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Phospholipase C (PLC) performs significant functions in a variety of biological processes, including plant growth and development. The PLC family of enzymes principally catalyze the hydrolysis of phospholipids in organisms. This exhaustive exploration of soybean GmPLC members using genome databases resulted in the identification of 15 phosphatidylinositol-specific PLC (GmPI-PLC) and 9 phosphatidylcholine-hydrolyzing PLC (GmNPC) genes. Chromosomal location analysis indicated that GmPLC genes mapped to 10 of the 20 soybean chromosomes. Phylogenetic relationship analysis revealed that GmPLC genes distributed into two groups in soybean, the PI-PLC and NPC groups. The expression patterns and tissue expression analysis showed that GmPLCs were differentially expressed in response to abiotic stresses. GmPI-PLC7 was selected to further explore the role of PLC in soybean response to drought and salt stresses by a series of experiments. Compared with the transgenic empty vector (EV) control lines, over-expression of GmPI-PLC7 (OE) conferred higher drought and salt tolerance in soybean, while the GmPI-PLC7-RNAi (RNAi) lines exhibited the opposite phenotypes. Plant tissue staining and physiological parameters observed from drought- and salt-stressed plants showed that stress increased the contents of chlorophyll, oxygen free radical (O2–), hydrogen peroxide (H2O2) and NADH oxidase (NOX) to amounts higher than those observed in non-stressed plants. This study provides new insights in the functional analysis of GmPLC genes in response to abiotic stresses.

GDSL-type esterase/lipase proteins (GELPs) belong to the SGNH hydrolase superfamily and contain a conserved GDSL motif at their N-terminus. GELPs are widely distributed in nature, from microbes to plants, and play crucial roles in growth and development, stress responses and pathogen defense. However, the identification and functional analysis of GELP genes are hardly explored in soybean. This study describes the identification of 194 GELP genes in the soybean genome and their phylogenetic classification into 11 subfamilies (A–K). GmGELP genes are disproportionally distributed on 20 soybean chromosomes. Large-scale WGD/segmental duplication events contribute greatly to the expansion of the soybean GDSL gene family. The Ka/Ks ratios of more than 70% of duplicated gene pairs ranged from 0.1–0.3, indicating that most GmGELP genes were under purifying selection pressure. Gene structure analysis indicate that more than 74% of GmGELP genes are interrupted by 4 introns and composed of 5 exons in their coding regions, and closer homologous genes in the phylogenetic tree often have similar exon-intron organization. Further statistics revealed that approximately 56% of subfamily K members contain more than 4 introns, and about 28% of subfamily I members consist of less than 4 introns. For this reason, the two subfamilies were used to simulate intron gain and loss events, respectively. Furthermore, a new model of intron position distribution was established in current study to explore whether the evolution of multi-gene families resulted from the diversity of gene structure. Finally, RNA-seq data were used to investigate the expression profiles of GmGELP gene under different tissues and multiple abiotic stress treatments. Subsequently, 7 stress-responsive GmGELP genes were selected to verify their expression levels by RT-qPCR, the results were consistent with RNA-seq data. Among 7 GmGELP genes, GmGELP28 was selected for further study owing to clear responses to drought, salt and ABA treatments. Transgenic Arabidopsis thaliana and soybean plants showed drought and salt tolerant phenotype. Overexpression of GmGELP28 resulted in the changes of several physiological indicators, which allowed plants to adapt adverse conditions. In all, GmGELP28 is a potential candidate gene for improving the salinity and drought tolerance of soybean.

Transcription factors play vital roles in plant growth and in plant responses to abiotic stresses. The RAV transcription factors contain a B3 DNA binding domain and/or an APETALA2 (AP2) DNA binding domain. Although genome-wide analyses of RAV family genes have been performed in several species, little is known about the family in soybean (Glycine max L.). In this study, a total of 13 RAV genes, named as GmRAVs, were identified in the soybean genome. We predicted and analyzed the amino acid compositions, phylogenetic relationships, and folding states of conserved domain sequences of soybean RAV transcription factors. These soybean RAV transcription factors were phylogenetically clustered into three classes based on their amino acid sequences. Subcellular localization analysis revealed that the soybean RAV proteins were located in the nucleus. The expression patterns of 13 RAV genes were analyzed by quantitative real-time PCR. Under drought stresses, the RAV genes expressed diversely, up- or down-regulated. Following NaCl treatments, all RAV genes were down-regulated excepting GmRAV-03 which was up-regulated. Under abscisic acid (ABA) treatment, the expression of all of the soybean RAV genes increased dramatically. These results suggested that the soybean RAV genes may be involved in diverse signaling pathways and may be responsive to abiotic stresses and exogenous ABA. Further analysis indicated that GmRAV-03 could increase the transgenic lines resistance to high salt and drought and result in the transgenic plants insensitive to exogenous ABA. This present study provides valuable information for understanding the classification and putative functions of the RAV transcription factors in soybean.

Fructose-1, 6-bisphosphate aldolase (FBA) is a key plant enzyme that is involved in glycolysis, gluconeogenesis, and the Calvin cycle. It plays significant roles in biotic and abiotic stress responses, as well as in regulating growth and development processes. In the present paper, 21 genes encoding TaFBA isoenzymes were identified, characterized, and categorized into three groups: class I chloroplast/plastid FBA (CpFBA), class I cytosol FBA (cFBA), and class II chloroplast/plastid FBA. By using a prediction online database and genomic PCR analysis of Chinese Spring nulli-tetrasomic lines, we have confirmed the chromosomal location of these genes in 12 chromosomes of four homologous groups. Sequence and genomic structure analysis revealed the high identity of the allelic TaFBA genes and the origin of different TaFBA genes. Numerous putative environment stimulus-responsive cis-elements have been identified in 1,500-bp regions of TaFBA gene promoters, of which the most abundant are the light-regulated elements (LREs). Phylogenetic reconstruction using the deduced protein sequence of 245 FBA genes indicated an independent evolutionary pathway for the class I and class II groups. Although, earlier studies have indicated that class II FBA only occurs in prokaryote and fungi, our results have demonstrated that a few class II CpFBAs exist in wheat and other closely related species. Class I TaFBA was predicted to be tetramers and class II to be dimers. Gene expression analysis based on microarray and transcriptome databases suggested the distinct role of TaFBAs in different tissues and developmental stages. The TaFBA 4–9 genes were highly expressed in leaves and might play important roles in wheat development. The differential expression patterns of the TaFBA genes in light/dark and a few abiotic stress conditions were also analyzed. The results suggested that LRE cis-elements of TaFBA gene promoters were not directly related to light responses. Most TaFBA genes had higher expression levels in the roots than in the shoots when under various stresses. Class I cytosol TaFBA genes, particularly TaFBA10/12/18 and TaFBA13/16, and three class II TaFBA genes are involved in responses to various abiotic stresses. Class I CpFBA genes in wheat are apparently sensitive to different stress conditions.