Novel R2R3-MYB transcription factor CsMYB39 influences epigallocatechin gallate biosynthesis in tea plant (Camellia sinensis) by regulating CsSCPL11-IA gene expression

Abstract

Epigallocatechin-3-gallate (EGCG) is the most bioactive galloylated catechin in tea ( Camellia sinensis (L.) O. Kuntze) plants. Its biosynthesis is precisely regulated by transcription factors. In this study, we used the promoter of the CsSCPL11-IA gene, which encodes a key enzyme in EGCG biosynthesis, as bait. Using yeast one-hybrid technology, we isolated and identified a novel R2R3-MYB transcription factor, CsMYB39 . Results revealed that the CsMYB39 gene was specifically localised in the nucleus. Transient overexpression of CsMYB39 in tea leaves caused a significant increase in total catechin content of 0.28-fold, and EGCG levels rose by 0.64-fold. After suppressing CsMYB39 gene expression using virus-induced gene silencing technology, catechin and EGCG contents in tea leaves decreased by 79% and 71%, respectively. Heterologous overexpression of CsMYB39 in tobacco activated the flavonoid metabolic pathway and significantly promoted the accumulation of flavonoid precursors. Correlation analysis revealed a highly significant positive correlation between the expression levels of CsMYB39 and CsSCPL11-IA (encoding a key enzyme for catechin acylation; R = 0.94, P < 0.01). Dual luciferase reporter, electrophoretic mobility shift, and yeast one-hybrid analyses confirmed that CsMYB39 binds directly to the specific promoter regions of CsSCPL11-IA , strongly activating CsSCPL11-IA transcription. In summary, CsMYB39 acts as a positive regulator of EGCG biosynthesis by activating CsSCPL11-IA gene expression. Our findings provide new genetic resources and theoretical foundations for improving tea quality through molecular breeding strategies.