CsBPC6-CsNACX2-CsTCS1 Module Positively Regulates Caffeine Biosynthesis Under PEG-Induced Drought Stress in Tea Plants (Camellia sinensis).
Caffeine biosynthesis in tea plants (Camellia sinensis) is dynamically regulated by environmental cues; however, the mechanisms linking early drought signals to caffeine accumulation remain unclear. Here, we show that acute PEG-induced osmotic stress induces rapid caffeine accumulation, accompanied by transcriptional activation of the key biosynthetic gene CsTCS1. Through integrative analyses, including GWAS and co-expression network modeling, we identified a CsBPC6-CsNACX2-CsTCS1 module as a central regulatory hub. Mechanistically, CsNACX2 directly binds the promoter of the caffeine synthase gene CsTCS1, activating its transcription and promoting caffeine accumulation. Moreover, the drought-responsive transcription factor CsBPC6 acts upstream to transactivate CsNACX2. Functional validation using gene silencing and transient overexpression assays confirmed that the CsBPC6-CsNACX2-CsTCS1 module positively regulates caffeine biosynthesis under PEG-induced drought conditions. Collectively, our findings uncover a previously uncharacterized drought-responsive transcriptional module governing caffeine biosynthesis, providing new mechanistic insights into how environmental cues regulate secondary metabolism in tea plants.