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Medicago glucosyltransferase UGT72L1: potential roles in proanthocyanidin biosynthesis

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Abstract

In the first reaction specific for proanthocyanidin (PA) biosynthesis in Arabidopsis thaliana and Medicago truncatula, anthocyanidin reductase (ANR) converts cyanidin to (−)-epicatechin. The glucosyltransferase UGT72L1 catalyzes formation of epicatechin 3′-O-glucoside (E3′OG), the preferred substrate for MATE transporters implicated in PA biosynthesis in both species. The mechanism of PA polymerization is still unclear, but may involve the laccase-like polyphenol oxidase TRANSPARENT TESTA 10 (TT10). We have employed a combination of cell biological, biochemical and genetic approaches to evaluate this PA pathway model. The promoter regions of UGT72L1 and MtANR share common cis-acting elements and direct overlapping, but partially distinct, expression patterns. UGT72L1 and MtANR are localized in the cytosol, whereas TT10 is localized to the vacuole. Over-expression of UGT72L1 in M. truncatula hairy roots results in increased accumulation of PA-like compounds, and loss of function of UGT72L1 partially reduces epicatechin, E3′OG and extractable PA levels in M. truncatula seeds. Expression of UGT72L1 in A. thaliana leads to a massive increase in E3′OG in immature seed, but reduced levels of extractable PAs. However, when UGT72L1 was expressed in the Arabidopsis tt10 mutant, extractable PA levels increased and seed coat browning was delayed. Our results suggest that glycosylation of epicatechin is important for both PA precursor transport and assembly, but that additional redundant pathways may exist.

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Abbreviations

ANR:

Anthocyanidin reductase

ANS:

Anthocyanidin synthase

DMACA:

Dimethylaminocinnamaldehyde

E3′OG:

Epicatechin 3′-O-glucoside

GFP:

Green fluorescent protein

GUS:

β-Glucuronidase

LAR:

Leucoanthocyanidin reductase

MATE:

Multidrug and toxic compound extrusion

NP-HPLC:

Normal-phase high-performance liquid chromatography

PA:

Proanthocyanidin

TT10:

Transparent testa 10

UGT:

Uridine diphosphate sugar glucosyltransferase

UPLC:

Ultra high-performance liquid chromatography

References

  • Abe M, Takahashi T, Komeda Y (2001) Identification of a cis-regulatory element for L1 layer-specific gene expression, which is targeted by an L1-specific homeodomain protein. Plant J 26:487–494

    Article  PubMed  CAS  Google Scholar 

  • Abrahams S, Tanner GJ, Larkin PJ, Ashton AR (2002) Identification and biochemical characterization of mutants in the proanthocyanidin pathway in Arabidopsis. Plant Physiol 130:561–576

    Article  PubMed  CAS  Google Scholar 

  • Abrahams S, Lee E, Walker AR, Tanner GJ, Larkin PJ, Ashton AR (2003) The Arabidopsis TDS4 gene encodes leucoanthocyanidin dioxygenase (LDOX) and is essential for proanthocyanidin synthesis and vacuole development. Plant J 35:624–636

    Article  PubMed  CAS  Google Scholar 

  • Cheng H, Li L, Cheng S, Cao F, Wang Y, Yuan H (2011) Molecular cloning and function assay of a chalcone isomerase gene (GbCHI) from Ginkgo biloba. Plant Cell Rep 30:49–62

    Article  PubMed  CAS  Google Scholar 

  • Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743

    Article  PubMed  CAS  Google Scholar 

  • Debeaujon I, Leon-Kloosterziel KM, Koornneef M (2000) Influence of the testa on seed dormancy, germination, and longevity in Arabidopsis. Plant Physiol 122:403–414

    Article  PubMed  CAS  Google Scholar 

  • Debeaujon I, Nesi N, Perez P, Devic M, Grandjean O, Caboche M, Lepiniec L (2003) Proanthocyanidin-accumulating cells in Arabidopsis testa: regulation of differentiation and role in seed development. Plant Cell 15:2514–2531

    Article  PubMed  CAS  Google Scholar 

  • Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: Version II. Plant Mol Biol Rep 1:19–21

    Article  CAS  Google Scholar 

  • Dixon RA, Xie DY, Sharma SB (2005) Proanthocyanidins: a final frontier in flavonoid research? New Phytol 165:9–28

    Article  PubMed  CAS  Google Scholar 

  • Fujita A, Soma N, Goto-Yamamoto N, Shindo H, Kakuta T, Koizumi T, Hashizume K (2005) Anthocyanidin reductase gene expression and accumulation of flavan-3-ols in grape berry. Am J Enol Viticult 56:336–342

    CAS  Google Scholar 

  • Hartmann U, Sagasser M, Mehrtens F, Stracke R, Weisshaar B (2005) Differential combinatorial interactions of cis-acting elements recognized by R2R3-MYB, BZIP, and BHLH factors control light-responsive and tissue-specific activation of phenylpropanoid biosynthesis genes. Plant Mol Biol 57:155–171

    Article  PubMed  CAS  Google Scholar 

  • Hartzfeld PW, Forkner R, Hunter MD, Hagerman AE (2002) Determination of hydrolyzable tannins (gallotannins and ellagitannins) after reaction with potassium iodate. J Agric Food Chem 50:1785–1790

    Article  PubMed  CAS  Google Scholar 

  • He F, Pan QH, Shi Y, Duan CQ (2008) Biosynthesis and genetic regulation of proanthocyanidins in plants. Molecules 13:2674–2703

    Article  PubMed  CAS  Google Scholar 

  • Hefner T, Arend J, Warzecha H, Siems K, Stockigt J (2002) Arbutin synthase, a novel member of the NRD1beta glycosyltransferase family, is a unique multifunctional enzyme converting various natural products and xenobiotics. Bioorg Med Chem 10:1731–1741

    Article  PubMed  CAS  Google Scholar 

  • Higo K, Ugawa Y, Iwamoto M, Korenaga T (1999) Plant cis-acting regulatory DNA elements (PLACE) database: 1999. Nucleic Acids Res 27:297–300

    Article  PubMed  CAS  Google Scholar 

  • Karimi M, Inze D, Depicker A (2002) GATEWAY vectors for Agrobacterium-mediated plant transformation. Trends Plant Sci 7:193–195

    Article  PubMed  CAS  Google Scholar 

  • Kennedy JA, Jones GP (2001) Analysis of proanthocyanidin cleavage products following acid-catalysis in the presence of excess phloroglucinol. J Agric Food Chem 49:1740–1746

    Article  PubMed  CAS  Google Scholar 

  • Kitamura S, Shikazono N, Tanaka A (2004) TRANSPARENT TESTA 19 is involved in the accumulation of both anthocyanins and proanthocyanidins in Arabidopsis. Plant J 37:104–114

    Article  PubMed  CAS  Google Scholar 

  • Kitamura S, Matsuda F, Tohge T, Yonekura-Sakakibara K, Yamazaki M, Saito K, Narumi I (2010) Metabolic profiling and cytological analysis of proanthocyanidins in immature seeds of Arabidopsis thaliana flavonoid accumulation mutants. Plant J 62:549–559

    Article  PubMed  CAS  Google Scholar 

  • Lepiniec L, Debeaujon I, Routaboul JM, Baudry A, Pourcel L, Nesi N, Caboche M (2006) Genetics and biochemistry of seed flavonoids. Annu Rev Plant Biol 57:405–430

    Article  PubMed  CAS  Google Scholar 

  • Limpens E, Ramos J, Franken C, Raz V, Compaan B, Franssen H, Bisseling T, Geurts R (2004) RNA interference in Agrobacterium rhizogenes-transformed roots of Arabidopsis and Medicago truncatula. J Exp Bot 55:983–992

    Article  PubMed  CAS  Google Scholar 

  • Logemann E, Parniske M, Hahlbrock K (1995) Modes of expression and common structural features of the complete phenylalanine ammonia-lyase gene family in parsley. Proc Natl Acad Sci USA 92:5905–5909

    Article  PubMed  CAS  Google Scholar 

  • Long JA, Moan EI, Medford JI, Barton MK (1996) A member of the KNOTTED class of homeodomain proteins encoded by the STM gene of Arabidopsis. Nature 379:66–69

    Article  PubMed  CAS  Google Scholar 

  • Marinova K, Pourcel L, Weder B, Schwarz M, Barron D, Routaboul JM, Debeaujon I, Klein M (2007) The Arabidopsis MATE transporter TT12 acts as a vacuolar flavonoid/H+-antiporter active in proanthocyanidin-accumulating cells of the seed coat. Plant Cell 19:2023–2038

    Article  PubMed  CAS  Google Scholar 

  • McKay DL, Blumberg JB (2002) The role of tea in human health: an update. J Am Coll Nutr 21:1–13

    Article  PubMed  CAS  Google Scholar 

  • Nassiri-Asl M, Hosseinzadeh H (2009) Review of the pharmacological effects of Vitis vinifera (grape) and its bioactive compounds. Phytother Res 23:1197–1204

    Article  PubMed  CAS  Google Scholar 

  • Nesi N, Jond C, Debeaujon I, Caboche M, Lepiniec L (2001) The Arabidopsis TT2 gene encodes an R2R3 MYB domain protein that acts as a key determinant for proanthocyanidin accumulation in developing seed. Plant Cell 13:2099–2114

    PubMed  CAS  Google Scholar 

  • Nonaka G (1989) Isolation and structure elucidation of tannins. Pure Appl Chem 61:357–360

    Article  CAS  Google Scholar 

  • Pang Y, Peel GJ, Wright E, Wang Z, Dixon RA (2007) Early steps in proanthocyanidin biosynthesis in the model legume Medicago truncatula. Plant Physiol 145:601–615

    Article  PubMed  CAS  Google Scholar 

  • Pang Y, Peel GJ, Sharma SB, Tang Y, Dixon RA (2008) A transcript profiling approach reveals an epicatechin-specific glucosyltransferase expressed in the seed coat of Medicago truncatula. Proc Natl Acad Sci USA 105:14210–14215

    Article  PubMed  CAS  Google Scholar 

  • Pang Y, Wenger JP, Saathoff K, Peel GJ, Wen J, Huhman D, Allen SN, Tang Y, Cheng X, Tadege M, Ratet P, Mysore KS, Sumner LW, Marks MD, Dixon RA (2009) A WD40 repeat protein from Medicago truncatula is necessary for tissue-specific anthocyanin and proanthocyanidin biosynthesis but not for trichome development. Plant Physiol 151:1114–1129

    Article  PubMed  CAS  Google Scholar 

  • Peel GJ, Dixon RA (2007) Detection and quantification of engineered proanthocyanidins in Transgenic plants. Nat Prod Commun 2:1009–1014

    CAS  Google Scholar 

  • Peng X, Ma J, Chao J, Sun Z, Chang RC, Tse I, Li ET, Chen F, Wang M (2010) Beneficial effects of cinnamon proanthocyanidins on the formation of specific advanced glycation endproducts and methylglyoxal-induced impairment on glucose consumption. J Agric Food Chem 58:6692–6696

    Article  PubMed  CAS  Google Scholar 

  • Pourcel L, Routaboul JM, Kerhoas L, Caboche M, Lepiniec L, Debeaujon I (2005) TRANSPARENT TESTA10 encodes a laccase-like enzyme involved in oxidative polymerization of flavonoids in Arabidopsis seed coat. Plant Cell 17:2966–2980

    Article  PubMed  CAS  Google Scholar 

  • Saito K, Kobayashi M, Gong Z, Tanaka Y, Yamazaki M (1999) Direct evidence for anthocyanidin synthase as a 2-oxoglutarate-dependent oxygenase: molecular cloning and functional expression of cDNA from a red forma of Perilla frutescens. Plant J 17:181–189

    Article  PubMed  Google Scholar 

  • Sharma SB, Dixon RA (2005) Metabolic engineering of proanthocyanidins by ectopic expression of transcription factors in Arabidopsis thaliana. Plant J 44:62–75

    Article  PubMed  CAS  Google Scholar 

  • Solano R, Nieto C, Avila J, Canas L, Diaz I, Paz-Ares J (1995) Dual DNA binding specificity of a petal epidermis-specific MYB transcription factor (MYB.Ph3) from Petunia hybrida. EMBO J 14:1773–1784

    PubMed  CAS  Google Scholar 

  • Sutoh K, Yamauchi D (2003) Two cis-acting elements necessary and sufficient for gibberellin-upregulated proteinase expression in rice seeds. Plant J 34:635–645

    Article  PubMed  CAS  Google Scholar 

  • Tadege M, Ratet P, Mysore KS (2005) Insertional mutagenesis: a Swiss Army knife for functional genomics of Medicago truncatula. Trends Plant Sci 10:229–235

    Article  PubMed  CAS  Google Scholar 

  • Tadege M, Wen J, He J, Tu H, Kwak Y, Eschstruth A, Cayrel A, Endre G, Zhao PX, Chabaud M, Ratet P, Mysore KS (2008) Large-scale insertional mutagenesis using the Tnt1 retrotransposon in the model legume Medicago truncatula. Plant J 54:335–347

    Article  PubMed  CAS  Google Scholar 

  • Tanner GJ, Francki KT, Abrahams S, Watson JM, Larkin PJ, Ashton AR (2003) Proanthocyanidin biosynthesis in plants. Purification of legume leucoanthocyanidin reductase and molecular cloning of its cDNA. J Biol Chem 278:31647–31656

    Article  PubMed  CAS  Google Scholar 

  • Wang HL, Grusak MA (2005) Structure and development of Medicago truncatula pod wall and seed coat. Ann Bot 95:737–747

    Article  PubMed  Google Scholar 

  • Wang S, Wang JW, Yu N, Li CH, Luo B, Gou JY, Wang LJ, Chen XY (2004) Control of plant trichome development by a cotton fiber MYB gene. Plant Cell 16:2323–2334

    Article  PubMed  CAS  Google Scholar 

  • Wang J, Ho L, Zhao W, Ono K, Rosensweig C, Chen L, Humala N, Teplow DB, Pasinetti GM (2008) Grape-derived polyphenolics prevent Abeta oligomerization and attenuate cognitive deterioration in a mouse model of Alzheimer’s disease. J Neurosci 28:6388–6392

    Article  PubMed  CAS  Google Scholar 

  • Winkel-Shirley B (2001) Flavonoid biosynthesis. A colorful model for genetics, biochemistry, cell biology, and biotechnology. Plant Physiol 126:485–493

    Article  PubMed  CAS  Google Scholar 

  • Xie DY, Sharma SB, Paiva NL, Ferreira D, Dixon RA (2003) Role of anthocyanidin reductase, encoded by BANYULS in plant flavonoid biosynthesis. Science 299:396–399

    Article  PubMed  CAS  Google Scholar 

  • Zhao J, Dixon RA (2009) MATE transporters facilitate vacuolar uptake of epicatechin 3′-O-glucoside for proanthocyanidin biosynthesis in Medicago truncatula and Arabidopsis. Plant Cell 21:2323–2340

    Article  PubMed  CAS  Google Scholar 

  • Zhao J, Dixon RA (2010) The ‘ins’ and ‘outs’ of flavonoid transport. Trends Plant Sci 15:72–80

    Article  PubMed  CAS  Google Scholar 

  • Zhao J, Pang Y, Dixon RA (2010) The mysteries of proanthocyanidin transport and polymerization. Plant Physiol 153:437–443

    Article  PubMed  CAS  Google Scholar 

  • Zhou R, Jackson L, Shadle G, Nakashima J, Temple S, Chen F, Dixon RA (2010) Distinct cinnamoyl CoA reductases involved in parallel routes to lignin in Medicago truncatula. Proc Natl Acad Sci USA 107:17803–17808

    Article  PubMed  CAS  Google Scholar 

  • Zhu Q, Ordiz MI, Dabi T, Beachy RN, Lamb C (2002) Rice TATA binding protein interacts functionally with transcription factor IIB and the RF2a bZIP transcriptional activator in an enhanced plant in vitro transcription system. Plant Cell 14:795–803

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank Drs. Jin Nakashima and Elison Blancaflor for assistance with microscopy, Drs Kiran Mysore, Million Tadege and Pascal Ratet for generation of the M. truncatula retrotransposon insertion population, and Drs JiHyung Jun and Stephen Temple for critical reading of the manuscript. The M. truncatula mutant plants utilized in this research project, which are jointly owned by the Centre National de la Recherche Scientifique, Gif-sur-Yvette, France and the Samuel Roberts Noble Foundation, Ardmore, OK, USA, were created through research funded, in part, by Grant # 703285 from the National Science Foundation. This work was supported by Forage Genetics International and the Samuel Roberts Noble Foundation.

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Author notes

  1. Yongzhen Pang

    Present address: Institute of Botany, The Chinese Academy of Sciences, Beijing, 100093, People’s Republic of China

  2. Gregory J. Peel

    Present address: Calgene/Monsanto, 1920 5th Street, Davis, CA, 95616, USA

  3. Richard A. Dixon

    Present address: Department of Biological Sciences, University of North Texas, 1155 Union Circle #305220, Denton, TX, 76203-5017, USA

Authors and Affiliations

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

    Yongzhen Pang, Xiaofei Cheng, David V. Huhman, Junying Ma, Gregory J. Peel, Guoan Shen, Lloyd W. Sumner, Yuhong Tang, Jianqi Wen, Jianfei Yun & Richard A. Dixon

  2. RIKEN Plant Science Center, 1-7-22, Suehiro, Tsurumi, Yokohama, 2300045, Japan

    Keiko Yonekura-Sakakibara & Kazuki Saito

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  1. Yongzhen Pang
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Correspondence to Richard A. Dixon.

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Pang, Y., Cheng, X., Huhman, D.V. et al. Medicago glucosyltransferase UGT72L1: potential roles in proanthocyanidin biosynthesis. Planta 238, 139–154 (2013). https://doi.org/10.1007/s00425-013-1879-z

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