The rhizobium–legume symbiotic system is crucial for nitrogen cycle balance in agriculture. Hydrogen sulfide (H₂S), a gaseous signaling molecule, may regulate various physiological processes in plants. However, whether H₂S has regulatory effect in this symbiotic system remains unknown. Herein, we investigated the possible role of H₂S in the symbiosis between soybean (Glycine max) and rhizobium (Sinorhizobium fredii). Our results demonstrated that an exogenous H₂S donor (sodium hydrosulfide [NaHS]) treatment promoted soybean growth, nodulation, and nitrogenase (Nase) activity. Western blotting analysis revealed that the abundance of Nase component nifH was increased by NaHS treatment in nodules. Quantitative real-time polymerase chain reaction data showed that NaHS treatment upregulated the expressions of symbiosis-related genes nodA, nodC, and nodD of S. fredii. In addition, expression of soybean nodulation marker genes, including early nodulin 40 (GmENOD40), ERF required for nodulation (GmERN), nodulation signaling pathway 2b (GmNSP2b), and nodulation inception genes (GmNIN1a, GmNIN2a, and GmNIN2b), were upregulated. Moreover, the expressions of glutamate synthase (GmGOGAT), asparagine synthase (GmAS), nitrite reductase (GmNiR), ammonia transporter (GmSAT1), leghemoglobin (GmLb), and nifH involved in nitrogen metabolism were upregulated in NaHS-treated soybean roots and nodules. Together, our results suggested that H₂S may act as a positive signaling molecule in the soybean–rhizobia symbiotic system and enhance the system’s nitrogen fixation ability.

Background: Hydrogen sulphide (H₂S) is involved in regulating physiological processes in plants. We investigated how H₂S ameliorates iron (Fe) deficiency in soybean (Glycine max L.) seedlings. Multidisciplinary approaches including physiological, biochemical and molecular, and transcriptome methods were used to investigate the H₂S role in regulating Fe availability in soybean seedlings. Results: Our results showed that H₂S completely prevented leaf interveinal chlorosis and caused an increase in soybean seedling biomass under Fe deficiency conditions. Moreover, H₂S decreased the amount of root-bound apoplastic Fe and increased the Fe content in leaves and roots by regulating the ferric-chelate reductase (FCR) activities and Fe homeostasis- and sulphur metabolism-related gene expression levels, thereby promoting photosynthesis in soybean seedlings. In addition, H₂S changed the plant hormone concentrations by modulating plant hormone-related gene expression abundances in soybean seedlings grown in Fe-deficient solution. Furthermore, organic acid biosynthesis and related genes expression also played a vital role in modulating the H₂S-mediated alleviation of Fe deficiency in soybean seedlings. Conclusion: Our results indicated that Fe deficiency was alleviated by H₂S through enhancement of Fe acquisition and assimilation, thereby regulating plant hormones and organic acid synthesis in plants.