Transcriptome and organic acid profiling uncover Cd tolerance mechanisms in Ilex asprella roots

Abstract Background Ilex asprella is a traditional Chinese medicinal herb of considerable economic value. However, it has a strong tendency to accumulate cadmium (Cd), resulting in potentially excessive heavy metal levels that severely compromise its medicinal quality and industrial development. The physiological and molecular mechanisms underlying Cd tolerance and accumulation in I. asprella remain unclear. Organic acids play crucial roles in plant responses to heavy metal stress. Therefore, this study aimed to elucidate the molecular mechanisms of Cd tolerance in I. asprella roots by integrating transcriptomic and organic acid profiling analyses. Results Under hydroponic treatment with 50 μmol/L CdCl₂ for different durations (0–72h), the total Cd content in I. asprella roots increased continuously and plateaued after 48h. Based on Mfuzz time-series clustering and lagged cross-correlation analysis, 6 Cd-responsive organic acids were identified: citric acid, malic acid, lactic acid, oxalic acid, acetic acid, and sinapic acid. These acids exhibited persistently increasing patterns in both roots and root exudates and were strongly positively correlated with root Cd content. Transcriptomic analysis revealed three major gene expression patterns, and weighted gene co-expression network analysis (WGCNA) identified core regulatory modules significantly associated with the six organic acids. Functional enrichment analysis showed that hub genes in these modules were mainly involved in pathways such as carbon metabolism, phenylpropanoid biosynthesis, and glyoxylate and dicarboxylate metabolism. Co-expression network construction further identified 15 key transcription factors (e.g., LBD31, WOX13, and MYB36) that may coordinately regulate organic acid biosynthesis and transport. Conclusions These processes may contribute to Cd tolerance and accumulation in I. asprella. Cd stress activates specific transcription factors, upregulating functional genes involved in carbon metabolism, glycolysis, phenylpropanoid biosynthesis, and related pathways, thereby promoting the synthesis of organic acids such as citrate and malate. These organic acids contribute to Cd detoxification through multiple synergistic mechanisms, including Cd chelation, vacuolar sequestration, reactive oxygen species scavenging, and rhizosphere chelation. Ultimately, this enhances Cd tolerance and accumulation in This study proposes a model of Cd tolerance and accumulation mediated by organic acid metabolism in I. asprella: I. asprella . These findings provide genetic resources and a theoretical foundation for further understanding plant metal detoxification and for breeding varieties with reduced Cd accumulation.