Abstract
Plant secondary metabolites, such as stilbenes, have fungicidal potential and have been found in several plant species. Stilbenes in grapevine, such as resveratrol and pterostilbene, have recently attracted much attention, they are not only helping the plant to fight against pathogen attack, but they are also being widely used as ingredients of fungicide, anti-inflammatory drugs, antioxidant, and anti-infective agents. However, resveratrol O-methyltransferase gene, related with the synthesis of pterostilbene from resveratrol, has not been characterized effectively from Chinese wild Vitis pseudoreticulata. In this study, a candidate of resveratrol O-methyltransferase gene designated as VpROMT was isolated from a powdery mildew-resistant Chinese wild V. pseudoreticulata ‘Baihe-35-1’, and characterization studies were performed. Expression studies showed that VpROMT was predominantly expressed in developing roots yet not found in the leaves, stems, nor tendrils when the plants are not challenged. Results of qRT-PCR showed that VpROMT was rapidly induced by Erysiphe necator in V. pseudoreticulata and by methyl-jasmonate, UV-irradiation in suspension culture cells of Vitis romanetii. The expression level varies in different tissues of grapevine, which MeJA and UV-C treatment significantly upregulated the expression of VpROMT gene while UV-B treatment failed to. Co-expression of VpROMT and grapevine stilbene synthase (VpSTS) gene leads to the accumulation of pterostilbene in leaves of tobacco (Nicotiana tabacum) indicating that VpROMT was able to catalyze the biosynthesis of pterostilbene from resveratrol in over-expression transgenic tobacco plants.
Similar content being viewed by others
Abbreviations
- qRT-PCR:
-
Quantitative reverse transcriptase-polymerase chain reaction
- VpSTS:
-
Vitis pseudoreticulata stilbene synthase
- VpROMT:
-
Vitis pseudoreticulata resveratrol O-methyltransferase
- OMTs:
-
O-methyltransferases
- MeJA:
-
Methyl jasmonate
- RT-PCR:
-
Reverse transcriptase-polymerase chain reaction
- GUS:
-
β-Glucuronidase
- hpi:
-
Hours post inoculation
- Hyg:
-
Hygromycin B
- HPLC:
-
High performance liquid chromatography
- PM:
-
Powdery mildew
References
Adrian M, Jeandet P, Douillet-Breuil AC, Tesson L, Bessis R (2000) Stilbene content of mature Vitis vinifera berries in response to UV-C elicitation. J Agric Food Chem 48:6103–6105
Asif MH, Dhawan P, Nath P (2000) A simple procedure for the isolation of high quality RNA from ripening banana fruit. Plant Mol Biol Rep 18:105–119
Bernhard D, Tinhofer I, Tonko M, Hubl H, Ausserlechner MJ, Greil R, Kofler R, Csordas A (2000) Resveratrol causes arrest in the S-phase prior to Fas-independent apoptosis in CEM-C7H2 acute leukemia cells. Cell Death and Differentiation 7:834–842
Chiron H, Drouet A, Claudot AC, Eckerskorn C, Trost M, Heller W, Ernst D, Sandermann H (2000a) Molecular cloning and functional expression of a stress-induced multifunctional O-methyltransferase with pinosylvin methyltransferase activity from Scots pine (Pinus sylvestris L.). Plant Mol Biol 44:733–745
Chiron H, Drouet A, Lieutier F, Payer HD, Ernst D, Sandermann H (2000b) Gene induction of stilbene biosynthesis in Scots pine in response to ozone treatment, wounding, and fungal infection. Plant Physiol 124:865–872
Daley M, Knauf VC, Summerfelt KR, Turner JC (1998) Co-transformation with one Agrobacterium tumefaciens strain containing two binary plasmids as a method for producing marker-free transgenic plants. Plant cell Rep 17:489–496
De Block M, Debrouwer D (1991) Two T-DNAs co-transformed into Brassica napus by a double Agrobacterium tumefaciens infection are mainly integrated at the same locus. Theor Appl Genet 82:257–263
De Framond AJ, Back EW, Chilton WS, Kayes L, Chilton M (1986) Two unlinked T-DNAs can transform the same tobacco plant cell and segregate in the F1 generation. Mol Gen Genet 202:125–131
Depicker A, Herman L, Jacobs A, Schdl J, Montagu MV (1985) Frequencies of simultaneous transformation with different T-DNAs and their relevance to the Agrobacterium/plant cell interaction. Mol Gen Genet 201:477–484
Dixon RA, Paiva NL (1995) Stress-induced phenylpropanoid metabolism. Plant Cell 7:1085–1097
Dudareva N, Murfitt LM, Mann CJ, Gorenstein N, Kolosova N, Kish CM, Bonham C, Wood K (2000) Developmental regulation of methyl benzoate biosynthesis and emission in snap dragon flowers. Plant Cell 12:949–961
Fan C, Pu N, Wang X, Wang Y, Fang L, Xu W, Zhang J (2008) Agrobacterium-mediated genetic transformation of grapevine (Vitis vinifera L.) with a novel stilbene synthase gene from Chinese wild Vitis pseudoreticulata. Plant Cell Tissue Organ Cult 92:197–206
Fang Y, Smith MAL, Pepin MF (1999) Effects of exogenous methyl jasmonate in elicited anthocyanin-producing cell cultures of ohelo (Vaccinium pahalae). In Vitro Cell Dev Biol -Plant 35:106–113
Fritzmeier KH, Kindl H (1981) Coordinate induction by UV light of stilbene synthase, phenylalanine ammonia-lyase and cinnamate 4-hydroxylase in leaves of Vitaceae. Planta 151:48–52
Fulton TM, Chunzoongse J, Tanksley SD (1995) Microprep protocol for extraction of DNA from tomato and other herbaceous plants. Plant Mol Biol Rep 13(3):207–209
Fung RWM, Gonzalo M, Fekete C, Kovacs LG, He Y, Marsh E, McIntyre LM, Schachtman DP, Qiu W (2008) Powdery mildew induces defense-oriented reprogramming of the transcriptome in a susceptible but not in a resistant grapevine. Plant Physiol 146:236–249
Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements of suspension cultures of soybean root cells. Exp Cell Res 50:151–156
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
Gastaminza P, Whitten-Bauer C, Chisari FV (2010) Unbiased probing of the entire hepatitis C virus life cycle identifies clinical compounds that target multiple aspects of the infection. P Natl Acad Sci USA 107:291–296
Gauthier A, Gulick PJ, Ibrahim RK (1998) Characterization of two cDNA clones which encode O-methyltransferases for the methylation of both flavonoid and phenylpropanoid compounds. Arch Biochem Biophys 351:243–249
Gindro K, Spring JL, Pezet R, Richter H, Viret O (2006) Histological and biochemical criteria for objective and early selection of grapevine cultivars resistant to Plasmopara viticola. Vitis 45:191–196
Hart JH (1981) Role of phytostilbenes in decay and disease resistance. Ann Rev Phytopathol 19:437–458
He XZ, Dixon RA (1996) Affinity chromatography, substrate/product specificity, and amino acid sequence analysis of an isoflavone O-methyl-transferase from alfalfa (Medicago sativa L.). Arch Biochem Biophys 336:121–129
Horsch RB, Fry JE, Eichlotz D, Rogers SG, Frakey RT (1985) A simple and general method for transferring genes into plants. Sci 227:1229–1231
Ibrahim RK, De Luca V, Khouri HE, Latchinian L, Brisson L (1987) Enzymology and compartmentalization of polymethylated flavonol glucosides in Chrysosplenium americanum. Phytochem 26:1237–1245
Jang M, Cai L, Udeani GO et al (1997) Cancer chemopreventive activity of resveratrol, a natural product derived from grapes. Science 275:218–220
Jeandet P, Douillet-Breuil AC, Bessis R, Debord S, Sbaghi M, Adrian M (2002) Phytoalexins from the vitaceae: biosynthesis, phytoalexin gene expression in transgenic plants, antifungal activity, and metabolism. J Agr Food Chem 50(10):2731–2741
Kakegawa K, Kaneko Y, Hattori E, Koike K, Takeda K (1987) Cell cultures of Centaurea cyanus produce malonated anthocyanin in UV light. Phytochem 26:2261–2263
Kim BG, Lee Y, Hur HG, Lim Y, Ahn JH (2006) Flavonoid 3′-O-methyltransferase from rice: cDNA cloning, characterization and functional expression. Phytochem 67:387–394
Kim JS, Ha TY, Ahn J, Kim HK, Kim S (2009) Pterostilbene from Vitis coignetiae protect H2O2-induced inhibition of gap junctional intercellular communication in rat liver cell line. Food Chem Toxicol 47:404–409
Komari T, Hiei Y, Saito Y, Murai N (1996) Vectors carrying two separate T-DNAs for co-transformation of higher plants mediated by Agrobacterium tumefaciens and segregation of transformants free from selection markers. Plant J 10:165–174
Krisa S, Larronde F, Budzinski H, Decendit A, Deffieux G, Mérillon JM (1999) Stilbene production by Vitis vinifera cell suspension cultures: methyl jasmonate induction and 13C biolabeling. J Nat Prod 62:1688–1690
La P, Cai WQ, Zhang JC, Zhang FL, Fang RX (2001) Function of resveratrol derived from transgenic plant expressing resveratrol synthase gene. Chinese Sci Bull 46(13):1103–1107
Langcake P (1981) Disease resistance of Vitis spp. and the production of the stress metabolites resveratrol, epsilon-viniferin, alpha-viniferin and pterostilbene. Physiol Plant Pathol 18(2):213–226
Langcake P, Cornford CA, Pryce RJ (1979) Identification of pterostilbene as a phytoalexin from Vitis vinifera leaves. Phytochem 18:1025–1028
Lavid N, Wang J, ShalitM GI, Bar E, Beuerle T, Menda N, Shafir S, Zamir D, Adam Z, Vainstein A, Weiss D, Pichersky E, Lewinsohn E (2002) O-methyltransferases involved in the biosynthesis of volatile phenolic derivatives in rose petals. Plant Physiol 129:1899–1907
Manickam M, Ramanathan M, Jahromi MA, Chansouria JP, Ray AB (1997) Antihyperglycemic activity of phenolics from Pterocarpus marsupium. J Nat Prod 60:609–610
McKnight TD, Lillis MT, Simpson RB (1987) Segregation of genes transferred to one plant cell from two separate Agrobacterium strains. Plant Mol Biol 8:439–445
Mikstacka R, Przybylska D, Rimando AM, Baer-Dubowska W (2007) Inhibition of human recombinant cytochromes P450 CYP1A1 and CYP1B1 by trans-resveratrol methyl ethers. Mol Nutr Food Res 51:517–524
Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plantarum 15:473–497
Muzac I, Wang J, Anzellotti D, Zhang H, Ibrahim RK (2000) Functional expression of an Arabidopsis cDNA clone encoding a flavonol 3′-O-methyltransferase and characterization of the gene product. Arch Biochem Biophys 375:385–388
Pezet R, Pont V (1988) Identification of pterostilbene in grape berries of Vitis vinifera. Plant Physiol Biochem 26:603–607
Pezet R, Perret C, Jean-Denis JB, Tabacchi R, Gindro K, Viret O (2003) Delta-viniferin, a resveratrol dehydrodimer: one of the major stilbenes synthesized by stressed grapevine leaves. J Agr Food Chem 51:5488–5492
RamanaRao MV, Veluthambi K (2010) Selectable marker elimination in the T0 generation by Agrobacterium-mediated co-transformation involving Mungbean yellow mosaic virus TrAP as a non-conditional negative selectable marker and bar for transient positive selection. Plant cell Rep 29:473–483
Remsberg CM, Yanez JA, Ohgami Y, Vega-Villa KR, Rimando AM, Davies NM (2008) Pharmacometrics of pterostilbene: preclinical pharmacokinetics and metabolism, anticancer, anti-inflammatory, antioxidant and analgesic activity. Phytother Res 22:169–179
Rimando AM, Cuendet M, Desmarchelier C, Mehta RG, Pezzuto JM, Duke SO (2002) Cancer chemo-preventive and antioxidant activities of pterostilbene, a naturally occurring analogue of resveratrol. J Agr Food Chem 50:3453–3457
Rimando AM, Kalt W, Magee JB, Dewey J, Ballington JR (2004) Resveratrol, pterostilbene, and piceatannol in vaccinium berries. J Agr Food Chem 52:4713–4719
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, New York
Saniewski M, Miszczak A, Kawa-Misczak L, Wegrzynowicz-Lesiak E, Miyamoto K et al (1998) Effects of methyl jasmonate on anthocyanin accumulation, ethylene production, and CO2 evolution in uncooled and cooled tulip bulbs. J Plant Growth Regul 17:33–37
Satheesh A, Pari L (2006) The antioxidant role of pterostilbene in streptozotocin nicotinamide-induced type 2 diabetes mellitus in Wistar rats. J Pharm Pharmacol 58:1483–1490
Sato K, Nakayama M, Shigeta J (1996) Culturing conditions affecting the production of anthocyanin in suspended cell cultures of strawberry. Plant Sci 113:91–98
Scalliet G, Journot N, Jullien F, Baudino S, Magnard JL, Channelière S, Vergne P, Dumas C, Bendahmane M, Cock JM, Hugueney P (2002) Biosynthesis of the major scent components 3,5-dimethoxytoluene and 1,3,5-trimethoxy-benzene by novel rose O-methyltransferases. FEBS Lett 523:113–118
Schmidlin L, Poutararaud A, Claudel P, Metre P, Prado E, Santos-Rosa M, Wiedemann-Merdinoglu S, Karst F, Merdinoglu D, Hugueney P (2008) A stress-inducible resveratrol O-methyltransferase involved in the biosynthesis of pterostilbene in grapevine. Plant Physiol 148:1630–1639
Schnee S, Viret O, Gindro K (2008) Role of stilbenes in the resistance of grapevine to powdery mildew. Physiol Mol Plant P 72:128–133
Schroder G, Wehingera E, Lukacin R, Wellmann F, Seefelder W, Schwab W, Schröder 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. Phytochem 65:1085–1094
Seshadri TR (1972) Polyphenols of Pterocarpus and Dalbergia woods. Phytochem 11:881–898
Sripriya R, Raghupathy V, Veluthambi K (2008) Generation of selectable marker-free sheath blight resistant transgenic rice plants by efficient co-transformation of a cointegrate vector T-DNA and a binary vector T-DNA in one Agrobacterium tumefaciens strain. Plant Cell Rep 27:1635–1644
Takeda J (1990) Light-induced synthesis of anthocyanin in carrot cells in suspension. II Effects of light and 2,4-D on induction and reduction of enzyme activities related to anthocyanin synthesis. J Exp Bot 41:749–755
Vignols F, Rigau J, Torres MA, Capeliades M, Puigdomenech P (1995) The brown midrib3 (bm3) mutation in maize occurs in the gene encoding caffeic acid O-methyltransferase. Plant Cell 7:407–416
Wang J, Pichersky E (1999) Identification of specific residues involved in substrate discrimination in two plant O-methyltransferases. Arch Biochem Biophys 368:172–180
Wang YJ, Liu Y, He P, Chen J, Lamicanra O, Lu J (1995) Evaluation of foliar resistance to Uncinula necator in Chinese wild Vitis species. Vitis 34:159–164
Wang QF, Ma LY, Huang MJ, Wang L, Luo K, Li JM (2008) Determination of five stilbenes composition in grape leaves by high performance liquid chromatography. Chinese J Anal Chem 36(10):1359–1363
Wu S, Watanabe N, Mita S, Dohra H, Ueda Y, Shibuya M, Ebizuka Y (2004) The key role of phloroglucinol O-methyltransferase in the biosynthesis of Rosa chinensis volatile 1,3,5-trimethoxybenzene. Plant Physiol 135:1–8
Xu W, Wang Y, Wang X, Hao W, Sun M (2005) Construction of the plant expression vectors carrying resistant genes to powdery mildew and adversities in wild species of Vitis in China. Acta Bot Boreal-Occident Sin 25(5):851–857
Xu Y, Zhu Z, Xiao Y, Wang Y (2009) Construction of a cDNA Library of Vitis pseudoreticulata native to China inoculated with Uncinula necator and the analysis of potential defence-related expressed sequence tags (ESTs). S Afr J Enol Viticult 30:65–71
Xu W, Yu Y, Ding J, Hua Z, Wang Y (2010) Characterization of a novel stilbene synthase promoter involved in pathogen- and stress-inducible expression from Chinese wild Vitis pseudoreticulata. Planta 231:475–487
Yu CKY, Springob K, Schmidt J, Nicholson RL, Chu IK, Yip WK, Lo C (2005) A stilbene synthase gene (SbSTS1) is involved in host and non-host defense in sorghum. Plant Physiol 138:3
Zhang W, Curtin C, Kikuchi M, Franco C (2002) Integration of jasmonic acid and light irradiation for enhancement of anthocyanin biosynthesis in Vitis inifera suspension cultures. Plant Sci 162:459–468
Zhang JJ, Wang YJ, Wang XP (2003) An improved method for rapidly extracting total RNA from Vitis. J Fruit Sci 20:178–181
Zhong JJ, Yoshida M, Fujiyama K, Seki T, Yoshida T (1993) Enhancement of anthocyanin production by Perilla frutescens cells in a stirred bioreactor with internal light irradiation. J Ferment Bioeng 75:299–303
Acknowledgments
We express our gratitude to the National Natural Science Foundation of China (Grant no. 30940051) for generous financial support of this work, Yan Xu is supported by the program New Century Excellent Talents in University (NCET-10-0692), and the National Public Benefit (Agricultural) Research Foundation of China (200903044).
Conflict of interest
The authors declare that they have no conflict of interest.
Author information
Authors and Affiliations
Corresponding authors
Additional information
Handling Editor: Peter Nick
Y. Xu and T.F. Xu contributed equally to this work.
Rights and permissions
About this article
Cite this article
Xu, Y., Xu, T.F., Zhao, X.C. et al. Co-expression of VpROMT gene from Chinese wild Vitis pseudoreticulata with VpSTS in tobacco plants and its effects on the accumulation of pterostilbene. Protoplasma 249, 819–833 (2012). https://doi.org/10.1007/s00709-011-0335-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00709-011-0335-9