Genetic variation in GmCRP contributes to nodulation in soybean(Glycine max Merr.)

Symbiosis between soybean and rhizobia contributes to soybean yield and quality. Although secreted rhizobial type Ⅲ effectors are known to regulate infection and promote nitrogen fixation, much about them remains unknown. Mutation of NopC, a type Ⅲ effector from Sinorhizobium fredii HH103, reduced nodule numbers and dry weights in 310 soybean accessions, and expression of NopC in soybean hairy roots promoted symbiosis. Based on observed differences in nodule traits between Suinong 14 and Zyd 00,006inoculated with HH103 and the NopC mutant, 11 QTL associated with rhizobia were identified in chromosome segment substitution lines(CSSLs) derived from Suinong 14 and Zyd 00006. Using chromosome fragment insertion, whole-genome sequencing of Suinong 14 and Zyd 00006, and qRT-PCR,Glyma.19G176300(GmCRP) was identified as a candidate gene associated with NopC, and GmCRP was found to be induced by NopC to positively regulate nodulation. SNPs located in the regulatory regions of GmCRP influenced its expression response to NopC, with SNPs contributing to nodulation having been selected during domestication. Our findings reveal the function of a soybean gene encoding a rhizobial type Ⅲ effector that contributes to symbiosis, and will facilitate the practical application of symbiotic nitrogen fixation in molecular breeding.

SPM-IS: An auto-algorithm to acquire a mature soybean phenotype based on instance segmentation

Mature soybean phenotyping is an important process in soybean breeding;however, the manual process is time-consuming and labor-intensive. Therefore, a novel approach that is rapid, accurate and highly precise is required to obtain the phenotypic data of soybean stems, pods and seeds. In this research, we propose a mature soybean phenotype measurement algorithm called Soybean Phenotype Measure-instance Segmentation(SPM-IS). SPM-IS is based on a feature pyramid network, Principal Component Analysis(PCA) and instance segmentation. We also propose a new method that uses PCA to locate and measure the length and width of a target object via image instance segmentation. After 60,000 iterations, the maximum mean Average Precision(m AP) of the mask and box was able to reach 95.7%. The correlation coefficients R^(2) of the manual measurement and SPM-IS measurement of the pod length, pod width, stem length, complete main stem length, seed length and seed width were 0.9755, 0.9872, 0.9692, 0.9803,0.9656, and 0.9716, respectively. The correlation coefficients R^(2) of the manual counting and SPM-IS counting of pods, stems and seeds were 0.9733, 0.9872, and 0.9851, respectively. The above results show that SPM-IS is a robust measurement and counting algorithm that can reduce labor intensity, improve efficiency and speed up the soybean breeding process.

GmGPDH12,a mitochondrial FAD-GPDH from soybean, increases salt and osmotic stress resistance by modulating redox state and respiration

In plants,glycerol-3-phosphate dehydrogenase(GPDH)catalyzes the interconversion of glycerol-3-phosphate(G3P)and dihydroxyacetone phosphate(DHAP)coupled to the reduction/oxidation of the nicotinamide adenine dinucleotide(NADH)pool,and plays a central role in glycerolipid metabolism and stress response.Previous studies have focused mainly on the NAD+-dependent GPDH isoforms,neglecting the role of flavin adenine dinucleotide(FAD)-dependent GPDHs.We isolated and characterized three mitochondrialtargeted FAD-GPDHs in soybean,of which one isoform(GmGPDH12)showed a significant transcriptional response to NaCl and mannitol treatments,suggesting the existence of a major FAD-GPDH isoform acting in soybean responses to salt and osmotic stress.An enzyme kinetic assay showed that the purified GmGPDH12 protein possessed the capacity to oxidize G3P to DHAP in the presence of FAD.Overexpression and RNA interference of GmGPDH12 in soybean hairy roots resulted in elevated tolerance and sensitivity to salt and osmotic stress,respectively.G3P contents were significantly lower in GmGPDH12-overexpressing hair roots and higher in knockdown hair roots,indicating that GmGPDH12 was essential for G3P catabolism.A significant perturbation in redox status of NADH,ascorbic acid(ASA)and glutathione(GSH)pools was observed in GmGPDH12-knockdown plants under stress conditions.The impaired redox balance was manifested by higher reactive oxygen species generation and consequent cell damage or death;however,overexpressing plants showed the opposite results for these traits.GmGPDH12 overexpression contributed to maintaining constant respiration rates under salt or osmotic stress by regulating mRNA levels of key mitochondrial respiratory enzymes.This study provides new evidence for the roles of mitochondria-localized GmGPDH12 in conferring resistance to salt or osmotic stress by maintaining cellular redox homeostasis,protecting cells and respiration from oxidative injury.

A soybean NAC homolog contributes to resistance to Phytophthora sojae mediated by dirigent proteins

Phytophthora sojae infection severely impairs soybean production. We previously identified a dirigent protein, Gm DRR1(Glycine max Disease Resistant Response 1), that increases soybean resistance to P.sojae. However, the molecular basis of Gm DRR1 function remained largely uncharacterized. In the present study, analysis of Gm DRR1-RNAi, Gm DRR1-overexpressing, and CRISPR/Cas9-derived Gmdrr1 mutant lines revealed that Gm DRR1 expression significantly restricted P. sojae growth. Combining coimmunoprecipitation with liquid chromatography–tandem mass spectrometry revealed a Gm DRR1-interacting protein, Gm DRR2, which is homologous to Gm DRR1. An E-coniferyl alcohol coupling assay indicated that Gm DRR1 promotes the synthesis of(+)-pinoresinol, which helps to protect plants from P. sojae. The Gm NAC1(Glyma.05 G025500) transcription factor bound to the Gm DRR1 promoter both in vitro and in vivo to upregulate Gm DRR1 expression. Soybean resistance to P. sojae was increased by overexpression of Gm NAC1. Our findings suggest a novel signaling pathway involving a NAC transcription factor that mediates soybean resistance to P. sojae. Specifically, Gm NAC1 directly induces Gm DRR1 expression to increase resistance of soybean plants to P. sojae.

Enhanced production of seed oil with improved fatty acid composition by overexpressing NAD^(+)-dependent glycerol-3-phosphate dehydrogenase in soybean

There is growing interest in expanding the production of soybean oils(mainly triacylglycerol, or TAG) to meet rising feed demand and address global energy concerns. We report that a plastidlocalized glycerol-3-phosphate dehydrogenase(GPDH), encoded by GmGPDHp1 gene, catalyzes the formation of glycerol-3-phosphate(G3 P), an obligate substrate required for TAG biosynthesis.Overexpression of GmGPDHp1 increases soybean seed oil content with high levels of unsaturated fatty acids(FAs), especially oleic acid(C18:1), without detectably affecting growth or seed protein content or seed weight. Based on the lipidomic analyses, we found that the increase in G3 P content led to an elevated diacylglycerol(DAG) pool, in which the Kennedy pathwayderived DAG was mostly increased, followed by PC-derived DAG, thereby promoting the synthesis of TAG containing relatively high proportion of C18:1. The increased G3 P levels induced several transcriptional alterations of genes involved in the glycerolipid pathways. In particular, genes encoding the enzymes responsible for de novo glycerolipid synthesis were largely upregulated in the transgenic lines, in-line with the identified biochemical phenotype. These results reveal a key role for GmGPDHp1-mediated G3 P metabolism in enhancing TAG synthesis and demonstrate a strategy to modify the FA compositions of soybean oils for improved nutrition and biofuel.