Proanthocyanidins Alleviate Cadmium Stress in Industrial Hemp (Cannabis sativa L.)

Industrial hemp (Cannabis sativa L.), an annual herbaceous cash crop, is widely used for the remediation of heavy metal-contaminated soils due to its short growth cycle, high tolerance, high biomass, and lack of susceptibility to transfer heavy metals into the human food chain. In this study, a significant increase in proanthocyanidins was found in Yunnan hemp no. 1 after cadmium stress. Proanthocyanidins are presumed to be a key secondary metabolite for cadmium stress mitigation. Therefore, to investigate the effect of proanthocyanidins on industrial hemp under cadmium stress, four experimental treatments were set up: normal environment, cadmium stress, proanthocyanidin treatment, and cadmium stress after pretreatment with proanthocyanidins. The phenotypes from the different treatments were compared. The experimental results showed that pretreatment with proanthocyanidins significantly alleviated cadmium toxicity in industrial hemp. The transcriptome and metabolome of industrial hemp were evaluated in the different treatments. Proanthocyanidin treatment and cadmium stress in industrial hemp mainly affected gene expression in metabolic pathways associated with glutathione metabolism, phenylpropanoids, and photosynthesis, which in turn altered the metabolite content in metabolic pathways of phenylalanine, vitamin metabolism, and carotenoid synthesis. The combined transcriptomic and metabolomic analysis revealed that proanthocyanidins mitigated cadmium toxicity by enhancing photosynthesis, secondary metabolite synthesis, and antioxidant synthesis. In addition, exogenous proanthocyanidins and cadmium ions acted simultaneously on EDS1 to induce the production of large amounts of salicylic acid in the plant. Finally, overexpression of CsANR and CsLAR, key genes for proanthocyanidins synthesis in industrial hemp, was established in Arabidopsis plants. The corresponding plants were subjected to cadmium stress, and the results showed that CsLAR transgenic plants were more tolerant to cadmium than the CsANR transgenic and wild-type Arabidopsis plants. The results showed that salicylic acid and jasmonic acid were increased in Arabidopsis overexpressing CsLAR compared to AT wild-type Arabidopsis, and levels of secondary metabolites were significantly higher in Arabidopsis overexpressing CsLAR than in AT wild-type Arabidopsis. These results revealed how proanthocyanidins alleviated cadmium stress and laid the foundation for breeding industrial hemp varieties with higher levels of proanthocyanidins and greater tolerance.