Exogenous nitrogen mitigation of cadmium stress in Pinellia ternata is associated with the regulation of arginine and proline metabolism

Abstract

Cadmium (Cd) contamination is a major constraint limiting the yield and quality of medicinal plants. In this study, the effects of exogenous nitrogen on alleviating Cd toxicity in Pinellia ternata seedlings were investigated, and the underlying mechanisms were explored using integrated physiological, transcriptomic, untargeted metabolomic, and rhizosphere microbial analyses. Nitrogen supplementation (150 mg N kg-¹ supplied as urea) mitigated Cd-induced growth inhibition, increasing root length by approximately 31% on days 6 and 9 and partially improving leaf area by day 9. Nitrogen supplementation also significantly decreased the root-to-shoot Cd translocation factor at all three sampling times, while root Cd concentrations remained largely unchanged; shoot Cd concentration was significantly reduced on day 9. Physiologically, nitrogen supplementation modulated antioxidant enzyme activity and reduced malondialdehyde (MDA) accumulation at the early stage, consistent with a time-dependent alleviation of membrane lipid peroxidation under Cd stress. Integrated multi-omics analyses revealed extensive transcriptional and metabolic reprogramming. Transcriptomic analysis identified 36,036, 9,373, and 33,136 differentially expressed genes on days 3, 6, and 9, respectively, whereas metabolomic profiling identified 947, 1,203, and 729 differentially accumulated metabolites, respectively. KEGG enrichment analysis of the differentially accumulated metabolites consistently highlighted arginine and proline metabolism across the three sampling times, accompanied by enrichment of related pathways, including ascorbate and aldarate metabolism, glutathione metabolism, ABC transporters, and nitrogen metabolism. Within the arginine and proline metabolism pathway, genes encoding pyrroline-5-carboxylate reductase, spermidine synthase, and aldehyde dehydrogenase family members showed treatment-dependent differential expression, accompanied by concurrent changes in the relative abundances of proline, glutamine, and polyamines. Collectively, these findings indicate that exogenous nitrogen is associated with improved Cd tolerance in P. ternata through coordinated regulation of Cd translocation, arginine and proline metabolism, antioxidant and transport-related processes, and rhizosphere-associated responses. This study provides integrated evidence for nitrogen-mediated Cd tolerance and offers insights into nutrient management for mitigating Cd stress during P. ternata cultivation.