The faba bean, a significant cool-season edible legume crop, is susceptible to drought during the germination stage. Research regarding the genetic regulation of drought tolerance throughout this stage in the faba bean is limited. The differentially expressed proteins (DEPs) in faba beans between the drought-tolerant variety C105 and the drought-sensitive variant E1 during seed germination were identified in this work, accomplished through isobaric tags for relative and absolute quantitation (iTRAQ) analysis. A total of 3827 proteins were identified in the two varieties of germinating seeds. Compared to those of variety E1, an increase in 108 DEPs and a decrease in 61 DEPs were observed in variety C105 under drought. Conversely, in the control group, variety C105 showed 108 significantly upregulated DEPs and 55 significantly downregulated DEPs. GO and KEGG analyses showed that the DEPs associated with glutathione metabolism and protein processing demonstrated significant increases in response to drought stress. Protein–protein interaction (PPI) analysis unveiled three closely connected functional modules of protein translation, DNA replication, and post-translational modification, originating from 22 DEPs derived from the germination period of two varieties under drought stress. To verify the proteomic function, we selected three differentially expressed protein coding genes, which were overexpressed or silenced in tobacco, thereby enhancing the drought resistance of tobacco. This was accompanied via altered levels of superoxide dismutase or peroxidase in transgenic plants under drought stress. The possible mechanism for drought tolerance in germinating seeds of faba bean involves increasing protein translation, decreasing DNA replication, and modifying chromatin. These findings offer invaluable insights into the reaction mechanism in response to drought stress in faba beans. The identified DEPs could be utilized in faba bean breeding initiatives to manage drought.

Faba bean is an important pulse. It provides proteins for the human diet and is used in industrial foodstuffs, such as flours. Drought stress severely reduces the yield of faba bean, and this can be efficiently overcome through the identification and application of key genes in response to drought. In this study, PacBio and Illumina RNA sequencing techniques were used to identify the key pathways and candidate genes involved in drought stress response. During seed germination, a total of 17,927 full-length transcripts and 12,760 protein-coding genes were obtained. There were 1676 and 811 differentially expressed genes (DEGs) between the varieties E1 and C105 at 16 h and 64 h under drought stress, respectively. Six and nine KEGG pathways were significantly enriched at 16 h and 64 h under drought stress, which produced 40 and 184 nodes through protein–protein interaction (PPI) analysis, respectively. The DEGs of the PPI nodes were involved in the ABA (abscisic acid) and MAPK (mitogen-activated protein kinase) pathways, N-glycosylation, sulfur metabolism, and sugar metabolism. Furthermore, the ectopic overexpression of a key gene, AAT, encoding aspartate aminotransferase (AAT), in tobacco, enhanced drought tolerance. The activities of AAT and peroxidase (POD), the contents of cysteine and isoleucine, were increased, and the contents of malonaldehyde (MDA) and water loss decreased in the overexpressed plants. This study provides a novel insight into genetic response to drought stress and some candidate genes for drought tolerance genetic improvements in this plant.

Crop productivity is severely constrained by abiotic and biotic stresses, necessitating innovative strategies to enhance stress resilience. Glycerol-3-phosphate (G3P) is a central metabolite in carbohydrate and lipid metabolism, playing crucial roles in stress responses. In this study, we engineered a novel glycerol-3-phosphate dehydrogenase (GPDH) gene, designated OEGD, by fusing the N-terminal NAD-binding domain of rice OsGPDH1 with the feedback-resistant C-terminal catalytic domain of Escherichia coli gpsA. Overexpression of OEGD in rice enhanced tolerance to drought, phosphorus deficiency, high temperature, and cadmium (Cd²⁺) stresses, while also improving plant growth and yield under drought stress at the adult stage. Notably, the accumulation of glycerol-3-phosphate (G3P) and activities of antioxidant enzymes (SOD, POD, CAT) were significantly elevated in the transgenic plants following osmotic stimuli, and fatty acid profiles were altered, favoring stress adaptation. Transcriptomic analyses revealed that OEGD modulates cell wall biogenesis, reactive oxygen species (ROS) scavenging, and lipid metabolism pathways, with minimal disruption to core G3P metabolic genes. These findings highlight the potential of OEGD as a valuable genetic resource for improving stress resistance in rice.