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Cadmium (Cd) is a highly toxic heavy metal that causes serious damage to plant and human health. <i>Phytolacca acinosa</i> Roxb. has a large amount of aboveground biomass and a rapid growth rate, and it has been identified as a novel type of Cd hyperaccumulator that can be harnessed for phytoremediation. However, the molecular mechanisms underlying the response of <i>P. acinosa</i> to Cd²⁺ stress remain largely unclear. In this study, the phenotype, biochemical, and physiological traits of <i>P. acinosa</i> seeds and seedlings were analyzed under different concentrations of Cd²⁺ treatments. The results showed higher Cd²⁺ tolerance of <i>P. acinosa</i> compared to common plants. Meanwhile, the Cd²⁺ content in shoots reached 449 mg/kg under 10 mg/L Cd²⁺ treatment, which was obviously higher than the threshold for Cd hyperaccumulators. To investigate the molecular mechanism underlying the adaptability of <i>P. acinosa</i> to Cd stress, RNA-Seq was used to examine transcriptional responses of <i>P. acinosa</i> to Cd stress. Transcriptome analysis found that 61 genes encoding TFs, 48 cell wall-related genes, 35 secondary metabolism-related genes, 133 membrane proteins and ion transporters, and 96 defense system-related genes were differentially expressed under Cd²⁺ stress, indicating that a series of genes were involved in Cd²⁺ stress, forming a complex signaling regulatory mechanism. These results provide new scientific evidence for elucidating the regulatory mechanisms of <i>P. acinosa</i> response to Cd²⁺ stress and new clues for the molecular breeding of heavy metal phytoremediation.