GiMYB76, a MeJA-inducible R2R3-type transcription factor, regulates licochalcone A biosynthesis in licorice (Glycyrrhiza inflata)

Licochalcone A (LCA) is a characteristic compound in licorice Glycyrrhiza inflata and is widely utilized in pharmaceutical and cosmetic industries. However, the biosynthetic pathway and regulatory mechanisms of LCA remain poorly understood. In this study, we first found the accumulation of LCA is induced by methyl jasmonate (MeJA). Given that MYB transcriptional factors are well-documented as key regulators of flavonoid biosynthesis, we identified a total of 147 GiR2R3-MYB genes in G. inflata, which were classified into 28 subgroups. The chromosome distributions, sequence characteristics, gene structures, duplication events and cis-acting elements were also investigated. Through integrated analysis of GiR2R3-MYBs expression patterns across different tissues and under MeJA treatment, along with phylogenetic relationship, we identified GiMYB76—a MeJA-inducible MYB transcription factor—as a potential regulator of LCA accumulation. Functional validation showed that transgenic hairy roots overexpressing GiMYB76 exhibited a significant increase in LCA content. DAP-seq analysis of GiMYB76 revealed potential target genes involved in flavonoid biosynthesis regulation. Subsequent promoter activity assay verified that GiMYB76 can bind to the promoter and activate the expression of GiCHS4. Consistently, overexpression of GiCHS4 in G. inflata hairy roots also significantly enhanced LCA production. This study not only clarifies that GiMYB76 transcriptionally activated GiCHS4 to promote LCA biosynthesis but also provides valuable insights for basic research on licorice and the development of related industries.