Abstract

Melatonin is a pleiotropic molecule that plays an important role in regulating plant growth, development and abiotic stress responses. Although melatonin biosynthesis has been extensively characterised, the catabolic pathways that regulate its content remain largely unexplored in many crops, including tomato ( Solanum lycopersicum L.). Here, we demonstrate that the tomato N -acetylserotonin deacetylase (SlASDAC) functions as a negative regulator of melatonin accumulation, likely involved in melatonin catabolism. We found that SlASDAC expression is strongly induced by exogenous melatonin treatment, and its transcript levels are inversely correlated with melatonin accumulation in fruit. The SlASDAC protein localises to the chloroplast, and protein–ligand docking combined with molecular dynamics simulations indicate a stable interaction between SlASDAC and melatonin. Functionally, overexpression of SlASDAC decreased melatonin levels and impaired leaf physiological traits. In contrast, virus‑induced gene silencing and CRISPR/Cas9‑mediated knockout of SlASDAC led to a substantial melatonin accumulation, thereby enhancing leaf pigmentation, trichome density, and carotenoid accumulation in fruit. Notably, the elevated melatonin levels in SlASDAC knockout lines contributed to the enhanced salt tolerance during germination and the seedling stage. Conversely, SlASDAC overexpression compromised salt tolerance, which could be rescued by exogenous melatonin treatment. Mechanistically, SlWRKY75 directly binds to W‑box elements in the SlASDAC promoter and represses its transcription. Consistently, knockout of SlWRKY75 resulted in upregulated SlASDAC expression and reduced melatonin accumulation, thereby establishing a SlWRKY75–SlASDAC module that modulates melatonin catabolism. Collectively, our findings demonstrate that this regulatory pathway fine-tunes melatonin levels to orchest