MYBA and MYBPA transcription factors co-regulate anthocyanin biosynthesis in blue-coloured berries.

Katja Karppinen,Hely Häggman,Richard V Espley,Tony Mcghie,Declan J Lafferty,Nick W Albert,Andrew C Allan,Nelli Mikkola,Laura Jaakola,Bilal M Afzal

doi:10.1111/nph.17669

Abstract

The regulatory network of R2R3 MYB transcription factors in anthocyanin biosynthesis is not fully understood in blue-coloured berries containing delphinidin compounds. We used blue berries of bilberry (Vaccinium myrtillus) to comprehensively characterise flavonoid-regulating R2R3 MYBs, which revealed a new type of co-regulation in anthocyanin biosynthesis between members of MYBA-, MYBPA1- and MYBPA2-subgroups. VmMYBA1, VmMYBPA1.1 and VmMYBPA2.2 expression was elevated at berry ripening and by abscisic acid treatment. Additionally, VmMYBA1 and VmMYBPA1.1 expression was strongly downregulated in a white berry mutant. Complementation and transient overexpression assays confirmed VmMYBA1 and VmMYBA2 to induce anthocyanin accumulation. Promoter activation assays showed that VmMYBA1, VmMYBPA1.1 and VmMYBPA2.2 had similar activity towards dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS), but differential regulation activity for UDP-glucose flavonoid 3-O-glucosyltransferase (UFGT) and flavonoid 3'5'-hydroxylase (F3'5'H) promoters. Silencing of VmMYBPA1.1 in berries led to the downregulation of key anthocyanin and delphinidin biosynthesis genes. Functional analyses of other MYBPA regulators, and a member of novel MYBPA3 subgroup, associated them with proanthocyanidin biosynthesis and F3'5'H expression. The existence of 18 flavonoid-regulating MYBs indicated gene duplication, which may have enabled functional diversification among MYBA, MYBPA1 and MYBPA2 subgroups. Our results provide new insights into the intricate regulation of the complex anthocyanin profile found in blue-coloured berries involving regulation of both cyanidin and delphinidin branches.

Highlights

Flavonoids are a large group of polyphenols in plants
Our results suggest a new type of abscisic acid (ABA)-induced co-regulation among MYBA, MYBPA1 and MYBPA2 transcription factors (TFs) in ripening-associated anthocyanin biosynthesis and provide functional evidence that MYBPA1-type TF contribute to anthocyanin biosynthesis during berry ripening by directly activating key biosynthetic genes
Our results revealed that the transcripts of VmMYBA1 and VmMYBPA1.1 were most highly associated with ripening fruit, showing a similar expression pattern to the structural genes related to anthocyanin biosynthesis (Figs 3, S2, S3), suggesting the involvement in regulation of anthocyanin accumulation in berry

Summary

Introduction

Anthocyanins, proanthocyanidins (PAs) and flavonols are the major classes of flavonoids found in almost all higher plants. Anthocyanins contribute to the red and blue colours in flowers and ripe fruits facilitating pollination and seed dispersal but they have a role in protecting plants against stress (Saigo et al, 2020). Chalcone synthase (CHS), chalcone isomerase (CHI) and flavanone 3-hydroxylase (F3H) are responsible for producing dihydroflavonol precursors for all flavonoid branches. Flavonol synthase (FLS) directs the dihydroflavonol precursors to the flavonol route, while the action of dihydroflavonol 4-reductase (DFR) following anthocyanidin synthase (ANS) and its homologue leucoanthocyanidin dioxygenase (LDOX) contributes to both anthocyanin and PA synthesis (Jun et al, 2018). The pathway to PAs involves leucoanthocyanidin reductase (LAR) producing 2,3-trans-2R,3S-flavan-3-ols (+)-catechin, (+)-gallocatechin) and anthocyanidin reductase (ANR) for production of 2,3-cis-2R,3R-flavan-3-ols

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Discussion

Conclusion