Skip to main content

Advertisement

Log in

Transgenic mimicry of pathogen attack stimulates growth and secondary metabolite accumulation

  • Original Paper
  • Published:
Transgenic Research Aims and scope Submit manuscript

Abstract

Plant secondary metabolites, including pharmaceuticals, flavorings and aromas, are often produced in response to stress. We used chemical inducers of the pathogen defense response (jasmonic acid, salicylate, killed fungi, oligosaccharides and the fungal elicitor protein, cryptogein) to increase metabolite and biomass production in transformed root cultures of the medicinal plant, Withania somnifera, and the weed, Convolvulus sepium. In an effort to genetically mimic the observed effects of cryptogein, we employed Agrobacterium rhizogenes to insert a synthetic gene encoding cryptogein into the roots of C. sepium, W. somnifera and Tylophora tanakae. This genetic transformation was associated with stimulation in both secondary metabolite production and growth in the first two species, and in growth in the third. In whole plants of Convolvulus arvensis and Arabidopsis thaliana, transformation with the cryptogein gene led, respectively, to increases in the calystegines and certain flavonoids. A similar transgenic mimicry of pathogen attack was previously employed to stimulate resistance to the pathogen and abiotic stress. In the present study of biochemical phenotype, we show that transgenic mimicry is correlated with increased secondary metabolite production in transformed root cultures and whole plants. We propose that natural transformation with genes encoding the production of microbial elicitors could influence interactions between plants and other organisms.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Ashtamker C, Kiss V, Sagi M, Davydov O, Fluhr R (2007) Diverse subcellular locations of cryptogein-induced ROS production in tobacco BY-2 cells. Plant Physiol 143(4):1817–1826

    Article  PubMed  CAS  Google Scholar 

  • Bandyopadhyay M, Jha S, Tepfer D (2007) Changes in morphological phenotypes and withanolide composition of Ri-transformed roots of Withania somnifera. Plant Cell Rep 26:599–609. doi:10.1007/s00299-006-0260-0

    Article  PubMed  CAS  Google Scholar 

  • Bechtold N, Ellis J, Pelletier G (1993) In planta Agrobacterium mediated gene transfer by infiltration of adult Arabidopsis thaliana plants. C R Acad Sci 316(3):1194–1199

    CAS  Google Scholar 

  • Bevan M (1984) Binary Agrobacterium vectors for plant transformation. Nucleic Acids Res 12:8711–8721. doi:10.1093/nar/12.22.8711

    Article  PubMed  CAS  Google Scholar 

  • Boissy G, de La Fortelle E, Kahn R, Huet JC, Bricogne G, Pernollet JC et al (1996) Crystal structure of a fungal elicitor secreted by Phytophthora cryptogea, a member of a novel class of plant necrotic proteins. Structure 4:1429–1439. doi:10.1016/S0969-2126(96)00150-5

    Article  PubMed  CAS  Google Scholar 

  • Buter B, Orlacchio C, Soldati A, Berger K (1998) Significance of genetic and environmental aspects in the field cultivation of Hypericum perforatum. Planta Med 64:431–437. doi:10.1055/s-2006-957475

    Article  PubMed  CAS  Google Scholar 

  • Chaudhuri KN, Ghosh B, Tepfer D, Jha S (2005) Genetic transformation of Tylophora indica with Agrobacterium rhizogenes A4: growth and tylophorine productivity in different transformed root clones. Plant Cell Rep 24:25–35. doi:10.1007/s00299-004-0904-x

    Article  PubMed  CAS  Google Scholar 

  • Coïc Y, Lesaint C (1973) La nutrition minérale en horticulture avancée. Rev Hortic Paris 2816:29–34

    Google Scholar 

  • Dellaporta S, Wood J, Hicks J (1983) A plant DNA minipreparation: version II. Plant Mol Biol Rep 1:19–21. doi:10.1007/BF02712670

    Article  CAS  Google Scholar 

  • Desfeux C, Clough S, Bent A (2000) Female reproductive tissues are the primary target of Agrobacterium-mediated transformation by the Arabidopsis floral-dip method. Plant Physiol 123:895–904. doi:10.1104/pp. 123.3.895

    Article  PubMed  CAS  Google Scholar 

  • Dueckershoff K, Unger M, Frank A, Gillam EM, Guengerich FP, Warzecha H (2005) Modified nicotine metabolism in transgenic tobacco plants expressing the human cytochrome P450 2A6 cDNA. FEBS Lett 579:2480–2484. doi:10.1016/j.febslet.2005.02.082

    Article  PubMed  CAS  Google Scholar 

  • Erwin DC, Ribeiro OK (1966) Phytophthora diseases worldwide. American Phytopathological Society Press, St. Paul

    Google Scholar 

  • Flores H, Hoy M, Pickard J (1987) Secondary metabolites from root cultures. Trends Biotechnol 5:64–69. doi:10.1016/S0167-7799(87)80013-6

    Article  CAS  Google Scholar 

  • Foissner I, Wendehenne D, Langebartels C, Durner J (2000) In vivo imaging of an elicitor-induced nitric oxide burst in tobacco. Plant J 23:817–824. doi:10.1046/j.1365-313X.2000.00835.x

    Article  PubMed  CAS  Google Scholar 

  • Glawischnig E (2007) Camalexin. Phytochemistry 68:401–406. doi:10.1016/j.phytochem.2006.12.005

    Article  PubMed  CAS  Google Scholar 

  • Goldmann A, Milat ML, Ducrot PH, Lallemand JY, Maille M, Lepingle A et al (1990) Tropane derivatives from Calystegia sepium. Phytochemistry 29:2125–2127. doi:10.1016/0031-9422(90)83019-W

    Article  CAS  Google Scholar 

  • Goldmann C, Message B, Lecoeur L, Delarue M, Maille M, Tepfer D (1993) Metabolic signals in the rhizosphere: catabolism of calystegins and betaines by Rhizobium meliloti. In: van Beek T, Breteler H (eds) Phytochemistry and agriculture. Oxford University Press, Oxford

    Google Scholar 

  • Goldmann A, Message B, Tepfer D, Molyneux R, Duclos O, Boyer F-D et al (1996) Biological activities of the nortropane alkaloid, calystegine B2, and analogs: structure–function relationships. J Nat Prod 59:1137–1142. doi:10.1021/np960409v

    Article  PubMed  CAS  Google Scholar 

  • Goossens A, Hakkinen ST, Laakso I, Seppanen-Laakso T, Biondi S, De Sutter V et al (2003) A functional genomics approach toward the understanding of secondary metabolism in plant cells. Proc Natl Acad Sci USA 100:8595–8600. doi:10.1073/pnas.1032967100

    Article  PubMed  CAS  Google Scholar 

  • Guillon S, Tremouillaux-Guiller J, Pati PK, Rideau M, Gantet P (2006) Hairy root research: recent scenario and exciting prospects. Curr Opin Plant Biol 9:341–346. doi:10.1016/j.pbi.2006.03.008

    Article  PubMed  CAS  Google Scholar 

  • Guo ZJ, Chen XJ, Wu XL, Ling JQ, Xu P (2004) Overexpression of the AP2/EREBP transcription factor OPBP1 enhances disease resistance and salt tolerance in tobacco. Plant Mol Biol 55:607–618. doi:10.1007/s11103-004-1521-3

    Article  PubMed  CAS  Google Scholar 

  • Harborne JB, Williams CA (2000) Advances in flavonoid research since 1992. Phytochemistry 55:481–504. doi:10.1016/S0031-9422(00)00235-1

    Article  PubMed  CAS  Google Scholar 

  • Holmberg N (1997) Stragedies for the production of secondary metabolites by pri-transformed regenerants. Plant Tissue Cult Biotechnol 3:128–137

    Google Scholar 

  • Inui T, Tamura K, Fujii N, Morishige T, Sato F (2007) Overexpression of Coptis japonica norcoclaurine 6-O-methyltransferase overcomes the rate-limiting step in Benzylisoquinoline alkaloid biosynthesis in cultured Eschscholzia californica. Plant Cell Physiol 48:252–262. doi:10.1093/pcp/pcl062

    Article  PubMed  CAS  Google Scholar 

  • Jiang DH, Chen XJ, Wu KL, Guo ZJ (2002) Site-directed mutagenesis of cryptogein gene (CryK13V) and generation of transgenic tobacco plants with nonspecific disease resistance. J Plant Physiol Mol Biol 28:399–406

    CAS  Google Scholar 

  • Jiang D, Chen X, Wu K, Guo Z (2004) Expression of cryptogein in tobacco plants exhibits enhanced disease resistance and tolerance to salt stress. Chin Sci Bull 49:803–809

    CAS  Google Scholar 

  • Jung G, Tepfer D (1987) Use of genetic transformation by the Ri T-DNA of Agrobacterium rhizogenes to stimulate biomass and tropane alkaloid production in Atropa belladonna and Calystegia sepium roots. Plant Sci 50:145–151. doi:10.1016/0168-9452(87)90151-8

    Article  CAS  Google Scholar 

  • Kasparovsky T, Blein J-P, Mikes V (2004) Ergosterol elicits oxidative burst in tobacco cells via phospholipase A2 and protein kinase C signal pathway. Plant Physiol Biochem 42:429–435. doi:10.1016/j.plaphy.2004.04.003

    Article  PubMed  CAS  Google Scholar 

  • Keller H, Pamboukdjian N, Ponchet M, Poupet A, Delon R, Verrier J et al (1999) Pathogen-induced elicitin production in transgenic tobacco generates a hypersensitive response and nonspecific disease resistance. Plant Cell 11:223–236

    Article  PubMed  CAS  Google Scholar 

  • Lamotte O, Gould K, Lecourieux D, Sequeira-Legrand A, Lebrun-Garcia A, Durner J et al (2004) Analysis of nitric oxide signaling functions in tobacco cells challenged by the elicitor cryptogein. Plant Physiol 135:516–529. doi:10.1104/pp.104.038968

    Article  PubMed  CAS  Google Scholar 

  • Lecourieux D, Mazars C, Pauly N, Ranjeva R, Pugin A (2002) Analysis and effects of cytosolic free calcium increases in response to elicitors in Nicotiana plumbaginifolia cells. Plant Cell 14:2627–2641. doi:10.1105/tpc.005579

    Article  PubMed  CAS  Google Scholar 

  • Li J, Ou-Lee TM, Raba R, Amundson RG, Last RL (1993) Arabidopsis flavonoid mutants are hypersensitive to UV-B irradiation. Plant Cell 5:171–179

    Article  PubMed  CAS  Google Scholar 

  • Lin L, Wu J (2002) Enhancement of shikonin production in single- and two-phase suspension cultures of Lithospermum erythrorhizon cells using low-energy ultrasound. Biotechnol Bioeng 78:81–88. doi:10.1002/bit.10180

    Article  PubMed  CAS  Google Scholar 

  • Liu R, Hu Y, Li J, Lin Z (2007) Production of soybean isoflavone genistein in non-legume plants via genetically modified secondary metabolism pathway. Metab Eng 9:1–7. doi:10.1016/j.ymben.2006.08.003

    Article  PubMed  Google Scholar 

  • Lochman J, Kasparovsky T, Damborsky J, Osman H, Marais A, Chaloupkova R et al (2005) Construction of cryptogein mutants, a proteinaceous elicitor from Phytophthora, with altered abilities to induce a defense reaction in tobacco cells. Biochemistry 44:6565–6572. doi:10.1021/bi0502285

    Article  PubMed  CAS  Google Scholar 

  • Long M, Millar DJ, Kimura Y, Donovan G, Rees J, Fraser PD et al (2006) Metabolite profiling of carotenoid and phenolic pathways in mutant and transgenic lines of tomato: identification of a high antioxidant fruit line. Phytochemistry 67:1750–1757. doi:10.1016/j.phytochem.2006.02.022

    Article  PubMed  CAS  Google Scholar 

  • Matthews D, Jones H, Gans P, Coates S, Smith LM (2005) Toxic secondary metabolite production in genetically modified potatoes in response to stress. J Agric Food Chem 53:7766–7776. doi:10.1021/jf050589r

    Article  PubMed  CAS  Google Scholar 

  • Mikes V, Milat M-L, Ponchet M, Ricci P, Blein J-P (1997) The fungal elicitor cryptogein is a sterol carrier protein. FEBS Lett 416:190–192. doi:10.1016/S0014-5793(97)01193-9

    Article  PubMed  CAS  Google Scholar 

  • Molyneux R, Pan Y, Goldmann A, Tepfer D, Elbein A (1993) Calystegins, a novel class of alkaloid glycosidase inhibitors. Arch Biochem Biophys 304:81–88. doi:10.1006/abbi.1993.1324

    Article  PubMed  CAS  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497. doi:10.1111/j.1399-3054.1962.tb08052.x

    Article  CAS  Google Scholar 

  • O’Donohue M, Gousseau H, Huet J-C, Tepfer D, Pernollet J-C (1995) Chemical synthesis, expression and mutagenesis of a gene encoding β-cryptogein, an elicitin produced by Phytophthora cryptogea. Plant Mol Biol 27:577–586. doi:10.1007/BF00019323

    Article  PubMed  CAS  Google Scholar 

  • O’Donohue MJ, Boissy G, Huet J-C, Nespoulous C, Brunie S, Pernollet J-C (1996) Overexpression in Phichia pastoris and crystallization of an elicitor protein secreted by the phytopathogenic fungus, Phytopthora cryptogea. Protein Expr Purif 8:254–261. doi:10.1006/prep.1996.0098

    Article  PubMed  CAS  Google Scholar 

  • Olsen CS (2005) Valuation of commercial central Himalayan medicinal plants. Ambio 34:607–610. doi:10.1639/0044-7447(2005)034[0607:VOCCHM]2.0.CO;2

    Article  PubMed  Google Scholar 

  • Pietta PG (2000) Flavonoids as antioxidants. J Nat Prod 63:1035–1042. doi:10.1021/np9904509

    Article  PubMed  CAS  Google Scholar 

  • Planchet E, Sonoda M, Zeier J, Kaiser WM (2006) Nitric oxide (NO) as an intermediate in the cryptogein-induced hypersensitive response—a critical re-evaluation. Plant Cell Environ 29:59–69. doi:10.1111/j.1365-3040.2005.01400.x

    Article  PubMed  CAS  Google Scholar 

  • Ren D, Yang KY, Li GJ, Liu Y, Zhang S (2006) Activation of Ntf4, a tobacco MAPK, during plant defense response and its involvement in hypersensitive response-like cell death. Plant Physiol 141(4):1482–1493

    Article  PubMed  CAS  Google Scholar 

  • Ricci P, Bonnnet P, Huet JC, Sallantin M, Beauvais-Cante F, Bruneteau M et al (1989) Structure and activity of proteins from pathogenic fungi Phytophthora eliciting necrosis and acquired resistance in tobacco. Eur J Biochem 183:555–563. doi:10.1111/j.1432-1033.1989.tb21084.x

    Article  PubMed  CAS  Google Scholar 

  • Routaboul J-M, Kerhoas L, Debeaujon I, Pourcel L, Caboche M, Einhorn J et al (2006) Flavonoid diversity and biosynthesis in seed of Arabidopsis thaliana. Planta 224:96–107. doi:10.1007/s00425-005-0197-5

    Article  PubMed  CAS  Google Scholar 

  • Shi M, Kwok KW, Wu JY (2007) Enhancement of tanshinone production in Salvia miltiorrhiza Bunge (red or Chinese sage) hairy-root culture by hyperosmotic stress and yeast elicitor. Biotechnol Appl Biochem 46:191–196. doi:10.1042/BA20060147

    Article  PubMed  CAS  Google Scholar 

  • Shinwari ZK, Gilani SS (2003) Sustainable harvest of medicinal plants at Bulashbar Nullah, Astore (Northern Pakistan). J Ethnopharmacol 84:289–298. doi:10.1016/S0378-8741(02)00333-1

    Article  PubMed  Google Scholar 

  • Slightom J, Durand-Tardif M, Jouanin L, Tepfer D (1986) Nucleotide sequence analysis of TL-DNA of Agrobacterium rhizogenes agropine type plasmid. J Biol Chem 261:108–121

    PubMed  CAS  Google Scholar 

  • Spiro MD, Kates KA, Koller AL, O’Neill MA, Albersheim P, Darvill AG (1993) Purification and characterization of biologically active 1,4-linked α-d-oligogalacturonides after partial digestion of polygalacturonic acid with endopolygalacturonase. Carbohydr Res 247:9–20. doi:10.1016/0008-6215(93)84237-Z

    Article  CAS  Google Scholar 

  • Tepfer D (1983a) The biology of genetic transformation of higher plants by Agrobacterium rhizogenes. In: Puhler A (ed) Molecular genetics of the bacteria plant interaction. Springer Verlag, Berlin

    Google Scholar 

  • Tepfer D (1983b) The potential uses of Agrobacterium rhizogenes in the genetic engineering of higher plants: nature got there first. In: Lurquin P, Kleinhofs A (eds) Genetic engineering in eukaryotes. Plenum Press, New York

    Google Scholar 

  • Tepfer D (1984) Transformation of several species of higher plants by Agrobacterium rhizogenes: sexual transmission of the transformed genotype and phenotype. Cell 37:959–967. doi:10.1016/0092-8674(84)90430-6

    Article  PubMed  CAS  Google Scholar 

  • Tepfer D (1989) Ri T-DNA from Agrobacterium rhizogenes: a source of genes having applications in rhizosphere biology and plant development, ecology, and evolution. In: Kosuge T, Nester E (eds) Plant–microbe interactions. McGraw Hill, New York

    Google Scholar 

  • Tepfer D, Tempé J (1981) Production d’agropine par des racines formées sous l’action d’Agrobacterium rhizogenes, souche A4. C R Acad Sci 292:153–156

    CAS  Google Scholar 

  • Tepfer D, Goldmann A, Fleury V, Maille M, Message B, Pamboukdjian N et al (1988a) Calystegins, nutritional mediators in plant-microbe interactions. In: Palacios R, Verma D (eds) Molecular genetics of plant–microbe interactions. APS Press, St Paul

    Google Scholar 

  • Tepfer D, Goldmann A, Pamboukdjian N, Maille M, Lépingle A, Chevalier D et al (1988b) A plasmid of Rhizobium meliloti 41 encodes catabolism of two compounds from root exudate of Calystegia sepium. J Bacteriol 170:1153–1161

    PubMed  CAS  Google Scholar 

  • Tepfer D, Metzger L, Prost R (1989) Use of roots transformed by Agrobacterium rhizogenes in rhizosphere research: applications in studies of cadmium assimilation from sewage sludges. Plant Mol Biol 13:295–302. doi:10.1007/BF00025317

    Article  PubMed  CAS  Google Scholar 

  • Tepfer D, Boutteaux C, Vigon C, Aymes S, Perez V, O’Donohue MJ, Huet J-C, Pernollet J-C (1998) Phytophtora resistance through production of a fungal protein elicitor (B-cryptogein) in tobacco. Mol Plant Microbe Interact 11(1):64–67

    Article  CAS  Google Scholar 

  • Tian L, Dixon RA (2006) Engineering isoflavone metabolism with an artificial bifunctional enzyme. Planta 224:496–507. doi:10.1007/s00425-006-0233-0

    Article  PubMed  CAS  Google Scholar 

  • de Vries J, Wackernagel W (1998) Detection of nptll (kanamycin resistance) genes in genomes of transgenic plants by marker-rescue transformation. Mol Gen Genet 257:606–613. doi:10.1007/s004380050688

    Article  PubMed  Google Scholar 

  • Wolucka BA, Goossens A, Inze D (2005) Methyl jasmonate stimulates the de novo biosynthesis of vitamin C in plant cell suspensions. J Exp Bot 56:2527–2538. doi:10.1093/jxb/eri246

    Article  PubMed  CAS  Google Scholar 

  • Ye H, Huang LL, Chen SD, Zhong JJ (2004) Pulsed electric field stimulates plant secondary metabolism in suspension cultures of Taxus chinensis. Biotechnol Bioeng 88:788–795. doi:10.1002/bit.20266

    Article  PubMed  CAS  Google Scholar 

  • Zhang C, Fevereiro PS (2007) The effect of heat shock on paclitaxel production in Taxus yunnanensis cell suspension cultures: role of abscisic acid pretreatment. Biotechnol Bioeng 96:506–514. doi:10.1002/bit.21122

    Article  PubMed  CAS  Google Scholar 

  • Zhao J, Davis LC, Verpoorte R (2005) Elicitor signal transduction leading to production of plant secondary metabolites. Biotechnol Adv 23:283–333. doi:10.1016/j.biotechadv.2005.01.003

    Article  PubMed  CAS  Google Scholar 

  • Zobayed SM, Afreen F, Kozai T (2005) Temperature stress can alter the photosynthetic efficiency and secondary metabolite concentrations in St. John’s wort. Plant Physiol Biochem 43:977–984. doi:10.1016/j.plaphy.2005.07.013

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

We thank Lucien Kerhoas for the LC-MS and Brigitte Message for general technical support. Aziz Aziz generously provided the oligogalacturonides, and Michael O’Donohue and Jean-Claude Pernollet donated the purified cryptogein protein. Seeds of T. tanakae and purified tylophorine were a kind gift from Fumiko Abe. This research was made possible by the Centre Franco-Indien pour la Promotion de la Recherche Avancée (Project 2503-2).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to David Tepfer.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chaudhuri, K., Das, S., Bandyopadhyay, M. et al. Transgenic mimicry of pathogen attack stimulates growth and secondary metabolite accumulation. Transgenic Res 18, 121–134 (2009). https://doi.org/10.1007/s11248-008-9201-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11248-008-9201-8

Keywords

Navigation