CsWRKY6 ‐2/ CsWRKY12 module regulates gallated catechin biosynthesis in tea plants

Tea cis-galloylated catechins, mainly epigallocatechin-3-gallate (EGCG) and epicatechin gallate (ECG), are important natural products that endow tea with flavor and health benefits. WRKY transcription factors play a crucial role in regulating plant metabolite biosynthesis, yet the mechanisms underlying their involvement in EGCG and ECG accumulation in tea plants (Camellia sinensis) remain unclear. Our results demonstrated that CsWRKY6-2 was negatively correlated with EGCG and ECG levels in different tea cultivars and leaves from distinct leaf positions. CsWRKY6-2 silencing resulted in increased EGCG and ECG content in tea leaves, whereas its overexpression had the opposite effect. Its regulation is tied to changes in key biosynthetic genes CsSCPL4, CsSCPL5, and their activator, CsWRKY12. Consistently, CsWRKY6-2 bound to the promoters of CsSCPL4, CsSCPL5, and CsWRKY12, inhibiting their expression. Additionally, CsWRKY6-2 interacted with CsWRKY12, which disrupted the regulation of CsSCPL4 and CsSCPL5 expression by CsWRKY6-2 and CsWRKY12. Further analyses revealed that CsWRKY6-2 and CsWRKY12 exerted varying regulatory effects on CsSCPL4 and CsSCPL5 expression across different leaf positions and tea cultivars, thereby jointly controlling EGCG and ECG accumulation. Collectively, these findings suggest a precise mechanism by which the CsWRKY6-2/CsWRKY12 module mediates the accumulation of galloylated catechins in tea plants, providing new insights for breeding tea cultivars.