Proanthocyanidin subunit composition determined by functionally diverged dioxygenases

Abstract

Proanthocyanidins (PAs) are primarily composed of the flavan-3-ol subunits (-)-epicatechin and/or (+)-catechin, but the basis for their different starter and extension unit compositions remains unclear. Genetic and biochemical analyses show that, in the model legume Medicago truncatula, two 2-oxoglutarate-dependent dioxygenases, anthocyanidin synthase (ANS) and its homologue leucoanthocyanidin dioxygenase (LDOX), are involved in parallel pathways to generate, respectively, the (-)-epicatechin extension and starter units of PAs, with (+) catechin being an intermediate in the formation of the (-)-epicatechin starter unit. The presence/absence of the LDOX pathway accounts for natural differences in PA compositions across species, and engineering loss of function of ANS or LDOX provides a means to obtain PAs with different compositions and degrees of polymerization for use in food and feed.

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Fig. 1: Anthocyanin and PA accumulation in ans-1, ldox-1 and ans-1 ldox-1 mutants of M. truncatula.
Fig. 2: Analysis of PAs in pods of ans-1 lar-1 and ldox-1 lar-1 double mutants.
Fig. 3: Activity of ANS and LDOX with (+)-catechin in vitro and in vivo.
Fig. 4: Association of LDOX expression with catechin incorporation into PAs in different plants.

Data availability

All data needed to evaluate the conclusions in the paper are present in the paper and/or the Supplementary Materials. The data sets supporting the results of this article are available in the NCBI Sequence Read Archive (SRA) repository, NCBI SRA accession No. PRJNA491470.

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Acknowledgements

We thank S. Temple for critical reading of the manuscript and V. Shulaev for assistance with high mass accuracy LCMS. This work was supported by Forage Genetics International Inc. and the University of North Texas.

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R.A.D., J.H.J. and X.R. conceived and designed the study, and analysed and interpreted data. J.H.J., X.X. and X.R. acquired data. J.H.J. and R.A.D. wrote the original draft. R.A.D reviewed and edited the final manuscript.

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Correspondence to Richard A. Dixon.

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R.A.D. and J.H.J. are inventors on a United States provisional patent application filed by the University of North Texas that describes methods for engineering PA composition.

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Jun, J.H., Xiao, X., Rao, X. et al. Proanthocyanidin subunit composition determined by functionally diverged dioxygenases. Nature Plants 4, 1034–1043 (2018). https://doi.org/10.1038/s41477-018-0292-9

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