Abstract
As one of the largest families of transcription factors (TFs) in plants, R2R3-MYB proteins play crucial roles in regulating a series of plant-specific biological processes. Although the diversity of plant R2R3-MYB TFs has been studied previously, the processes and mechanisms underlying the expansion of these proteins remain unclear. Here, we performed evolutionary analyses of plant R2R3-MYB TFs with dense coverage of streptophyte algae and embryophytes. Our analyses revealed that ancestral land plants exhibited 10 subfamilies of R2R3-MYB proteins, among which orthologs of seven subfamilies were present in chlorophytes and charophycean algae. We found that asymmetric gene duplication events in different subfamilies account for the expansion of R2R3-MYB proteins in embryophytes. We further discovered that the largest subfamily of R2R3-MYBs in land plants, subfamily VIII, emerged in the common ancestor of Zygnematophyceae and embryophytes. During plant terrestrialization, six duplication events gave rise to seven clades of subfamily VIII. Subsequently, this TF subfamily showed a tendency for expansion in bryophytes, lycophytes, and ferns and extensively diversified in ancestral gymnosperms and angiosperms in clades VIII-A-1, VIII-D, and VIII-E. In contrast to subfamily VIII, other subfamilies of R2R3-MYB TFs have remained less expanded across embryophytes. The findings regarding phylogenetic analyses, auxiliary motifs, and DNA-binding specificities provide insight into the evolutionary history of plant R2R3-MYB TFs and shed light on the mechanisms underlying the extensive expansion and subsequent sub- and neofunctionalization of these proteins.