2. Whetten, R.W., MacKay, J.J. & Sederoff, R.R. Recent advances in understanding lignin biosynthesis. Annu. Rev. Plant Physiol. Plant Mol. Biol. 49, 585–609 (1998).

    Article  CAS  Google Scholar 

  3. Van Doorsselaere, J. et al. A novel lignin in poplar trees with a reduced caffeic acid/5-hydroxyferulic acid O-methyltransferase activity. Plant J. 8, 855–864 (1995).

    Article  CAS  Google Scholar 

  4. Tsai, C.-J. et al. Suppression of O-methyltransferase gene by homologous sense transgene in quaking aspen causes red-brown wood phenotypes. Plant Physiol. 117, 101–112 (1998).

    Article  CAS  Google Scholar 

  5. Baucher, M. et al. Red xylem and higher lignin extractability by down-regulating a cinnamyl alcohol dehydrogenase in poplar. Plant Physiol. 112, 1479–1490 (1996).

    Article  CAS  Google Scholar 

  6. Dwivedi, U.N. et al. Modification of lignin biosynthesis in transgenic Nicotiana through expression of an antisense O-methyltransferase gene from Populus. Plant Mol. Biol. 26, 61– 71 (1994).

    Article  CAS  Google Scholar 

  7. Halpin, C. et al. Manipulation of lignin quality by downregulation of cinnamyl alcohol dehydrogenase. Plant J. 6, 339– 350 (1994).

    Article  CAS  Google Scholar 

  8. Atanassova, R. et al. Altered lignin composition in transgenic tobacco expressing O-methyltransferase sequences in sense and antisense orientation. Plant J. 8, 465–477 ( 1995).

    Article  CAS  Google Scholar 

  9. Hibino, T., Takabe, K., Kawazu, T., Shibata, D. & Higuchi, T. Increase of cinnamaldehyde groups in lignin of transgenic tobacco plants carrying an antisense gene for cinnamyl alcohol dehydrogenase. Biosci. Biotech. Biochem. 59, 929– 931 (1995).

    Article  CAS  Google Scholar 

  10. Elkind, Y. et al. Abnormal plant development and down-regulation of phenylpropanoid biosynthesis in transgenic tobacco containing a heterologous phenylalanine ammonia-lyase gene. Proc. Natl. Acad. Sci USA 87, 9057–9061 (1990).

    Article  CAS  Google Scholar 

  11. Bate, N. et al. Quantitative relationship between phenylalanine ammonia-lyase levels and phenylpropanoid accumulation in transgenic tobacco identifies a rate-limiting step in natural product synthesis. Proc. Natl. Acad. Sci. USA 91, 7608–7612 ( 1994).

    Article  CAS  Google Scholar 

  12. Kajita, S., Katayama, Y. & Omori, S. Alterations in the biosynthesis of lignin in transgenic plants with chimeric genes for 4-coumarate:coenzyme A ligase. Plant Cell Physiol. 37, 957–965 (1996).

    Article  CAS  Google Scholar 

  13. Kajita, S., Hishiyama, S., Tomimura, Y., Katayama, Y. & Omori, S. Structural characterization of modified lignin in transgenic tobacco plants in which the activity of 4-coumarate:coenzyme A ligase is depressed. Plant Physiol. 114, 871–879 (1997).

    Article  CAS  Google Scholar 

  14. Lee, D., Meyer, K., Chapple, C. & Douglas, C.J. Antisense suppression of 4-coumarate:coenzyme A ligase activity in Arabidopsis leads to altered lignin subunit composition. Plant Cell 9, 1985–1998 (1997).

    Article  CAS  Google Scholar 

  15. Piquemal, J. et al. Down-regulation of cinnamoyl-CoA reductase induces significant changes of lignin profiles in transgenic tobacco plants. Plant J. 13, 71–83 ( 1998).

    Article  CAS  Google Scholar 

  16. Hu, W.-J. et al. Compartmentalized expression of two structurally and functionally distinct 4-coumarate:CoA ligase genes in aspen (Populus tremuloides). Proc. Natl. Acad. Sci. USA 95, 5407– 5412 (1998).

    Article  CAS  Google Scholar 

  17. Hakomori, S.I. A rapid permethylation of glycolipids and polysaccharides catalyzed by methylsulfinyl carbanion in dimethyl sulphoxide. J. Biochem. Tokyo 55, 205–208 (1964).

    CAS  PubMed  Google Scholar 

  18. Ciucanu, I. & Kerek, F. A simple and rapid method for the permethylation of carbohydrates. Carbohydr. Res. 131 , 209–217 (1984).

    Article  CAS  Google Scholar 

  19. Arioli, T. et al. Molecular analysis of cellulose biosynthesis in Arabidopsis . Science 279, 717– 720 (1997).

    Article  Google Scholar 

  20. Delmer, D.P. Biosynthesis of cellulose. Annu. Rev. Plant Physiol. 38, 259–290 (1987).

    Article  CAS  Google Scholar 

  21. Sewalt, V.J.H. et al. Reduced lignin content and altered lignin composition in transgenic tobacco down-regulated in expression of L-phenylalanine ammonia-lyase or cinnamate 4-hydroxylase. Plant Physiol. 115, 41–50 (1997).

    Article  CAS  Google Scholar 

  22. Tan, K.S., Hoson, T., Masuda, Y. & Kamisaka, S. Effects of ferulic and p-coumaric acids on Oryza coleoptiole growth and the mechanical properties of cell walls. J. Plant Physiol. 140, 460–465 (1992).

    Article  CAS  Google Scholar 

  23. Mock H.-P. & Strack, D. Energetics of the uridine 5´-diphosphoglucose:hydroxycinnamic acid acyl-glucosyltransferase reaction. Phytochem. 32, 575–579 (1993).

    Article  CAS  Google Scholar 

  24. Timell, T.E. Compression wood in gymnosperms. (Springer, New York; 1986).

    Book  Google Scholar 

  25. Pear, J.R., Kawagoe, Y., Schreckengost, W.E., Delmer, D.P. & Stalker, D.M. Higher plants contain homologs of the bacterial celA genes encoding the catalytic subunit of cellulose synthase. Proc. Natl. Acad. Sci. USA 93, 12637–12642 (1996).

    Article  CAS  Google Scholar 

  26. Creelman, R.A. & Mullet, J.E. Oligosaccharins, brassinolides, and jasmonates: nontraditional regulators of plant growth, development, and gene expression. Plant Cell 9, 1211–1223 (1997).

    Article  CAS  Google Scholar 

  27. Logemann, E. et al. Gene activation by UV light, fungal elicitor or fungal infection in Petroselinum crispum is correlated with repression of cell cycle-related genes. Plant J. 8, 865– 876 (1995).

    Article  CAS  Google Scholar 

  28. Lee, T.T. & Skoog, S. Effects of substituted phenols on bud formation and growth of tobacco tissue cultures Physiol. Plant. 18, 386–402 ( 1965).

    Article  CAS  Google Scholar 

  29. Nitsch, J.P. & Nitsch, C. Composes phenolique et croissance vegetale. Ann. Physiol. Veg. 4, 211– 225 (1962).

    CAS  Google Scholar 

  30. Zenk, M.H. & Muller, G.Z. In vivo destruction of exogenously applied indole-acetic acid as influenced by phenolic acids. Nature 200, 761–763 ( 1963).

    Article  CAS  Google Scholar 

  31. Hahlbrock, K. & Grisebach, H. Enzymic controls in the biosynthesis of lignin and flavonoids. Ann. Rev. Plant Physiol. 30, 105–130 (1979).

    Article  CAS  Google Scholar 

  32. Shirley, B.W. Flavonoid biosynthesis: new functions for an old pathway. Trends Plant Sci. 1, 377–382 ( 1996).

    Google Scholar 

  33. Jacobs, M. & Rubery, P.H. Naturally occurring auxin transport regulators. Science 241, 346– 349 (1988).

    Article  CAS  Google Scholar 

  34. Ruegger, M. et al. Reduced naphthylphthalamic acid binding in the tir3 mutant of Arabidopsis is associated with a reduction in polar auxin transport and diverse morphological defects. Plant Cell 9, 745–757 (1997).

    Article  CAS  Google Scholar 

  35. Tsai, C.-J., Podila, G.K. & Chiang, V.L. Agrobacterium-mediated transformation of quaking aspen (Populus tremuloides) and regeneration of transgenic plants. Plant Cell Rep. 14, 94– 97 (1994).

    CAS  PubMed  Google Scholar 

  36. Datla, R.S.S., et al. Improved high-level constitutive foreign gene expression in plants using an AMV RNA4 untranslated leader sequence. Plant Sci. 94, 139–149 ( 1993).

    Article  CAS  Google Scholar 

  37. Bugos, R.C. et al. RNA isolation from plant tissues recalcitrant to extraction in guanidine. Biotechniques 19, 734– 737 (1995).

    CAS  PubMed  Google Scholar 

  38. Chiang, V.L. & Funaoka, M. The dissolution and condensation reactions of guaiacyl and syringyl units in residual lignin during kraft delignification of sweetgum. Holzforschung 44, 147– 155 (1990).

    Article  CAS  Google Scholar 

  39. Rolando, C., Monties, B. & Lapierre, C. in Methods in lignin chemistry (eds Lin, S.Y. & Dence, C.W.) 334–349 (Springer, New York; 1992).

    Book  Google Scholar 

  40. Ralph, J. et al. Pathway of p-coumaric acid incorporation into maize lignin as revealed by NMR. J. Am. Chem. Soc. 116, 9448–9456 (1994).

    Article  CAS  Google Scholar 

  41. Palmer, A.G., Cavanagh, J., Wright, P.E. & Rance, M. Sensitivity improvement in proton-detected two-dimensional heteronuclear correlation NMR spectroscopy. J. Magn. Reson. A 93, 151–170 (1991).

    CAS  Google Scholar 

  42. Braunschweiler, L. & Ernst, R.R. Coherence transfer by isotropic mixing: application to proton correlation spectroscopy. J. Magn. Reson. 53, 521–528 (1983).

    CAS  Google Scholar 

  43. Ruiz-Cabello, J. et al. Gradient-enhanced heteronuclear correlation spectroscopy. Theory and experimental aspects. J. Magn. Reson. 100 , 282–302 (1992).

    CAS  Google Scholar 

  44. Hartley, R.D. Improved methods for the estimation by gas-liquid chromatography of lignin degradation products from plants. Chromatography 54 , 335–344 (1971).

    Article  CAS  Google Scholar 

  45. Chiang, V.L. & Sarkanen, K.V. Ammonium sulfide organosolv pulping. Wood Sci. Technol. 17, 217– 226 (1983).

    Article  CAS  Google Scholar 

  46. Davis, M.W. A rapid modified method for compositional carbohydrate analysis of lignocellulosics by high pH anion-exchange chromatography with pulsed amperometric detection (HPAEC/PAD). J. Wood Chem. Technol. 18, 235–252 (1988).

    Article  Google Scholar 

Download references

Acknowledgements

We thank R.R. Sederoff and G.D. McGinnis for critical reading of the manuscript; C.P. Joshi and A. Kawaoka for helpful discussions; H. Suzuki for preparing the antisense Pt4CL1 gene construct; J. Marita, F. Lu, and R. Hatfield for assistance and discussions on NMR work; M.W. Davis for compositional carbohydrate analysis; and P. Azadi for polysaccharide linkage analysis. Supported in part by grants from the National Science Foundation, the USDA-National Research Initiative Competitive Grants Program, and the USDA-McIntire-Stennis Forestry Research Program.

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Correspondence to Vincent L. Chiang.

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Hu, WJ., Harding, S., Lung, J. et al. Repression of lignin biosynthesis promotes cellulose accumulation and growth in transgenic trees. Nat Biotechnol 17, 808–812 (1999). https://doi.org/10.1038/11758

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