Nanoplastics (NPs) can adversely affect living organisms. However, the uptake of NPs by plants and the physiological and molecular mechanisms underlying NP-mediated plant growth remain unclear, particularly in the presence of iron minerals and humic acid (HA). In this study, we investigated NP accumulation in rice (Oryza sativa L.) and the physiological effects of exposure to polystyrene NPs (0, 20, and 100 mg L−1) in the presence of iron plaque (IP) and HA. NPs were absorbed on the root surface and entered cells, and confocal laser scanning microscopy confirmed NP uptake by the roots. NP treatments decreased root superoxide dismutase (SOD) activity (28.9–44.0%) and protein contents (31.2–38.6%). IP and HA (5 and 20 mg L−1) decreased the root protein content (20.44–58.3% and 44.2–45.2%, respectively) and increased the root lignin content (22.3–27.5% and 19.2–29.6%, respectively) under NP stress. IP inhibited the NP-induced decreasing trend of SOD activity (19.2–29.5%), while HA promoted this trend (48.7–50.3%). Transcriptomic and metabolomic analysis (Control, 100NPs, and IP-100NPs-20HA) showed that NPs inhibited arginine biosynthesis, and alanine, aspartate, and glutamate metabolism and activated phenylpropanoid biosynthesis related to lignin. The coexistence of IP and HA had positive effects on the amino acid metabolism and phenylpropanoid biosynthesis induced by NPs. Regulation of genes and metabolites involved in nitrogen metabolism and secondary metabolism significantly altered the levels of protein and lignin in rice roots. These findings provide a scientific basis for understanding the environmental risk of NPs under real environmental conditions.
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