The NF-Y-PYR module integrates the abscisic acid signal pathway to regulate plant stress tolerance.

Drought and salt stresses impose major constraints on soybean production worldwide. However, improving agronomically valuable soybean traits under drought conditions can be challenging due to trait complexity and multiple factors that influence yield. Here, we identified a nuclear factor Y C subunit (NF-YC) family transcription factor member, GmNF-YC14, which formed a heterotrimer with GmNF-YA16 and GmNF-YB2 to activate the GmPYR1-mediated abscisic acid (ABA) signalling pathway to regulate stress tolerance in soybean. Notably, we found that CRISPR/Cas9-generated GmNF-YC14 knockout mutants were more sensitive to drought than wild-type soybean plants. Furthermore, field trials showed that overexpression of GmNF-YC14 or GmPYR1 could increase yield per plant, grain plumpness, and stem base circumference, thus indicating improved adaptation of soybean plants to drought conditions. Taken together, our findings expand the known functional scope of the NF-Y transcription factor functions and raise important questions about the integration of ABA signalling pathways in plants. Moreover, GmNF-YC14 and GmPYR1 have potential for application in the improvement of drought tolerance in soybean plants.

Functional Analysis of the Soybean GmCDPK3 Gene Responding to Drought and Salt Stresses.

Plants have a series of response mechanisms to adapt when they are subjected to external stress. Calcium-dependent protein kinases (CDPKs) in plants function against a variety of abiotic stresses. We screened 17 CDPKs from drought- and salt-induced soybean transcriptome sequences. The phylogenetic tree divided CDPKs of rice, Arabidopsis and soybean into five groups (I-V). Cis-acting element analysis showed that the 17 CDPKs contained some elements associated with drought and salt stresses. Quantitative real-time PCR (qRT-PCR) analysis indicated that the 17 CDPKs were responsive after different degrees of induction under drought and salt stresses. GmCDPK3 was selected as a further research target due to its high relative expression. The subcellular localization experiment showed that GmCDPK3 was located on the membrane of Arabidopsis mesophyll protoplasts. Overexpression of GmCDPK3 improved drought and salt resistance in Arabidopsis. In the soybean hairy roots experiment, the leaves of GmCDPK3 hairy roots with RNA interference (GmCDPK3-RNAi) soybean lines were more wilted than those of GmCDPK3 overexpression (GmCDPK3-OE) soybean lines after drought and salt stresses. The trypan blue staining experiment further confirmed that cell membrane damage of GmCDPK3-RNAi soybean leaves was more severe than in GmCDPK3-OE soybean lines. In addition, proline (Pro) and chlorophyll contents were increased and malondialdehyde (MDA) content was decreased in GmCDPK3-OE soybean lines. On the contrary, GmCDPK3-RNAi soybean lines had decreased Pro and chlorophyll content and increased MDA. The results indicate that GmCDPK3 is essential in resisting drought and salt stresses.

Genome-Wide Analysis of the C2 Domain Family in Soybean and Identification of a Putative Abiotic Stress Response Gene GmC2-148.

Plant C2 domain proteins play essential biological functions in numerous plants. In this study, 180 soybean C2 domain genes were identified by screening. Phylogenetic relationship analysis revealed that C2 domain genes fell into three distinct groups with diverged gene structure and conserved functional domain. Chromosomal location analysis indicated that C2 domain genes mapped to 20 chromosomes. The transcript profiles based on RNA-seq data showed that GmC2-58, GmC2-88, and GmC2-148 had higher levels of expression under salt, drought, and abscisic acid (ABA) treatments. GmC2-148, encoding a cell membrane-localized protein, had the highest level of response to various treatments according to real-time quantitative polymerase chain reaction (RT-qPCR) analysis. Under salt and drought stresses, the soybean plants with GmC2-148 transgenic hairy roots showed delayed leaf rolling, a higher content of proline (Pro), and lower contents of H2O2, O2- and malondialdehyde (MDA) compared to those of the empty vector (EV) plants. The results of transgenic Arabidopsis in salt and drought treatments were consistent with those in soybean treatments. In addition, the soybean plants with GmC2-148 transgenic hairy roots increased transcript levels of several abiotic stress-related marker genes, including COR47, NCDE3, NAC11, WRKY13, DREB2A, MYB84, bZIP44, and KIN1 which resulted in enhanced abiotic stress tolerance in soybean. These results indicate that C2 domain genes are involved in response to salt and drought stresses, and this study provides a genome-wide analysis of the C2 domain family in soybean.

The Ankyrin-Repeat Gene GmANK114 Confers Drought and Salt Tolerance in Arabidopsis and Soybean.

Ankyrin repeat (ANK) proteins are essential in cell growth, development, and response to hormones and environmental stresses. In the present study, 226 ANK genes were identified and classified into nine subfamilies according to conserved domains in the soybean genome (Glycine max L.). Among them, the GmANK114 was highly induced by drought, salt, and abscisic acid. The GmANK114 encodes a protein that belongs to the ANK-RF subfamily containing a RING finger (RF) domain in addition to the ankyrin repeats. Heterologous overexpression of GmANK114 in transgenic Arabidopsis improved the germination rate under drought and salt treatments compared to wild-type. Homologous overexpression of GmANK114 improved the survival rate under drought and salt stresses in transgenic soybean hairy roots. In response to drought or salt stress, GmANK114 overexpression in soybean hairy root showed higher proline and lower malondialdehyde contents, and lower H2O2 and O2- contents compared control plants. Besides, GmANK114 activated transcription of several abiotic stress-related genes, including WRKY13, NAC11, DREB2, MYB84, and bZIP44 under drought and salt stresses in soybean. These results provide new insights for functional analysis of soybean ANK proteins and will be helpful for further understanding how ANK proteins in plants adapt to abiotic stress.

Rapid identification of a stripe rust resistant gene in a space-induced wheat mutant using specific locus amplified fragment (SLAF) sequencing.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most devastating diseases of wheat. Resistant cultivars are the preferred strategy to control the disease. Space-induced wheat mutant R39 has adult-plant resistance (APR) to Pst. Genetic analysis indicated that a single recessive gene, designated YrR39, was responsible for the APR of R39 to Pst. Bulked segregant analysis (BSA) combined with a SLAF sequencing (SLAF-seq) strategy was used to fine-map YrR39 to a 17.39 Mb segment on chromosome 4B. The region was confirmed by analysis with simple sequence repeat (SSR) markers. A total of 126 genes were annotated in the region and 21 genes with annotations associated with disease response were selected for further qRT-PCR analysis. The candidate gene Traes_4BS_C868349E1 (annotated as an F-box/LRR-repeat protein) was up-regulated after 12, 24, 48, and 96 hours post inoculation with Pst, suggesting it is likely involved in the resistance. The current study demonstrated that BSA combined with SLAF-seq for SNP discovery is an efficient approach for mapping and identifying candidate functional gene.

Ectopic expression of FaesAP3, a Fagopyrum esculentum (Polygonaceae) AP3 orthologous gene rescues stamen development in an Arabidopsis ap3 mutant.

Arabidopsis thaliana APETALA3 (AP3) and Antirrhinum majus DEFICIENS (DEF) MADS box genes are required to specify petal and stamen identity. AP3 and DEF are members of the euAP3 lineage, which arose by gene duplication coincident with radiation of the core eudicots. In order to investigate the molecular mechanisms underlying organ development in early diverging clades of core eudicots, we isolated and identified an AP3 homolog, FaesAP3, from Fagopyrum esculentum (buckwheat, Polygonaceae), a multi-food-use pseudocereal with healing benefits. Protein sequence alignment and phylogenetic analyses revealed that FaesAP3 grouped into the euAP3 lineage. Expression analysis showed that FaesAP3 was transcribed only in developing stamens, and differed from AP3 and DEF, which expressed in developing petals and stamens. Moreover, ectopic expression of FaesAP3 rescued stamen development without complementation of petal development in an Arabidopsis ap3 mutant. Our results suggest that FaesAP3 is involved in the development of stamens in buckwheat. These results also suggest that FaesAP3 holds some potential for biotechnical engineering to create a male sterile line of F. esculentum.