Advertisement

Antisense expression of a caffeic acidO-methyltransferase ofStylosanthes humilis in transgenic poplar: Effect of expression onO-methyltransferase activity and lignin composition

  • Kasim Bajrovic
  • Kemal Kazan
  • Zeliha İpekçi
  • Nermin Gözükırmızı
Original Articles

Abstract

Poplar (Populus tremula) was transformed with a construct carrying an antisense caffeic acidO-methyltransferase (COMT) cDNA (pOMT8) from a tropical pasture legume,Stylosanthes humilis. pOMT8 shows 83% overall homology to the corresponding COMT gene (pPCLA) of poplar. Of the 200 putatively-transformed plants regenerated on selective media after co-cultivation of poplar stem explants withAgrobacterium tumefaciens harbouring a CaMV 35S-antisensepOMT8 construct, a subset of 20 plants were randomly chosen for further analysis. PCR and Southern blot analysis demonstrated the stable integration of T-DNA into the genome of these plants. Antisense expression ofpOMT8 resulted in reductions in total COMT activity in the majority of the transgenic plants with the lowest total COMT activities (61–70% of untransformed control plants) being observed in four transgenic plants. The composition of lignin in transgenic plants was also changed, as detected by reductions in the content of syringyl units using infrared spectroscopy. However, no changes were found in the amount of insoluble lignin in transgenic plants as compared to untransformed control plants. These results indicate the potential of thepOMT8 gene to partially suppress COMT activity and modify the composition of lignin in transgenic poplar.

Key words

caffeic acidO-methyltransferase lignin Populus tremula Stylosanthes humilis 

Literature cited

  1. An, G., Watson, B.D., Stacheh, S., Gordon, M.P., and Nester, E.W. (1985) New cloning vehicles for transformation of higher plants. EMBO 4: 277–284.Google Scholar
  2. Atanassova, R., Favet, N., Martz, F., Chabbert, B., Tollier, M.T., Monties, B., Fritig, B., and Legrand, M. (1995) Altered lignin composition in transgenic tobacco expressingO-methyltransferase sequences in sense and antisense orientation. Plant J. 8: 465–477.CrossRefGoogle Scholar
  3. Boudet, A.M. and Grima-Pettenati, J. (1996) Lignin genetic engineering. Mol. Breeding 2: 25–39.CrossRefGoogle Scholar
  4. Boudet, A.M., Lapierre, C., and Grima-Pettenati, J. (1995) Transley review No. 80. Biochemistry and molecular biology of lignification. New Phytol. 129: 203–236.CrossRefGoogle Scholar
  5. Bradford, M.M. (1976) A rapid and sensitive method for quantitation of microgram quantities of protein utilising the principle of proteindye binding. Anal. Biochem. 72: 248–254.PubMedCrossRefGoogle Scholar
  6. Bugos, R.C., Chiang, V.L.C., and Campbell, W.H. (1991) cDNA cloning, sequence analysis and seasonal expression of lignin-bispecific caffeic acid/5-hydroxyferulic acidO-methyltransferase of aspen. Plant Mol. Biol. 17: 1203–1215.PubMedCrossRefGoogle Scholar
  7. Campbell, M.M. and Ellis, B.E. (1992) Elicited phenylpropanoid metabolism in pine cultures. Planta 186: 409–417.CrossRefGoogle Scholar
  8. Ditta, G., Stanfield, D., Corbin, D., and Helinski, D.R. (1980) Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank ofRhizobium melioti. Proc. Natl. Acad. Sci. USA 77: 7347–7351.PubMedCrossRefGoogle Scholar
  9. Dumas, B., Van Doorsselaere, J., Gielen, J., Legrand, M., Fritig, B., Van Montagu, M., and Inze, D. (1992) Nucleotide sequence of a complementary DNA encodingO-methyltransferase from poplar. Plant Physiol. 98: 796–797.PubMedGoogle Scholar
  10. Dwivedi, U.N., Capbell, W.H., Yu, J., Datla, R.S.S., Bugos, R.C., Chiang, V.L., and Dodila, G.K. (1994) Modification of lignin biosynthesis in transgenicNicotiana through expression of an antisenseO-methyltransferase gene fromPopulus. Plant Mol. Biol. 26: 61–71.PubMedCrossRefGoogle Scholar
  11. Edwards, R. and Dixon, R.A. (1991) Purification and characterisation ofS-adenosyl-l methionine: caffeic acid 3-O-methyltransferase from suspension cultures of alfalfa (Medicago sativa L.). Arch. Biochem. Biophys. 287: 372–379.PubMedCrossRefGoogle Scholar
  12. Effland, M.J. (1977) Modified procedure to determine acid-insoluble lignin in Wood and Pulp. TAPPI 60: 143–144.Google Scholar
  13. Gözükırmızı, N., Bajrovic, K., Ipekçi, Z., Boydak, M., Akalp, T., Tunçtaner, K., Balkan, H., Tanriyar, H., Çalikoglu, M., Oğraş, T., Özden, Ö., Tulukçu, M., and Tank, T. (1998) Genotype differences in direct plant regeneration from stem explants ofPopulus tremula in Turkey. J. For. Res. 3: 123–126.Google Scholar
  14. Halpin, C., Knight, M.E., Foxon, G.A., Campbell, M.M., Boudet, A.M., Boon, J.J., Chappert, B., Tollier, M-T., and Schuch, W. (1994) Manipulation of lignin quality by downregulating of cinnamyl alcohol dehydrogenase. Plant J. 3: 339–350.CrossRefGoogle Scholar
  15. Ibrahim, R.K., Bruneau, A., and Bantignies, B. (1998) PlantO-methyltransferases: molecular analysis, common signature and classification. Plant Mol. Biol. 36: 1–10.PubMedCrossRefGoogle Scholar
  16. Inoune, K., Sewalt, V.J.H., Ballance, G.M., Ni, W., Stürzer, C., and Dixon, R.A. (1998) Developmental expression and substrate specificities of alfalfa caffeic acid 3-O-methyltransferase and caffeoyl coenzyme a 3-O-methyltransferase in relation to lignification. Plant Physiol. 117: 761–770.CrossRefGoogle Scholar
  17. Joseleau, J.P. and Ruel, K. (1997) Study of lignification by non-invasive techniques in growing maize internodes. Plant Physiol. 114: 1123–1133.PubMedCrossRefGoogle Scholar
  18. Kajita, S., Katayama, Y., and Omori, S. (1996) Alterations in the biosynthesis of lignin in transgenic plants with chimeric genes for 4-coumarate:coenzyme a ligase. Plant Cell Physiol. 37: 957–965.PubMedGoogle Scholar
  19. Lagrimini, M.L. (1991) Wound-induced deposition of polyphenols in transgenic plants overexpressing peroxidase. Plant Physiol. 96: 577–583.PubMedGoogle Scholar
  20. Lee, D., Meyer, K., Chapple, C., and Douglas, C.J. (1997) Antisense suppression of 4-coumarate: coenzyme a ligase activity in arabidopsis leads to altered lignin subunit composition. Plant Cell 9: 1985–1998.PubMedCrossRefGoogle Scholar
  21. Legrand, M., Atanassova, R., Favet, N., Martz, F., Chabbert, B., Tolier, M.T., Monties, B., and Fritig, B. (1994) Inhibition ofO-methyltransferase (OMT) activity in transgenic tobacco plants modified lignin monomeric composition. International Plant Molecular Biology Meeting, Amsterdam.Google Scholar
  22. Leple, J.C., Miranda Brasileiro, A.C., Michel, M.F., Delmotte, F., and Jouanin, L. (1992) Transgenic poplars: expression of chimerical genes using four different constructs. Plant Cell Rep. 11: 137–141.CrossRefGoogle Scholar
  23. Lewis, N.G. and Yamamoto, E. (1990) Occurrence, biogenesis and biodegration. Ann. Rev. Plant Physiol. Lignin: Plant Mol. Biol. 41: 455–496.CrossRefGoogle Scholar
  24. Lloyd, G.B. and McCown, B.H. (1981) Commercially feasible micropropagation of mountain Laurel,Kalmia latifolia, by use of shoot-tip culture. Proc. Intl. Plant Prop. Soc. 30: 421–427.Google Scholar
  25. McIntyre, C.L., Rae, A.L., Curtis, M.D., and Manners, J.M. (1995) Sequence and expression of a caffeic acidO-methyltransferase cDNA homologue in the tropical forage legumeStylosanthes humilis. Aust. J. Plant Physiol. 22: 471–478.CrossRefGoogle Scholar
  26. Ni, W., Paiva, N.L., and Dixon, R.A. (1994) Reduced lignin in transgenic plants containing a caffeic acidO-methyltransferase antisense gene. Transgenic Res. 3: 120–126.CrossRefGoogle Scholar
  27. O’malley, D.M., Whetten, R., Bao, W., Chen, C-L., and Sedereoff, R.R. (1993) The role of laccase in lignification. Plant J. 4: 751–757.CrossRefGoogle Scholar
  28. Sewalt, V.J.H., Ni, W., Blount, J.W., Jung, H.G., Masoud, H.A., Howels, P.A., Lamb, C., and Dixon, R.A. (1997) Reduced lignin content and altered lignin composition in transgenic tobacco down-regulated in expression ofl-phenylalanine ammonia-lyase or cinnamate-4-hydroxylase. Plant Physiol. 115: 41–50.PubMedGoogle Scholar
  29. Tien, M. (1987) Properties of lignin asses fromPhanerochaete chrysosporium and their possible application. C.R.C. Crit. Rev. Microbiol. 15: 141–168.CrossRefGoogle Scholar
  30. Van Doorsselaere, J., Baucher M., Chognot, E., Chabbert, B., Tollier, M.T., Petit-Conil, M., Leple, J.C., Pilate, G., Cornu, D., Monties, B., Van Montagu, M., Inze, D., Boerjan, W., and Jouanin, L. (1995) A novel lignin in poplar trees with a reduced caffeic acid/5-hydroxyferulic acidO-methyltransferase activity. Plant J. 8: 855–864.Google Scholar
  31. Walbot, V. (1988) Preparation of DNA from single rice seedlings. Rice Genet. Newslett. 5: 149–151.Google Scholar
  32. Whetten, R.W. and Sederoff, R.R. (1995) Lignin biosynthesis. Plant Cell 7: 1001–1013.PubMedCrossRefGoogle Scholar

Copyright information

© The Japanese Forest Society and Springer 1999

Authors and Affiliations

  • Kasim Bajrovic
    • 1
  • Kemal Kazan
    • 2
  • Zeliha İpekçi
    • 1
  • Nermin Gözükırmızı
    • 1
    • 2
  1. 1.Research Institute for Genetic Engineering and BiotechnologyTÜBITAK, MRCGebze-KocaeliTurkey
  2. 2.Cooperative Research Center for Tropical Plant PathologyUniversity of Queensland BrisbaneAustralia

Personalised recommendations