Evolution of R2R3-MYB regulators of plant pigmentation

The family of MYB-domain transcription factors expanded greatly during plant evolution, reaching over 200 members in some species. To assess how MYBs were co-opted into distinct processes, we studied MYB regulators of distinct branches of the flavonoid pathway. To this end, we expressed different MYBs in Petunia hybrida and assessed the capacity of each one to induce anthocyanin accumulation. Study of MYBs from different species in the same expression system did not reveal the functions of individual proteins in the plants from where they originated, but clearly pinpointed functional differences. Pigmentation MYBs from different species induced transcription of different but overlapping sets of target genes when expressed in the same cells, while yeast two-hybrid assays revealed divergence even among MYBs from the same phylogenetic subgroup for their capacity to bind specific bHLH and WD40 protein partners. These results suggest that diversification of proteins occurred after gene duplication, resulting in specificity for different target genes. Such evolution of proteins parallels the acquisition of new branches of the flavonoid pathway in different species. The overlap in the sets of induced genes could be indicative of their ancestral function and supports stepwise evolution toward regulating new pathway branches and acquiring new products in plant tissues.