Phosphorus (P) is necessary for nitrogen fixation in the root nodules of soybeans, a symbiotic process whereby plants support bacterial nitrogen fixation to obtain nitrogen needed for plant growth. Nitrogen accumulation, quantity, weight, specific nitrogenase activity (SNA) and acetylene reduction activity (ARA) of root soybean nodules were analyzed, using a broadly targeted metabolomics method incorporating liquid chromatography-mass spectrometry (LC-MS) to study the effects of P level (1, 11, 31, 61 mg/L denoted by P1, P11, P31, P61) on the types and abundance of various metabolites and on the expression of associated metabolic pathways in soybean root nodules. Nitrogen accumulation, quantity, weight, SNA and ARA of root nodules were inhibited by P stress. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that root nodules responded to P stress by increasing the number of amino acids and derivatives. Down-regulation of ABA, phosphorylcholine, and D-glucose 6-phosphate affected carotenoid biosynthesis, glycerophospholipid metabolism and sugar metabolism which inhibited nodule nitrogen fixation under P stress. More flavonoids were involved in metabolic processes in soybean root nodules under P stress that regulated the nodulation and nitrogen fixation. The pathway ascorbate and aldarate metabolism, and associated metabolites, were involved in the adaptation of the symbiotic soybean root nodule system to P starvation. This work provides a foundation for future investigations of physiological mechanisms that underly phosphorus stress on soybeans.

Phosphorus (P) is an essential elemental nutrient required in high abundance for robust soybean growth and development. Low P stress negatively impacts plant physiological and biochemical processes, such as photosynthesis, respiration, and energy transfer. Soybean roots play key roles in plant adaptive responses to P stress and other soil-related environmental stressors. Study the changes in soybean root exudates and differences in related metabolic pathways under low phosphorus stress, analyzing the response mechanism of soybean roots to phosphorus stress from the perspective of root exudates, which provide a theoretical basis for further analyzing the physiological mechanism of phosphorus stress on soybean. In this study, soybean roots were exposed to three phosphate levels: 1 mg/L (P stress), 11 mg/L (P stress) and 31 mg/L (Normal P) for 10 days and 20 days, then root exudates were analyzed via ultra-high-performance liquid chromatography-mass spectrometry to identify effects of P stress on root metabolite profiles and associated metabolic pathways. Our results revealed that with increasing P stress severity and/or duration, soybean roots produced altered types, quantities, and increased numbers of exudate metabolites (DMs in the P1 group were primarily upregulated, whereas those in the P11 group were predominately downregulated) caused by changes in regulation of activities of numerous metabolic pathways. These pathways had functions related to environmental adaptation, energy metabolism, and scavenging of reactive oxygen species and primarily included amino acid, flavonoid, and nicotinate and nicotinamide metabolic pathways and pathways related to isoquinoline alkaloid biosynthesis, sugar catabolism, and phospholipid metabolism. These metabolites and metabolic pathways lay a foundation to support further investigations of physiological mechanisms underlying the soybean root response to P deficiency.

In agroecosystems, a plant-usable form of nitrogen is mainly generated by legume-based biological nitrogen fixation, a process that requires phosphorus (P) as an essential nutrient. To investigate the physiological mechanism whereby phosphorus influences soybean nodule nitrogen fixation, soybean root nodules were exposed to four phosphate levels: 1 mg/L (P stress), 11 mg/L (P stress), 31 mg/L (Normal P), and 61 mg/L (High P) then proteome analysis of nodules was conducted to identify phosphorus-associated proteome changes. We found that phosphorus stress-induced ribosomal protein structural changes were associated with altered key root nodule protein synthesis profiles. Importantly, up-regulated expression of peroxidase was observed as an important phosphorus stress-induced nitrogen fixation-associated adaptation that supported two nodule-associated activities: scavenging of reactive oxygen species (ROS) and cell wall growth. In addition, phosphorus transporter (PT) and purple acid phosphatase (PAPs) were up-regulated that regulated phosphorus transport and utilization to maintain phosphorus balance and nitrogen fixation function in phosphorus-stressed root nodules.

Phosphorus is an indispensable nutrient for nitrogen metabolism in soybeans. In this study, two P levels were established, 1 mg/L (low-P stress) and 31 mg/L (normal P, CK), by combining ¹⁵N labeling with real-time quantitative PCR and the UHPLC-MS/MS method, to analyze soybean nitrogen accumulation, ¹⁵N abundance, nodule nitrogen fixation accumulation, nodule nitrogen fixation rate, soluble protein content, the relative expression of phosphorus transporters, amino acid changes, and metabolic pathways. The impacts of phosphorus stress on soybean nitrogen metabolism were explored from the perspectives of nitrogen accumulation and protein metabolism. The results demonstrated that low-P stress promoted the absorption of fertilizer nitrogen by aboveground parts, roots, and nodules of soybeans. However, it significantly inhibited nitrogen accumulation (11.09–95.41%), nodule nitrogen fixation accumulation (21.54–96.21%), and nodule nitrogen fixation rate (2.95–37.75%). The soluble protein content in both leaves and nodules decreased remarkably, while the relative expression of GmPT7 was upregulated in leaves, roots, and nodules under low-P stress. A total of 70 amino acids exhibited alterations, among which 26 amino acids were involved in 37 metabolic pathways, playing a crucial role in regulating the effects of low-P stress on soybean nitrogen metabolism. This study identifies significant alterations in nitrogen accumulation, nodule nitrogen fixation, and the expression of phosphorus transporter genes, providing insights into the metabolic pathways involved in soybean’s adaptation to phosphorus deficiency. This research provides a solid theoretical foundation for further in-depth investigations into the physiological and molecular mechanisms of soybean response to low-P stress.

Low phosphorus (P) stress impacts nitrogen (N) metabolism in soybeans. This study investigated the effects of exogenous cytokinin (Trans-Zeatin) on soybean N metabolism under low P stress by treating seeds with Trans-Zeatin and analyzing N accumulation, ¹⁵N abundance, nodule N fixation accumulation, nodule N fixation rate, nodule nitrogenase activity, soluble protein content, and free amino acid profiles. The results showed that exogenous cytokinin enhanced N accumulation in aboveground tissues, roots, and nodules, as well as nodule N fixation accumulation and fixation rate (from day 35 onward) under low P stress. Additionally, it promoted both acetylene reduction activity (ARA) and specific nitrogenase activity (SNA) in soybean nodules. By increasing the absorption of fertilizer-derived N, exogenous cytokinin alleviated the inhibitory effects of low P stress on the early growth and development of soybeans. Notably, under low P conditions, exogenous cytokinin significantly elevated the soluble protein content in nodules. However, the underlying mechanisms governing changes in free amino acid profiles require further investigation. This study provides a theoretical foundation for developing strategies to regulate soybean N metabolism under low P stress.

Soybean (Glycine max L.) is an essential food and economic crop in China, yet its growth and yield are severely constrained by saline–alkali stress. A saline–alkali soil exacerbates root absorption barriers, leading to 30–50% yield losses. Understanding the mechanisms underlying alkali tolerance is therefore crucial for developing stress-resilient soybean varieties and improving the productivity of saline–alkali land. In our previous study, we evaluated 99 soybean germplasms from Northeast China and obtained the alkali-tolerant varieties HN48 and HN69, along with the alkali-sensitive varieties HNWD4 and HN83. In this study, fifteen-day-old soybean seedlings were subjected to (30 mM NaHCO₃) alkali stress for 72 h, and whole plants were sampled to assess their morphology and physiology, while leaf tissues were harvested for biochemical analysis. For transcriptomic analysis, soybean seedlings were exposed to alkali stress (50 mM NaHCO₃, pH 9.0) for 6 h, and leaf and root tissues were harvested for RNA sequencing. The results showed that alkali-tolerant varieties mitigated these effects by suppressing excessive ROS generation by 55–63%, decreasing malondialdehyde (MDA) accumulation by 37–39%, and increasing photosynthetic efficiency by 18.3%, as well as accumulating more osmoprotectants and activating antioxidant enzymes such as superoxide dismutase (SOD) and catalase (CAT) under alkaline stress. Transcriptome analysis showed that the alkali-tolerant variety HN69 exhibited cultivar-specific enrichment of metabolism cytochrome P450, estrogen signaling, and GnRH signaling pathways under alkali stress. These results collectively indicate that alkali-tolerant soybean varieties adapt to alkali stress through coordinated multi-pathway responses, with differential pathway enrichment potentially underlying the variation in alkali tolerance between cultivars. Overall, this study elucidates the physiological and molecular mechanisms of alkali tolerance in soybean, providing a theoretical foundation for breeding stress-tolerant germplasms.