Citric acid-driven cadmium uptake and growth promotion mechanisms in Brassica napus

Citric acid (CA) is well-known for mitigating cadmium (Cd) toxicity in plants. Yet, the underlying mechanisms driving growth promotion, Cd detoxification/tolerance, and enhanced phytoremediation processes remain incompletely understood. This study investigated the effects of CA application (2.5 mM) on Brassica napus grown in Cd-contaminated (30 mg kg^-1) growth medium through a controlled pot experiment. Cd exposure alone significantly impaired various plant physiological parameters in B. napus. Whereas CA application significantly (p < 0.05) enhanced physiological attributes, Cd detoxification and tolerance by modulating key genes involved in photosynthesis and Cd transport, including the metal-transporting P_1B-type ATPases (Cd/zinc heavy metal-transporting ATPase 1; HMA1) and light-harvesting chlorophyll a/b-binding 3 (LHCB3). Notably, CA application increased Cd accumulation in stems and leaves by 4% and 35%, respectively, enhancing bioconcentration factors (BCF) by 12% in stems and 40% in leaves while reducing root BCF by 10%. This translocation was facilitated by the upregulation of HMA4, HMA2, and plant Cd resistance (PCR2) genes in plant leaves, improving Cd mobility within the plant. Furthermore, CA induced a 34% increase in phytochelatins and a 32% upregulation in metallothioneins, accompanied by a significant reduction in oxidative stress markers, including a 40% decrease in hydrogen peroxide and a 44% decline in malondialdehyde levels in leaves. Enhanced antioxidant enzyme activity and osmolyte accumulation further contributed to improved Cd detoxification/sequestration in leaves, reduced oxidative stress, and improved photosynthetic efficiency, resulting in enhanced plant biomass production and Cd tolerance. Transcriptomic analysis showed that CA treatment substantially influenced the expression of 12,291 differentially expressed genes (DEGs), with 750 common genes consistently downregulated in CK vs Cd treatment group but upregulated in Cd vs Cd-CA treatment group. Additionally, CA modulated 11 DEGs associated with 32 gene ontologies in the citrate pathway under Cd stress, highlighting its targeted regulatory effect on metabolic pathways involved in Cd stress response. This study offers novel insights into the synergistic role of CA in promoting plant growth and regulating Cd uptake in B. napus, highlighting its potential to enhance phytoremediation strategies.