Abstract
The biosynthesis of flavonoids such as anthocyanin and stilbenes has attracted increasing attention because of their potential health benefits. Anthocyanins and stilbenes share common phenylpropanoid precursor pathways. We previously reported that the overexpression of sweetpotato IbMYB1a induced anthocyanin pigmentation in transgenic tobacco (Nicotiana tabacum) plants. In the present study, transgenic tobacco (Nicotiana tabacum SR1) plants (STS-OX and ROST-OX) expressing the RpSTS gene encoding stilbene synthase from rhubarb (Rheum palmatum L. cv. Jangyeop) and the RpSTS and VrROMT genes encoding resveratrol O-methyltransferase from frost grape (Vitis riparia) were generated under the control of 35S promoter. Phenotypic alterations in floral organs, such as a reduction in floral pigments and male sterility, were observed in STS-OX transgenic tobacco plants. However, we failed to obtain STS-OX and ROST-OX plants with high levels of resveratrol compounds. Therefore, to improve the production of resveratrol derivatives in plants, we cross-pollinated flowers of STS-OX or ROST-OX and IbMYB1a-OX transgenic lines (SM and RSM). Phenotypic changes in vegetative and reproductive development of SM and RSM plants were observed. Furthermore, by HPLC and LC-MS analyses, we found enhanced production of resveratrol derivatives such as piceid, piceid methyl ether, resveratrol methyl ether O-hexoside, and 5-methyl resveratrol-3,4′-O-β-d-diglucopyranoside in SM and RSM cross-pollinated lines. Here, total contents of trans- and cis-piceids ranged from approximately 104–240 µg/g fresh weight in SM (F2). Collectively, we suggest that coexpression of RpSTS and IbMYB1a via cross-pollination can induce enhanced production of resveratrol compounds in plants by increasing metabolic flux into stilbenoid biosynthesis.
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Acknowledgments
This work was supported by grants from the KRIBB Initiative Program and the Next-Generation BioGreen 21 Program (SSAC, Grant #: PJ01107004), Rural Development Administration, Republic of Korea.
Author contributions
C.Y. Kim and Y.J. Jeong: conceived and designed the experiments. S.G. Woo, C.H. An, J.H. Park and S.-H. Lee: performed the experiments. Y. Rim and H.J. Jeong: analyzed the data. Y.B. Ryu: contributed reagents/materials/analysis tools. C.Y. Kim and Y.J. Jeong: wrote the paper.
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Yu Jeong Jeong and Chul Han An have contributed equally to this work.
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11103_2016_497_MOESM2_ESM.tif
Supplementary Fig. S1 Expression analysis of RpSTS transgene in tobacco (Nicotiana benthamiana) leaves by transient Agro-infiltration assays. Agrobacterium [GV3101 (pSoup)] cells were infiltrated into the leaves of 4-week-old tobacco plants. Leaf discs were harvested at 3 and 5 days post-inoculation of Agrobacterium cells, immediately frozen in liquid nitrogen, and stored at −80C until use. Total proteins were isolated from tobacco leaf samples harvested after Agro-infiltrartion and analyzed by Western blot analysis with anti-Flag antibody. Con, empty vector as a control; STS, 35S-RpSTS; ROST, 35S-VrROMT-RpSTS; M, protein molecule marker; 3, 5; 3 and 5 days after Agro-infiltration. (TIF 2403 KB)
11103_2016_497_MOESM3_ESM.tif
Supplementary Fig. S2 Expression analysis of RpSTS transgene in transgenic tobacco (Nicotiana tabacum SR1) plants (T2 generation) expressing the RpSTS alone (STS-OX) or both the RpSTS and VrROMT (ROST-OX). Both transgenic lines of STS-OX and ROST-OX were screened by Flag-tagged RpSTS protein expression following Western blot analysis with anti-Flag antibody. (TIF 385 KB)
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Jeong, Y.J., An, C.H., Woo, S.G. et al. Enhanced production of resveratrol derivatives in tobacco plants by improving the metabolic flux of intermediates in the phenylpropanoid pathway. Plant Mol Biol 92, 117–129 (2016). https://doi.org/10.1007/s11103-016-0497-0
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DOI: https://doi.org/10.1007/s11103-016-0497-0