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The R2R3-MYB TT2b and the bHLH TT8 genes are the major regulators of proanthocyanidin biosynthesis in the leaves of Lotus species

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Abstract

Main conclusion

By exploiting interspecific hybrids and their progeny, we identified key regulatory and transporter genes intimately related to proanthocyanidin biosynthesis in leaves of Lotus spp.

Proanthocyanidins (PAs), known as condensed tannins, are polymeric flavonoids enriching forage legumes of key nutritional value to prevent bloating in ruminant animals. Unfortunately, major forage legumes such as alfalfa and clovers lack PAs in edible tissues. Therefore, engineering the PA trait in herbage of forage legumes is paramount to improve both ecological and economical sustainability of cattle production system. Progresses on the understanding of genetic determinants controlling PA biosynthesis and accumulation have been mainly made studying mutants of Arabidopsis, Medicago truncatula and Lotus japonicus, model species unable to synthesize PAs in the leaves. Here, we exploited interspecific hybrids between Lotus corniculatus, with high levels of PAs in the leaves, and Lotus tenuis, with no PAs in these organs, and relative F2 progeny, to identify among candidate PA regulators and transporters the genes mainly affecting this trait. We found that the levels of leaf PAs significantly correlate with the expression of MATE1, the putative transporter of glycosylated PA monomers, and, among the candidate regulatory genes, with the expression of the MYB genes TT2a, TT2b and MYB14 and the bHLH gene TT8. The expression levels of TT2b and TT8 also correlated with those of all key structural genes of the PA pathways investigated, MATE1 included. Our study unveils a different involvement of the three Lotus TT2 paralogs to the PA trait and highlights differences in the regulation of this trait in our Lotus genotypes with respect to model species. This information opens new avenues for breeding bloat safe forage legumes.

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Abbreviations

ANR :

Anthocyanidin reductase

ANS :

Anthocyanidin synthase

CHS :

Chalcone synthase

DFR :

Dihydroflavonol 4-reductase

DMACA:

4-Dimethylaminocinnamaldehyde

LAR :

Leucoanthocyanidin reductase

LDOX :

Leucoanthocyanidin synthase

PAL :

Phenylalanine ammonia-lyase

PAs:

Proanthocyanidins

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Acknowledgements

This work was supported by FONCYT-PICT 2011-1612, PICT 2014-3718 and 2014-3648 grants from the Agencia Nacional de Promoción Científica y Tecnológica (ANPCYT-Argentina); PIP 0980 grant from Consejo Nacional de Ciencia y Tecnologìa (CONICET-Argentina), CNR-CONICET bilateral project 2014–2016, and by Fondazione Cassa di Risparmio di Perugia grant n° 2015.0303 021. The authors are grateful to the anonymous reviewers for their insightful comments and suggestions.

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Authors and Affiliations

  1. Unidad de Biotecnología 1, Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico de Chascomús (IIB-INTECh) / Universidad Nacional de San Martín. Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Chascomús, Argentina

    Francisco José Escaray, Ana Perea-García, Cristian Javier Antonelli & Oscar Adolfo Ruiz

  2. Institute of Biosciences and BioResources (CNR-IBBR), Perugia, Italy

    Valentina Passeri, Francesco Damiani & Francesco Paolocci

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  1. Francisco José Escaray
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  2. Valentina Passeri
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  3. Ana Perea-García
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  4. Cristian Javier Antonelli
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  5. Francesco Damiani
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  6. Oscar Adolfo Ruiz
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  7. Francesco Paolocci
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Correspondence to Francesco Paolocci.

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Escaray, F.J., Passeri, V., Perea-García, A. et al. The R2R3-MYB TT2b and the bHLH TT8 genes are the major regulators of proanthocyanidin biosynthesis in the leaves of Lotus species. Planta 246, 243–261 (2017). https://doi.org/10.1007/s00425-017-2696-6

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