Salt and drought stresses are two primary abiotic stresses that inhibit growth and reduce the activity of photosynthetic apparatus in plants. Abscisic acid (ABA) plays a key role in abiotic stress regulation in plants. Some aldo–keto reductases (AKRs) can enhance various abiotic stresses resistance by scavenging cytotoxic aldehydes in some plants. However, there are few comprehensive reports of plant AKR genes and their expression patterns in response to abiotic stresses. In this study, we identified 30 putative AKR genes from Medicago truncatula. The gene characteristics, coding protein motifs, and expression patterns of these MtAKRs were analyzed to explore and identify candidate genes in regulation of salt, drought, and ABA stresses. The phylogenetic analysis result indicated that the 52 AKRs in Medicago truncatula and Arabidopsis thaliana can be divided into three groups and six subgroups. Fifteen AKR genes in M. truncatula were randomly selected from each group or subgroup, to investigate their response to salt (200 mM of NaCl), drought (50 g·L⁻¹ of PEG 6000), and ABA (100 µM) stresses in both leaves and roots. The results suggest that MtAKR1, MtAKR5, MtAKR11, MtAKR14, MtAKR20, and MtAKR29 may play important roles in response to these stresses.

Bioactive gibberellic acids (GAs) are diterpenoid plant hormones that are biosynthesized through complex pathways and control various aspects of growth and development. Although GA biosynthesis has been intensively studied, the downstream metabolic pathways regulated by GAs have remained largely unexplored. We investigated Tnt1 retrotransposon insertion mutant lines of Medicago truncatula with a dwarf phenotype by forward and reverse genetics screening and phylogenetic, molecular, biochemical, proteomic and metabolomic analyses. Three Tnt1 retrotransposon insertion mutant lines of the gibberellin 3-beta-dioxygenase 1 gene (GA3ox1) with a dwarf phenotype were identified, in which the synthesis of GAs (GA₃ and GA₄) was inhibited. Phenotypic analysis revealed that plant height, root and petiole length of ga3ox1 mutants were shorter than those of the wild type (Medicago truncatula ecotype R108). Leaf size was also much smaller in ga3ox1 mutants than that in wild-type R108, which is probably due to cell-size diminution instead of a decrease in cell number. Proteomic and metabolomic analyses of ga3ox1/R108 leaves revealed that in the ga3ox1 mutant, flavonoid isoflavonoid biosynthesis was significantly up-regulated, while nitrogen metabolism was down-regulated. Additionally, we further demonstrated that flavonoid and isoflavonoid biosynthesis was induced by prohexadione calcium, an inhibitor of GA3ox enzyme, and inhibited by exogenous GA₃. In contrast, nitrogen metabolism was promoted by exogenous GA₃ but inhibited by prohexadione calcium. The results of this study further demonstrated that GAs play critical roles in positively regulating nitrogen metabolism and transport and negatively regulating flavonoid biosynthesis through GA-mediated signaling pathways in leaves.

Saline-alkaline stress is a universal abiotic stress that adversely affects plant growth and productivity. Saline-alkaline conditions results in plant abnormal transcriptome expression finally manifesting as defective phenotypes. Considerable research has revealed the active role of microRNA in various stress conditions. This study was aimed to identify novel miRNAs and the miRNA expression patterns in the leguminous model plant R108 (Medicago truncatula). The miRNA contained in the total RNA extracted from Medicago truncatula seedlings (72 h) that had been treated with solutions mimicking saline and alkaline soils was subjected to miRNA deep sequencing. The Illumina HiSeq sequencing platform was used to analyze nine small RNA libraries of three treatment groups: distilled water, 20 mM NaCl + Na₂SO₄ and 5 mM Na₂CO₃ + NaHCO₃. Sequencing revealed that 876 miRNAs including 664 known miRNAs and 212 potential novel miRNAs were present in all the libraries. The miR159 family, miR156 family, miR2086-3p, miR396, miR166, miR319, miR167, miR5213-5p, miR1510 and miR2643 were among the most expressed miRNAs in all libraries. The results of miRNAs expression under treatments were validated by reverse-transcription quantitative PCR (RT-qPCR). Target gene prediction through computational analysis and pathway annotation analysis revealed that the primary pathways affected by stress were related to plant development, including metabolic processes, single-organism processes and response to the stimulus. Our results provide valuable information towards elucidating the molecular mechanisms of salt/alkali tolerance in Medicago truncatula and provide insight into the putative role of miRNAs in plant stress resistance.

Drought stress is a predominant abiotic factor contributing to reduced crop yields globally. Therefore, exploring the molecular mechanism of drought control is of great significance to improve drought resistance and ultimately achieve crop yield increase. As a plant endogenous hormone, melatonin plays a key role in the regulation of abiotic stress, but the key genes and metabolic pathways of melatonin mediated drought resistance regulation in alfalfa have not been fully revealed. Based on transcriptomics and physiological index detection, this study aimed to explore the regulatory mechanism of melatonin in alleviating drought stress during alfalfa germination. The findings revealed that alfalfa seedlings treated with melatonin exhibited higher germination rates, increased shoot length, and greater fresh weight compared to those exposed solely to drought stress. Additionally, there was a reduction in the levels of malondialdehyde (MDA) and superoxide anion (O2−), while the activity and concentration of superoxide dismutase (SOD), peroxidase (POD), and glutathione (GSH) were enhanced to varying extents. To investigate the molecular mechanism underlying melatonin-mediated drought resistance in alfalfa, we performed a transcriptomic analysis on the seedlings. In the drought treatment group, we identified a total of 1,991 differentially expressed genes (DEGs), comprising 778 up-regulated and 1,213 down-regulated genes. Conversely, in the melatonin-treated group, we discovered 2,336 DEGs, including 882 up-regulated and 1,454 down-regulated genes. Through the application of GO functional annotation and KEGG pathway enrichment analysis, we discovered that DEGs were predominantly enriched in pathways related to flavonoid and isoflavone biosynthesis, plant hormone biosynthesis and signal transduction, glutathione metabolism, and MAPK signaling, and the ABC transporter signaling. Notably, the DEGs added to the MT group showed greater enrichment in these pathways. This suggests that MT mitigates drought stress by modulating the expression of genes associated with energy supply and antioxidant capacity. These findings hold significant reference value for breeding drought-tolerant alfalfa and other crops.