Flavonoids - flowers, fruit, forage and the future

Abstract

Flavonoids are plant-specific secondary metabolites that arose early during land-plant colonisation, most likely evolving for protection from UV-B and other abiotic stresses. As plants increased in complexity, so too did the diversity of flavonoid compounds produced and their physiological roles. The most conspicuous are the pigments, including yellow aurones and chalcones, and the red/purple/blue anthocyanins, which provide colours to flowers, fruits and foliage. Anthocyanins have been particularly well studied, prompted by the ease of identifying mutants of genes involved in biosynthesis or regulation, providing an important model system to study fundamental aspects of genetics, gene regulation and biochemistry. This has included identifying the first plant transcription factor, and later resolving how multiple classes of transcription factor coordinate in regulating the production of various flavonoid classes - each with different activities and produced at differing developmental stages. In addition, dietary flavonoids from fruits/vegetables and forage confer human- and animal-health benefits, respectively. This has prompted strong interest in generating new plant varieties with increased flavonoid content through both traditional breeding and plant biotechnology. Gene-editing technologies provide new opportunities to study how flavonoids are regulated and produced and to improve the flavonoid content of flowers, fruits, vegetables and forages.

Keywords: Flavonoid; MYB; anthocyanin; proanthocyanidin; transcription factor.

Figure 1.

Physiological roles of flavonoids in plants and their biosynthesis. A, Flavonoids perform numerous physiological roles in plants. Flavones or flavonols act as UV-B sunscreens and are induced by high irradiance white light as well as UV-B and UV-A. Isoflavonoids are signalling molecules for rhizobia, forming root nodules (nitrogen fixation) in legumes. Anthocyanins provide visual cues to pollinators and seed distributers. Bees have preferences for purple-blue flowers and/or flowers with high contrast patterns with other pigments, including UV-B absorbing flavones or flavonols; bird-pollinated flowers are less attractive to bees and are typically red with low UV-B absorption, while moth-pollinated flowers are typically white with high UV-B absorption. Anthocyanins are often also produced during senescence, or by abiotic stresses such as nutrient deficiency (N, P), high light intensities or cold. Proanthocyanidins are unpalatable, deterring insect pests in vegetative tissues and consumption of immature fruit. B, The flavonoid biosynthetic pathway begins with chalcone synthase (CHS). Many tissues contain multiple classes of flavonoids, which share common biosynthetic steps. For example, CHS, CHI and F3H are common to flavonol (green), proanthocyanidin (brown) and anthocyanin (purple) biosynthesis. Coordinated expression of genes in each these pathways is necessary for metabolite biosynthesis to occur. The activities of flavonoid 3′hydroxylase (F3′H), flavonoid 3′5′hydroxylase (F3′5′H) add hydroxyl groups to the 3 and 5 position of the B-ring of dihydroflavonols which can progress through towards anthocyanins, proanthocyanidins, but also to generate flavonols with different hydroxylation patterns (not shown). The three main anthocyanidins are pelargonidin (Pel), cyanidin (Cy) and delphinidin (Del), which can become glycosylated and decorated to produce anthocyanins, or undergo reduction to become flavan 3-ols for proanthocyanidins. Some biosynthetic steps are not fully resolved (?), such as those involved in converting aurones into auronidins, and for polymerising flavan 3-ols into oligomeric and polymeric proanthocyanidins. Abbreviations: chalcone synthase (CHS), chalcone Isomerase-Like (CHI-L), chalcone isomerase (CHI), flavone synthase (FNS), flavanone 3-hydroxylase (F3H), flavonoid 3′-hydroxylase (F3′H), flavonoid 3′5′-hydroxylase (F3′5′H), flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS)/leucoanthocyanidin dioxygenase (LDOX), leucoanthocyanidin reductase (LAR), anthocyanidin reductase (ANR), flavonoid 3-O-glucosyltransferase (3GT), flavonoid 5-O-glucosyltransferase (5GT), acyltransferase (AT), methyltransferase (MT), stilbene synthase (STS), aurone synthase (AS), isoflavone synthase (IFS).

Figure 2.

R2R3 MYB genes regulate flavonoid biosynthesis. A, MrBayes Phylogenetic tree of R2R3-MYB subgroups (posterior probabilities shown), rooted on MpMYB14 from Marchantia polymorpha. Notable examples of each subgroup (SG) are indicated. * indicate subgroups that form MYB-bHLH-WDR (MBW) complexes. B, Several branches of the flavonoid biosynthetic pathway are regulated by R2R3-MYB proteins as part of MBW, in particular those belonging to SG5 (proanthocyanidins) and 6 (anthocyanins) are well characterised. The stoichiometry isn’t clear, but at least two different MYB proteins can be present, bound through their interactions with dimerised bHLH proteins. (1) If all three components are present, they can form a complex, which (2) can then bind the promoters of target genes and activate transcription, ultimately resulting in phenotypes such as metabolite production. (3) bHLH and WDR genes are themselves regulated by MBW complexes (reinforcement). (4) R2R3- and R3-MYB repressor genes are regulated by MBW (feed-back inhibition). They can interfere with the correct assembly of the MBW complex, by competing with MYB activator proteins for bHLHs. R2R3-MYB repressors can also be incorporated into MBW complexes and recruited to promoters where C-terminal motifs (e.g. EAR, TLLFR) inhibit transcription.