MYB transcription factors regulate O-methylated EGCG biosynthesis in different leaf positions of tea plants (Camellia sinensis)

(−)-Epigallocatechin-3-O-(3-O-methyl)gallate (EGCG3″Me) and (−)-epigallocatechin-3-O-(4-O-methyl)gallate (EGCG4″Me) are the crucial O-methylated EGCG compounds in tea plants, with potential bioactivity and potential medicinal uses. The maturity of fresh tea leaves in different leaf positions is an important factor in determining the quality of tea made from various types of tea processing. However, the underlying molecular mechanism of the differential accumulation of O-methylated EGCG in leaf positions that differ regarding maturity are still unclear. In this study, ‘Lingtou Dancong (LTDC)’ was revealed to be a tea cultivar with high EGCG3″Me and EGCG4″Me contents. Specifically, in LTDC tea variety, EGCG3″Me predominantly accumulated in the third and fourth leaves, while EGCG4″Me was primarily found in the stem. The accumulation of these compounds was significantly positively correlated with the expression patterns of their biosynthetic genes (CsFAOMT1 or CsFAOMT2), rather than with their synthetic precursor EGCG. R2R3-MYB transcription factors, were identified by WGCNA and differential expression analysis. Furthermore, dual-luciferase reporter assay, EMSA, and Agrobacterium-mediated gene overexpression assay revealed that CsMYB123 and CsMYB86 promote EGCG3″Me and EGCG4″Me biosynthesis by activating their biosynthetic CsFAOMT1 and CsFAOMT2 genes expression, respectively; whereas CsMYB73 inhibits EGCG3″Me biosynthesis. This study elucidated that MYB transcription factors regulate the differential accumulation of O-methylated EGCG in different leaf positions of tea plants, which providing new technical insights relevant to regulation mechanism of O-methylated catechins.