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Functional analysis of members of the isoflavone and isoflavanone O-methyltransferase enzyme families from the model legume Medicago truncatula

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Abstract

Previous studies have identified two distinct O-methyltransferases (OMTs) implicated in isoflavonoid biosynthesis in Medicago species, a 7-OMT methylating the A-ring 7-hydroxyl of the isoflavone daidzein and a 4’-OMT methylating the B-ring 4′-hydroxyl of 2,7,4′-trihydroxyisoflavanone. Genes related to these OMTs from the model legume Medicago truncatula cluster as separate branches of the type I plant small molecule OMT family. To better understand the possible functions of these related OMTs in secondary metabolism in M. truncatula, seven of the OMTs were expressed in E. coli, purified, and their in vitro substrate preferences determined. Many of the enzymes display promiscuous activities, and some exhibit dual regio-specificity for the 4′ and 7-hydroxyl moieties of the isoflavonoid nucleus. Protein structure homology modeling was used to help rationalize these catalytic activities. Transcripts encoding the different OMT genes exhibited differential tissue-specific and infection- or elicitor-induced expression, but not always in parallel with changes in expression of confirmed genes of the isoflavonoid pathway. The results are discussed in relation to the potential in vivo functions of these OMTs based on our current understanding of the phytochemistry of M. truncatula, and the difficulties associated with gene annotation in plant secondary metabolism.

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Abbreviations

EST:

expressed sequence tag

IFS:

isoflavone synthase

MeJA:

methyl jasmonate

OMT:

O-methyltransferase

TC:

tentative consensus

YE:

yeast elicitor

References

  • Akashi T, Sawada Y, Aoki T, Ayabe S-I (2000) New scheme of the biosynthesis of formononetin involving 2,7,4′-trihydroxyisoflavanone but not daidzein as the methyl acceptor. Biosci Biotechnol Biochem 64:2276–2279

    Article  PubMed  CAS  Google Scholar 

  • Akashi T, Sawada Y, Shimada H, Sakurai N, Aoki T, Ayabe S (2003) cDNA cloning and biochemical characterization of S-adenosyl-L-methionine: 2,7,4′-trihydroxyisoflavanone 4′-O-methyltransferase, a critical enzyme of the legume isoflavonoid phytoalexin pathway. Plant Cell Physiol 44:103–112

    Article  PubMed  CAS  Google Scholar 

  • Bisby FA, Buckingham J, Harborne JB (Eds) (1994) Phytochemical dictionary of the Leguminosae, Vol I. Plants and their constituents. Chapman and Hall, New York

    Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  PubMed  CAS  Google Scholar 

  • Deavours BE, Dixon RA (2005) Metabolic engineering of isoflavonoid biosynthesis in alfalfa. Plant Physiol 138:2245–2259

    Article  PubMed  CAS  Google Scholar 

  • Dixon RA, Steele CL (1999) Flavonoids and isoflavonoids- a gold mine for metabolic engineering. Trends Plant Sci 4:394–400

    Article  PubMed  Google Scholar 

  • Edwards R, Dixon RA (1991) Isoflavone O-methyltransferase activities in elicitor-treated cell suspension cultures of Medicago sativa. Phytochemistry 30:2597–2606

    Article  CAS  Google Scholar 

  • Ferrer J-L, Zubieta C, Dixon RA, Noel JP (2005) Crystal structure of alfalfa caffeoyl coenzyme A 3-O-methyltransferase. Plant Physiol 137:1009–1017

    Article  PubMed  CAS  Google Scholar 

  • Frick S, Kutchan TM (1999) Molecular cloning and functional expression of O-methyltransferases common to isoquinoline alkaloid and phenylpropanoid biosynthesis. Plant J 17:329–339

    Article  PubMed  CAS  Google Scholar 

  • Frick S, Ounaroon A, Kutchan TM (2001) Combinatorial biochemistry in plants: the case of O-methyltransferases. Phytochemistry 56:1–4

    Article  PubMed  CAS  Google Scholar 

  • Gang DR, Lavid N, Zubieta C, Chen F, Beuerle T, Lewinsohn E, Noel JP, Pichersky E (2002) Characterization of phenylpropene O-methyltransferases from sweet basil: facile change of substrate specificity and convergent evolution within a plant O-methyltransferase family. Plant Cell 14:505–519

    Article  PubMed  CAS  Google Scholar 

  • Harborne JB (Eds.), (1994) The Flavonoids, advances in research since 1986. Chapman & Hall, London

    Google Scholar 

  • Harrison MJ, Dixon RA (1993) Isoflavonoid accumulation and expression of defense gene transcripts during the establishment of vesicular arbuscular mycorrhizal associations in roots of Medicago truncatula. Mol Plant-Microbe Interact 6:643–654

    CAS  Google Scholar 

  • He X-Z, Dixon RA (1996) Affinity chromatography, substrate/product specificity and amino acid sequence analysis of an isoflavone O-methyltransferase from alfalfa (Medicago sativa L.). Arch Biochem Biophys 336:121–129

    Article  PubMed  CAS  Google Scholar 

  • He X-Z, Dixon RA (2000) Genetic manipulation of isoflavone 7-O-methyltransferase enhances the biosynthesis of 4′-O-methylated isoflavonoid phytoalexins and disease resistance in alfalfa. Plant Cell 12:1689–1702

    Article  PubMed  CAS  Google Scholar 

  • He X-Z, Reddy JT, Dixon RA (1998) Stress responses in alfalfa (Medicago sativa L.) XXII. cDNA cloning and characterization of an elicitor-inducible isoflavone 7-O-methyltransferase. Plant Mol Biol 36:43–54

    Article  PubMed  CAS  Google Scholar 

  • Hoffmann L, Maury S, Bergdoll M, Thion L, Erard M, Legrand M (2001) Identification of the enzymatic active site of tobacco caffeoyl-coenzyme A O-methyltransferase by site-directed mutagenesis. J Biol Chem 276:36831–36838

    Article  PubMed  CAS  Google Scholar 

  • Ibrahim RK, De Luca V, Khouri H, Latchinian L, Brisson L, Charest PM (1987) Enzymology and compartmentation of polymethylated flavonol glucosides in Chrysosplenium americanum. Phytochemistry 26:1237–1245

    Article  Google Scholar 

  • Jones TA, Zou J-Y, Cowan SW, Kjeldgaard M (1991) Improved methods for building protein models in electron density maps and the location of errors in these models. Acta. Crystallogr A49:148–157

    Google Scholar 

  • Joshi CP, Chiang VL (1998) Conserved sequence motifs in plant S-adenosyl-L-methionine-dependent methyltransferases. Plant Mol. Biol 37:663–674

    Article  PubMed  CAS  Google Scholar 

  • Kim DH, Kim B-G, Lee Y, Ryu JY, Lim Y, Hur H-G, Ahn JH (2005) Regiospecific methylation of naringenin to ponciretin by soybean O-methyltransferase expressed in Escherichia coli. J Biotech 119:155--162

    Google Scholar 

  • Kornblatt J, Muzac I, Lim Y, Ahn JH, Ibrahim RK (2004) Role of serine 286 in cosubstrate binding and catalysis of a flavonol O-methyltransferase. Biochem. Cell Biol 82:531–537

    Article  PubMed  CAS  Google Scholar 

  • Laskowski RA, MacArthur MW, Moss DS, Throrton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures J. Appl. Cryst 26:283–291

    Article  CAS  Google Scholar 

  • Liu C-J, Deavours BE, Richard SB, Ferrer J-L, Dixon RA, Noel JP (2005) Dual functionality of isoflavonoid O-methyltransferases in the evolution of plant defense responses. Plant Cell (in review)

  • Liu C-J, Dixon RA (2001) Elicitor-induced association of isoflavone O-methyltransferase with endomembranes prevents formation and 7-O-methylation of daidzein during isoflavonoid phytoalexin biosynthesis. Plant Cell 13:2643–2658

    Article  PubMed  CAS  Google Scholar 

  • Marti-Renom MA, Stuart A, Fiser A, Sanchez R, Melo F, Sali A (2000) Comparative protein structure modeling of genes and genomes. Annu Rev Biophys Biomol Struct 29:291–325

    Article  PubMed  CAS  Google Scholar 

  • Maxwell CA, Harrison MJ, Dixon RA (1993) Molecular characterization and expression of alfalfa isoliquiritigenin 2′-O-methyltransferase, an enzyme specifically involved in the biosynthesis of an inducer of Rhizobium meliloti nodulation genes. Plant J 4:971–981

    Article  PubMed  CAS  Google Scholar 

  • Noel JP, Dixon RA, Pichersky E, Zubieta C, Ferrer J-L (2003) Structural, functional, and evolutionary basis for methylation of plant small molecules. Rec Adv Phytochemistry 37:37–58

    Article  CAS  Google Scholar 

  • Otwinowski Z, Minor W (1997) Processing of X-ray diffraction data collected in oscillation mode. In: Carter CWJ, Sweet RM (Eds) Methods in enzymology: macromolecular crystallography, part A., Vol 276. Academic Press, New York, pp 307–326

    Chapter  Google Scholar 

  • Paiva NL, Oommen A, Harrison MJ, Dixon RA (1994) Regulation of isoflavonoid metabolism in alfalfa. Plant Cell Tissue Organ Cult 38:213–220

    Article  CAS  Google Scholar 

  • Phillips DA, Kapulnik Y (1995) Plant isoflavonoids, pathogens and symbionts. Trends Microbiol 3:58–64

    Article  PubMed  CAS  Google Scholar 

  • Preisig CL, Matthews DE, VanEtten HD (1989) Purification and characterization of S-adenosyl-L-methionine:6a-hydroxymaackiain 3-O-methyltransferase from Pisum sativum. Plant Physiol 91:559–566

    Article  PubMed  CAS  Google Scholar 

  • Schroder G, Wehinger E, Lukacin R, Wellmann F, Seefelder W, Schwab W, Schroder J (2004) Flavonoid methylation: a novel 4′-O-methyltransferase from Catharanthus roseus, and evidence that partially methylated flavanones are substrates of four different flavonoid dioxygenases. Phytochemistry 65:1085–1094

    Article  PubMed  CAS  Google Scholar 

  • Schroder G, Wehinger E, Schroder J (2002) Predicting the substrates of cloned plant O-methyltransferases. Phytochemistry 59:1–8

    Article  PubMed  CAS  Google Scholar 

  • Suzuki H, Reddy MSS, Naoumkina M, Aziz N, May GD, Huhman DV, Sumner LW, Blount JW, Mendes P, Dixon RA (2005) Methyl jasmonate and yeast elicitor induce differential genetic and metabolic re-programming in cell suspension cultures of the model legume Medicago truncatula. Planta 220:698–707

    Article  CAS  Google Scholar 

  • Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680

    PubMed  CAS  Google Scholar 

  • VanEtten HD, Matthews DE, Smith DA (1982) Metabolism of phytoalexins. In: Bailey JA, Mansfield JW (Eds) Phytoalexins. Blackie, Glasgow, pp 180–217

    Google Scholar 

  • Wengenmayer H, Ebel J, Grisebach H (1974) Purification and properties of a S-adenosylmethionine: isoflavone 4′-O-methyltransferase from cell suspension cultures of Cicer arietinum L. Eur J Biochem 50:135–143

    Article  PubMed  CAS  Google Scholar 

  • Yang H, Ahn JH, Ibrahim RK, Lee S, Lim Y (2004) The three-dimensional structure of Arabidopsis thaliana O-methyltransferase predicted by homology-based modeling. J. Mol Graph Model 23:77–87

    Article  PubMed  CAS  Google Scholar 

  • Zubieta C, He X-Z, Dixon RA, Noel JP (2001) Structures of two natural product methyltransferases reveal the basis for substrate specificity in plant O-methyltransferases. Nature Struct Biol 8:271–279

    Article  PubMed  CAS  Google Scholar 

  • Zubieta C, Kota P, Ferrer J-L, Dixon RA, Noel J (2002) Structural basis for the modulation of lignin monomer methylation by caffeic acid/5-hydroxyferulic acid 3/5-O-methyltransferase. Plant Cell 14:1265–1277

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

We thank Drs Luis Marquez and Marilyn Roossinck for help with PAUP analysis, and Drs Xiaoqiang Wang and Luzia Modolo for critical reading of the manuscript. This work was supported by grants from the Oklahoma Center for the Advancement of Science and Technology Health Sciences Program to RAD, the National Science Foundation under Grant No. 0236027 to JPN and under Grant No. DBI 0109732 to RAD, and a Noble Foundation Postdoctoral Fellowship at the Salk Institute and a Laboratory Directed Research and Development Award (LDRD) at Brookhaven National Laboratory under contract with U.S. Department of Energy to C-JL. JPN is an investigator of the Howard Hughes Medical Institute.

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Author notes

  1. Bettina E. Deavours

    Present address: Department of Biology, Colorado State University, Fort Collins, CO, 80523, USA

  2. Mohamed A. Farag

    Present address: Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, Kasr El Aini St., Cairo, Egypt

Authors and Affiliations

  1. Plant Biology Division, Samuel Roberts Noble Foundation, 2510 Sam Noble Parkway, Ardmore, OK, 73401, USA

    Bettina E. Deavours, Marina A. Naoumkina, Yuhong Tang, Mohamed A. Farag, Lloyd W. Sumner & Richard A. Dixon

  2. Howard Hughes Medical Institute, Jack Skirball Chemical Biology and Proteomics Laboratory, The Salk Institute for Biological Studies, 10010 N. Torrey Pines Road, La Jolla, CA, 92036, USA

    Chang-Jun Liu & Joseph P. Noel

  3. Biology Department, Brookhaven National Laboratory, Upton, NY, 11973, USA

    Chang-Jun Liu

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  1. Bettina E. Deavours
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Correspondence to Richard A. Dixon.

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Deavours, B.E., Liu, CJ., Naoumkina, M.A. et al. Functional analysis of members of the isoflavone and isoflavanone O-methyltransferase enzyme families from the model legume Medicago truncatula . Plant Mol Biol 62, 715–733 (2006). https://doi.org/10.1007/s11103-006-9050-x

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  • DOI: https://doi.org/10.1007/s11103-006-9050-x

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