Complexity and robustness of the flavonoid transcriptional regulatory network revealed by comprehensive analyses of MYB–bHLH–WDR complexes and their targets in Arabidopsis seed

Abstract

SummaryIn Arabidopsis thaliana, proanthocyanidins (PAs) accumulate in the innermost cell layer of the seed coat (i.e. endothelium, chalaza and micropyle). The expression of the biosynthetic genes involved relies on the transcriptional activity of R2R3-MYB and basic helix-loop-helix (bHLH) proteins which form ternary complexes ('MBW') with TRANSPARENT TESTA GLAB-RA1 (TTG1) (WD repeat protein). The identification of the direct targets and the determination of the nature and spatio-temporal activity of these MBW complexes are essential steps towards a comprehensive understanding of the transcriptional mechanisms that control flavonoid biosynthesis.In this study, various molecular, genetic and biochemical approaches were used. Here, we have demonstrated that, of the 12 studied genes of the pathway, only dihydroflavonol-4-reductase (DFR), leucoanthocyanidin dioxygenase (LDOX), BANYULS (BAN), TRANSPARENT TESTA 19 (TT19), TT12 and H + -ATPase isoform 10 (AHA10) are direct targets of the MBW complexes. Interestingly, although the TT2-TT8-TTG1 complex plays the major role in developing seeds, three additional MBW complexes (i.e. MYB5-TT8-TTG1, TT2-EGL3-TTG1 and TT2-GL3-TTG1) were also shown to be involved, in a tissuespecific manner. Finally, a minimal promoter was identified for each of the target genes of the MBW complexes. Altogether, by answering fundamental questions and by demonstrating or invalidating previously made hypotheses, this study provides a new and comprehensive view of the transcriptional regulatory mechanisms controlling PA and anthocyanin biosynthesis in Arabidopsis.

MYB5 and MYB14 Play Pivotal Roles in Seed Coat Polymer Biosynthesis in Medicago truncatula

“…As shown in Supplemental Figure S4, both MtPAR and MtMYB14 rescued the Arabidopsis tt2 phenotype (leading to dark seed coats), supporting the conclusion that MtPAR and MtMYB14 are functional orthologs of Arabidopsis TT2. Although Xu et al (2014) observed that AtMYB5 could partially rescue the tt2 phenotype, we did not observe such rescue with MtMYB5.…”

The MYB182 Protein Down-Regulates Proanthocyanidin and Anthocyanin Biosynthesis in Poplar by Repressing Both Structural and Regulatory Flavonoid Genes

“…The MBW complex interacts directly with promoter DNA via multiple cis-elements. Known target sequences include the Myb-response element and AC elements for R2R3-MYB proteins and the E-box or bHLH-binding motif, which is bound by the bHLH proteins (Feller et al, 2011;Lai et al, 2013;Xu et al, 2014). In contrast, WDR proteins have not been shown to bind DNA but are thought to function by interacting with MYB and bHLH proteins (Baudry et al, 2004).…”

The Phenylpropanoid Pathway Is Controlled at Different Branches by a Set of R2R3-MYB C2 Repressors in Grapevine

“…Depending on the plant lineage, these transcriptional regulatory proteins may affect all biosynthetic genes in a single module (as in maize [Zea mays]) or separately, regulating the so-called late structural genes, like in petunia (Petunia hybrida), snapdragon (Antirrhinum majus), and Arabidopsis (Arabidopsis thaliana; Mol et al, 1998). In these species, the expression of the early biosynthetic genes is controlled by other regulators (sometimes redundantly with the MBW complex; Tornielli et al, 2009;Xu et al, 2014). In some cases (e.g.…”

“…The phenylpropanoid pathway is finely tuned and tightly controlled by transcription factors (Albert et al, 2011;Xu et al, 2014). In this control, several protein families participate and mutually interact, giving rise to regulatory specificity (Grotewold et al, 2000).…”