The MYB Transcription Factor GmMYB78 Negatively Regulates Phytophthora sojae Resistance in Soybean.

Phytophthora root rot is a devastating disease of soybean caused by Phytophthora sojae. However, the resistance mechanism is not yet clear. Our previous studies have shown that GmAP2 enhances sensitivity to P. sojae in soybean, and GmMYB78 is downregulated in the transcriptome analysis of GmAP2-overexpressing transgenic hairy roots. Here, GmMYB78 was significantly induced by P. sojae in susceptible soybean, and the overexpressing of GmMYB78 enhanced sensitivity to the pathogen, while silencing GmMYB78 enhances resistance to P. sojae, indicating that GmMYB78 is a negative regulator of P. sojae. Moreover, the jasmonic acid (JA) content and JA synthesis gene GmAOS1 was highly upregulated in GmMYB78-silencing roots and highly downregulated in overexpressing ones, suggesting that GmMYB78 could respond to P. sojae through the JA signaling pathway. Furthermore, the expression of several pathogenesis-related genes was significantly lower in GmMYB78-overexpressing roots and higher in GmMYB78-silencing ones. Additionally, we screened and identified the upstream regulator GmbHLH122 and downstream target gene GmbZIP25 of GmMYB78. GmbHLH122 was highly induced by P. sojae and could inhibit GmMYB78 expression in resistant soybean, and GmMYB78 was highly expressed to activate downstream target gene GmbZIP25 transcription in susceptible soybean. In conclusion, our data reveal that GmMYB78 triggers soybean sensitivity to P. sojae by inhibiting the JA signaling pathway and the expression of pathogenesis-related genes or through the effects of the GmbHLH122-GmMYB78-GmbZIP25 cascade pathway.

GmMKK4-activated GmMPK6 stimulates GmERF113 to trigger resistance to Phytophthora sojae in soybean.

Phytophthora root and stem rot is a worldwide soybean (Glycine max) disease caused by the soil-borne pathogen Phytophthora sojae. This disease is devastating to soybean production, so improvement of resistance to P. sojae is a major target in soybean breeding. Mitogen-activated protein kinase (MAPK) cascades are important signaling modules that convert environmental stimuli into cellular responses. Compared with extensive studies in Arabidopsis, the molecular mechanism of MAPK cascades in soybean disease resistance is barely elucidated. In this work, we found that the gene expression of mitogen-activated protein kinase 6 (GmMPK6) was potently induced by P. sojae infection in the disease-resistant soybean cultivar 'Suinong 10'. Overexpression of GmMPK6 in soybean resulted in enhanced resistance to P. sojae and silencing of GmMPK6 led to the opposite phenotype. In our attempt to dissect the role of GmMPK6 in soybean resistance to phytophthora disease, we found that MAPK kinase 4 (GmMKK4) and the ERF transcription factor GmERF113 physically interact with GmMPK6, and we determined that GmMKK4 could phosphorylate and activate GmMPK6, which could subsequently phosphorylate GmERF113 upon P. sojae infection, suggesting that P. sojae can stimulate the GmMKK4-GmMPK6-GmERF113 signaling pathway in soybean. Moreover, phosphorylation of GmERF113 by the GmMKK4-GmMPK6 module promoted GmERF113 stability, nuclear localization and transcriptional activity, which significantly enhanced expression of the defense-related genes GmPR1 and GmPR10-1 and hence improved disease resistance of the transgenic soybean seedlings. In all, our data reveal that the GmMKK4-GmMPK6-GmERF113 cascade triggers resistance to P. sojae in soybean and shed light on functions of MAPK kinases in plant disease resistance.

GmWAK1, Novel Wall-Associated Protein Kinase, Positively Regulates Response of Soybean to Phytophthora sojae Infection.

Phytophthora root rot is a destructive soybean disease worldwide, which is caused by the oomycete pathogen Phytophthora sojae (P. sojae). Wall-associated protein kinase (WAK) genes, a family of the receptor-like protein kinase (RLK) genes, play important roles in the plant signaling pathways that regulate stress responses and pathogen resistance. In our study, we found a putative Glycine max wall-associated protein kinase, GmWAK1, which we identified by soybean GmLHP1 RNA-sequencing. The expression of GmWAK1 was significantly increased by P. sojae and salicylic acid (SA). Overexpression of GmWAK1 in soybean significantly improved resistance to P. sojae, and the levels of phenylalanine ammonia-lyase (PAL), SA, and SA-biosynthesis-related genes were markedly higher than in the wild-type (WT) soybean. The activities of enzymatic superoxide dismutase (SOD) and peroxidase (POD) antioxidants in GmWAK1-overexpressing (OE) plants were significantly higher than those in in WT plants treated with P. sojae; reactive oxygen species (ROS) and hydrogen peroxide (H2O2) accumulation was considerably lower in GmWAK1-OE after P. sojae infection. GmWAK1 interacted with annexin-like protein RJ, GmANNRJ4, which improved resistance to P. sojae and increased intracellular free-calcium accumulation. In GmANNRJ4-OE transgenic soybean, the calmodulin-dependent kinase gene GmMPK6 and several pathogenesis-related (PR) genes were constitutively activated. Collectively, these results indicated that GmWAK1 interacts with GmANNRJ4, and GmWAK1 plays a positive role in soybean resistance to P. sojae via a process that might be dependent on SA and involved in alleviating damage caused by oxidative stress.

Evaluation of Short-Season Soybean Genotypes for Resistance and Partial Resistance to Phytophthora sojae.

Phytophthora root and stem rot caused by Phytophthora sojae Kaufmann and Gerdemann is a soil-borne disease severely affecting soybean production worldwide. Losses caused by P. sojae can be controlled by both major genes and quantitative trait locus. Here, we tested 112 short-season soybean cultivars from Northeast China for resistance to P. sojae. A total of 58 germplasms were resistant to 7-11 P. sojae strains. Among these, Mengdou 28 and Kejiao 10-262 may harbor either Rps3a or multiple Rps genes conferring resistance to P. sojae. The remaining 110 germplasms produced 91 reaction types and may contain new resistance genes or gene combinations. Partial resistance evaluation using the inoculum layer method revealed that 34 soybean germplasms had high partial resistance, with a mean disease index lower than 30. Combining the results of resistance and partial resistance analyses, we identified 35 excellent germplasm resources as potential elite materials for resistance and tolerance in future breeding programs. In addition, we compared the radicle inoculation method with the inoculum layer method to screen for partial resistance to P. sojae. Our results demonstrate that the radicle inoculation method could potentially replace the inoculum layer method to identify partial resistance against P. sojae, and further verification with larger samples is required in the future.

The AP2/ERF GmERF113 Positively Regulates the Drought Response by Activating GmPR10-1 in Soybean.

Ethylene response factors (ERFs) are involved in biotic and abiotic stress; however, the drought resistance mechanisms of many ERFs in soybeans have not been resolved. Previously, we proved that GmERF113 enhances resistance to the pathogen Phytophthora sojae in soybean. Here, we determined that GmERF113 is induced by 20% PEG-6000. Compared to the wild-type plants, soybean plants overexpressing GmERF113 (GmERF113-OE) displayed increased drought tolerance which was characterized by milder leaf wilting, less water loss from detached leaves, smaller stomatal aperture, lower Malondialdehyde (MDA) content, increased proline accumulation, and higher Superoxide dismutase (SOD) and Peroxidase (POD) activities under drought stress, whereas plants with GmERF113 silenced through RNA interference were the opposite. Chromatin immunoprecipitation and dual effector-reporter assays showed that GmERF113 binds to the GCC-box in the GmPR10-1 promoter, activating GmPR10-1 expression directly. Overexpressing GmPR10-1 improved drought resistance in the composite soybean plants with transgenic hairy roots. RNA-seq analysis revealed that GmERF113 downregulates abscisic acid 8'-hydroxylase 3 (GmABA8'-OH 3) and upregulates various drought-related genes. Overexpressing GmERF113 and GmPR10-1 increased the abscisic acid (ABA) content and reduced the expression of GmABA8'-OH3 in transgenic soybean plants and hairy roots, respectively. These results reveal that the GmERF113-GmPR10-1 pathway improves drought resistance and affects the ABA content in soybean, providing a theoretical basis for the molecular breeding of drought-tolerant soybean.

The BTB/POZ domain protein GmBTB/POZ promotes the ubiquitination and degradation of the soybean AP2/ERF-like transcription factor GmAP2 to regulate the defense response to Phytophthora sojae.

Phytophthora root and stem rot in soybean (Glycine max) is a destructive disease worldwide, and hence improving crop resistance to the causal pathogen, P. sojae, is a major target for breeders. However, it remains largely unclear how the pathogen regulates the various affected signaling pathways in the host, which consist of complex networks including key transcription factors and their targets. We have previously demonstrated that GmBTB/POZ enhances soybean resistance to P. sojae and the associated defense response. Here, we demonstrate that GmBTB/POZ interacts with the transcription factor GmAP2 and promotes its ubiquitination. GmAP2-RNAi transgenic soybean hairy roots exhibited enhanced resistance to P. sojae, whereas roots overexpressing GmAP2 showed hypersensitivity. GmWRKY33 was identified as a target of GmAP2, which represses its expression by directly binding to the promoter. GmWRKY33 acts as a positive regulator in the response of soybean to P. sojae. Overexpression of GmBTB/POZ released the GmAP2-regulated suppression of GmWRKY33 in hairy roots overexpressing GmAP2 and increased their resistance to P. sojae. Taken together, our results indicate that GmBTB/POZ-GmAP2 modulation of the P. sojae resistance response forms a novel regulatory mechanism, which putatively regulates the downstream target gene GmWRKY33 in soybean.

Effect of Drought Stress during Soybean R2-R6 Growth Stages on Sucrose Metabolism in Leaf and Seed.

Sucrose is the main photosynthesis product of plants and the fundamental carbon skeleton monomer and energy supply for seed formation and development. Drought stress induces decreased photosynthetic carbon assimilation capacity, and seriously affects seed weight in soybean. However, little is known about the relationship between decreases in soybean seed yield and disruption of sucrose metabolism and transport balance in leaves and seeds during the reproductive stages of crop growth. Three soybean cultivars with similar growth periods, "Shennong17", "Shennong8", and "Shennong12", were subjected to drought stress during reproductive growth for 45 days. Drought stress significantly reduced leaf photosynthetic rate, shoot biomass, and seed weight by 63.93, 33.53, and 41.65%, respectively. Drought stress increased soluble sugar contents, the activities of sucrose phosphate synthase, sucrose synthase, and acid invertase enzymes, and up-regulated the expression levels of GmSPS1, GmSuSy2, and GmA-INV, but decreased starch content by 15.13% in leaves. Drought stress decreased the contents of starch, fructose, and glucose in seeds during the late seed filling stages, while it induced sucrose accumulation, which resulted in a decreased hexose-to-sucrose ratio. In developing seeds, the activities of sucrose synthesis and degradation enzymes, the expression levels of genes related to metabolism, and the expression levels of sucrose transporter genes were enhanced during early seed development under drought stress; however, under prolonged drought stress, all of them decreased. These results demonstrated that drought stress enhances the capacity for unloading sucrose into seeds and activated sucrose metabolism during early seed development. At the middle and late seed filling stages, sucrose flow from leaves to seeds was diminished, and the balance of sucrose metabolism was impaired in seeds, resulting in seed mass reduction. The different regulation strategies in sucrose allocation, metabolism, and transport during different seed development stages may be one of the physiological mechanisms for soybean plants to resist drought stress.

The bHLH transcription factor GmPIB1 facilitates resistance to Phytophthora sojae in Glycine max.

Phytophthora sojae Kaufmann and Gerdemann causes Phytophthora root rot, a destructive soybean disease worldwide. A basic helix-loop-helix (bHLH) transcription factor is thought to be involved in the response to P. sojae infection in soybean, as revealed by RNA sequencing (RNA-seq). However, the molecular mechanism underlying this response is currently unclear. Here, we explored the function and underlying mechanisms of a bHLH transcription factor in soybean, designated GmPIB1 (P. sojae-inducible bHLH transcription factor), during host responses to P. sojae. GmPIB1 was significantly induced by P. sojae in the resistant soybean cultivar 'L77-1863'. Analysis of transgenic soybean hairy roots with elevated or reduced expression of GmPIB1 demonstrated that GmPIB1 enhances resistance to P. sojae and reduces reactive oxygen species (ROS) accumulation. Quantitative reverse transcription PCR and chromatin immunoprecipitation-quantitative PCR assays revealed that GmPIB1 binds directly to the promoter of GmSPOD1 and represses its expression; this gene encodes a key enzyme in ROS production. Moreover, transgenic soybean hairy roots with GmSPOD1 silencing through RNA interference exhibited improved resistance to P. sojae and reduced ROS generation. These findings suggest that GmPIB1 enhances resistance to P. sojae by repressing the expression of GmSPOD1.

Overexpression of GmERF5, a new member of the soybean EAR motif-containing ERF transcription factor, enhances resistance to Phytophthora sojae in soybean.

Phytophthora root and stem rot of soybean [Glycine max (L.) Merr.], caused by Phytophthora sojae Kaufmann and Gerdemann, is a destructive disease throughout the soybean planting regions in the world. Here, we report insights into the function and underlying mechanisms of a novel ethylene response factor (ERF) in soybean, namely GmERF5, in host responses to P. sojae. GmERF5-overexpressing transgenic soybean exhibited significantly enhanced resistance to P. sojae and positively regulated the expression of the PR10, PR1-1, and PR10-1 genes. Sequence analysis suggested that GmERF5 contains an AP2/ERF domain of 58 aa and a conserved ERF-associated amphiphilic repression (EAR) motif in its C-terminal region. Following stress treatments, GmERF5 was significantly induced by P. sojae, ethylene (ET), abscisic acid (ABA), and salicylic acid (SA). The activity of the GmERF5 promoter (GmERF5P) was upregulated in tobacco leaves with ET, ABA, Phytophthora nicotianae, salt, and drought treatments, suggesting that GmERF5 could be involved not only in the induced defence response but also in the ABA-mediated pathway of salt and drought tolerance. GmERF5 could bind to the GCC-box element and act as a repressor of gene transcription. It was targeted to the nucleus when transiently expressed in Arabidopsis protoplasts. GmERF5 interacted with a basic helix-loop-helix transcription factor (GmbHLH) and eukaryotic translation initiation factor (GmEIF) both in yeast cells and in planta. To the best of our knowledge, GmERF5 is the first soybean EAR motif-containing ERF transcription factor demonstrated to be involved in the response to pathogen infection.

Isolation and characterization of a pathogenesis-related protein 10 gene (GmPR10) with induced expression in soybean (Glycine max) during infection with Phytophthora sojae.

In previous study, a cDNA library enriched for mRNAs encoding ESTs that increased in abundance during infection with Phytophthora sojae was constructed by suppression subtractive hybridization from leaf tissues of a high resistant soybean, and an EST homologous to the class 10 of pathogenesis-related (PR) proteins was identified to be up-regulated by microarray and real-time PCR. Here, the full-length cDNA (termed GmPR10, GenBank accession number FJ960440; ADC31789.1) of the EST was isolated by rapid amplification of cDNA ends, and contains an open reading frame of 474 bp. The GmPR10 protein included a "P-loop'' motif. The constitutive transcript abundance of GmPR10 in soybean was the highest in leaves, followed by roots and stems. Further analysis showed that GmPR10 mRNA abundance was increased during infection with P. sojae following leaf treatments with gibberellin (GA3), hydrogen peroxide (H2O2), salicylic acid (SA), and abscisic acid (ABA). The dialytically renatured GmPR10 protein significantly inhibited P. sojae hyphal growth and exhibited RNase activity. Transgenic tobacco and soybean plants overexpressing GmPR10 showed increased resistance to P. nicotianae Breda and P. sojae, respectively. These results suggest that the GmPR10 protein plays an important role in host defense against P. sojae infection. To the best of our knowledge, this is the first report on the functional characterization of a PR10 protein from soybean in defense against P. sojae.

The EIN3 transcription factor GmEIL1 improves soybean resistance to Phytophthora sojae.

Phytophthora root and stem rot of soybean (Glycine max), caused by the oomycete Phytophthora sojae, is an extremely destructive disease worldwide. In this study, we identified GmEIL1, which encodes an ethylene-insensitive3 (EIN3) transcription factor. GmEIL1 was significantly induced following P. sojae infection of soybean plants. Compared to wild-type soybean plants, transgenic soybean plants overexpressing GmEIL1 showed enhanced resistance to P. sojae and GmEIL1-silenced RNA-interference lines showed more severe symptoms when infected with P. sojae. We screened for target genes of GmEIL1 and confirmed that GmEIL1 bound directly to the GmERF113 promoter and regulated GmERF113 expression. Moreover, GmEIL1 positively regulated the expression of the pathogenesis-related gene GmPR1. The GmEIL1-regulated defence response to P. sojae involved both ethylene biosynthesis and the ethylene signalling pathway. These findings suggest that the GmEIL1-GmERF113 module plays an important role in P. sojae resistance via the ethylene signalling pathway.

Stimulation of primed carbon under climate change corresponds with phosphorus mineralization in the rhizosphere of soybean.

Elevated CO2 and temperature likely alter photosynthetic carbon inputs to soils, which may stimulate soil microbial activity to accelerate the decomposition of soil organic carbon (SOC), liberating more phosphorus (P) into the soil solution. However, this hypothesis on the association of SOC decomposition and P transformation in the plant rhizosphere requires robust soil biochemical evidence, which is critical to nutrient management for the mitigation of soil quality against climate change. This study investigated the microbial functional genes relevant to P mineralization together with priming processes of SOC in the rhizosphere of soybean grown under climate change. Soybean plants were grown under elevated CO2 (eCO2, 700 ppm) combined with warming (+ 2 °C above ambient temperature) in open-top chambers. Photosynthetic carbon flow in the plant-soil continuum was traced with 13CO2 labeling. The eCO2 plus warming treatment increased the primed carbon (C) by 43 % but decreased the NaHCO3-extratable organic P by 33 %. Furthermore, NaHCO3-Po was negatively correlated with phosphatase activity and microbial biomass C. Elevated CO2 increased the abundances of C degradation genes, such as abfA and ManB, and P mineralization genes, such as gcd, phoC and phnK. The results suggested that increased photosynthetic carbon inputs to the rhizosphere of plants under eCO2 plus warming stimulated the microbial population and metabolic functions of both SOC and organic P mineralization. There is a positive relationship between the rhizosphere priming effect and P mineralization. The response of microorganisms to plant-C flow is decisive for coupled C and P cycles, which are likely accelerated under climate change.

Fluorination Enables Tunable Molecular Interaction and Photovoltaic Performance in Non-Fullerene Solar Cells Based on Ester-Substituted Polythiophene.

Owing to the advantages of low synthetic cost and high scalability of synthesis, polythiophene and its derivatives (PTs) have been of interest in the community of organic photovoltaics (OPVs). Nevertheless, the typical efficiency of PT based photovoltaic devices reported so far is much lower than those of the prevailing push-pull type conjugated polymer donors. Recent studies have underscored that the excessively low miscibility between PT and nonfullerene acceptor is the major reason accounting for the unfavorable active layer morphology and the inferior performance of OPVs based on a well-known PT, namely PDCBT-Cl and a non-halogenated nonfullerene acceptor IDIC. How to manipulate the miscibility between PT and acceptor molecule is important for further improving the device efficiency of this class of potentially low-cost blend systems. In this study, we introduced different numbers of F atoms to the end groups of IDIC to tune the intermolecular interaction of the hypo-miscible blend system (PDCBT-Cl:IDIC). Based on calorimetric, microscopic, and scattering characterizations, a clear relationship between the number of F atoms, miscibility, and device performance was established. With the increased number of F atoms in IDIC, the resulting acceptors exhibited enhanced miscibility with PDCBT-Cl, and the domain sizes of the blend films were reduced substantially. As a result, distinctively different photovoltaic performances were achieved for these blend systems. This study demonstrates that varying the number of F atoms in the acceptors is a feasible way to manipulate the molecular interaction and the film morphology toward high-performance polythiophene:nonfullerene based OPVs.

GmBTB/POZ promotes the ubiquitination and degradation of LHP1 to regulate the response of soybean to Phytophthora sojae.

Phytophthora sojae is a pathogen that causes stem and root rot in soybean (Glycine max [L.] Merr.). We previously demonstrated that GmBTB/POZ, a BTB/POZ domain-containing nuclear protein, enhances resistance to P. sojae in soybean, via a process that depends on salicylic acid (SA). Here, we demonstrate that GmBTB/POZ associates directly with soybean LIKE HETEROCHROMATIN PROTEIN1 (GmLHP1) in vitro and in vivo and promotes its ubiquitination and degradation. Both overexpression and RNA interference analysis of transgenic lines demonstrate that GmLHP1 negatively regulates the response of soybean to P. sojae by reducing SA levels and repressing GmPR1 expression. The WRKY transcription factor gene, GmWRKY40, a SA-induced gene in the SA signaling pathway, is targeted by GmLHP1, which represses its expression via at least two mechanisms (directly binding to its promoter and impairing SA accumulation). Furthermore, the nuclear localization of GmLHP1 is required for the GmLHP1-mediated negative regulation of immunity, SA levels and the suppression of GmWRKY40 expression. Finally, GmBTB/POZ releases GmLHP1-regulated GmWRKY40 suppression and increases resistance to P. sojae in GmLHP1-OE hairy roots. These findings uncover a regulatory mechanism by which GmBTB/POZ-GmLHP1 modulates resistance to P. sojae in soybean, likely by regulating the expression of downstream target gene GmWRKY40.

The 26S Proteasome Regulatory Subunit GmPSMD Promotes Resistance to Phytophthora sojae in Soybean.

Phytophthora root rot, caused by Phytophthora sojae is a destructive disease of soybean (Glycine max) worldwide. We previously confirmed that the bHLH transcription factor GmPIB1 (P. sojae-inducible bHLH transcription factor) reduces accumulation of reactive oxygen species (ROS) in cells by inhibiting expression of the peroxidase-related gene GmSPOD thus improving the resistance of hairy roots to P. sojae. To identify proteins interacting with GmPIB1 and assess their participation in the defense response to P. sojae, we obtained transgenic soybean hairy roots overexpressing GmPIB1 by Agrobacterium rhizogenes mediated transformation and examined GmPIB1 protein-protein interactions using immunoprecipitation combined with mass spectrometry. We identified 392 proteins likely interacting with GmPIB1 and selected 20 candidate genes, and only 26S proteasome regulatory subunit GmPSMD (Genbank accession no. XP_014631720) interacted with GmPIB1 in luciferase complementation and pull-down experiments and yeast two-hybrid assays. Overexpression of GmPSMD (GmPSMD-OE) in soybean hairy roots remarkably improved resistance to P. sojae and RNA interference of GmPSMD (GmPSMD -RNAi) increased susceptibility. In addition, accumulation of total ROS and hydrogen peroxide (H2O2) in GmPSMD-OE transgenic soybean hairy roots were remarkably lower than those of the control after P. sojae infection. Moreover, in GmPSMD-RNAi transgenic soybean hairy roots, H2O2 and the accumulation of total ROS exceeded those of the control. There was no obvious difference in superoxide anion (O2 -) content between control and transgenic hairy roots. Antioxidant enzymes include peroxidase (POD), glutathione peroxidase (GPX), superoxide dismutase (SOD), catalase (CAT) are responsible for ROS scavenging in soybean. The activities of these antioxidant enzymes were remarkably higher in GmPSMD-OE transgenic soybean hairy roots than those in control, but were reduced in GmPSMD-RNAi transgenic soybean hairy roots. Moreover, the activity of 26S proteasome in GmPSMD-OE and GmPIB1-OE transgenic soybean hairy roots was significantly higher than that in control and was significantly lower in PSMD-RNAi soybean hairy roots after P. sojae infection. These data suggest that GmPSMD might reduce the production of ROS by improving the activity of antioxidant enzymes such as POD, SOD, GPX, CAT, and GmPSMD plays a significant role in the response of soybean to P. sojae. Our study reveals a valuable mechanism for regulation of the pathogen response by the 26S proteasome in soybean.

Races of Phytophthora sojae and Their Virulences on Soybean Cultivars in Heilongjiang, China.

Phytophthora root and stem rot, caused by Phytophthora sojae, is an economically important disease of soybean (Glycine max) in Heilongjiang Province, China. The objectives of this research were to determine the race profile of P. sojae in Heilongjiang and evaluate soybean cultivars for reactions to the pathogen races. A total of 96 single-zoospore P. sojae isolates were obtained from soil samples collected from 35 soybean fields in 18 counties in Heilongjiang from 2005 to 2007. Eight races of P. sojae, including races 1, 3, 4, 5, 9, 13, 44, and 54, were identified on a set of eight differentials, each containing a single resistance Rps gene, from 80 of the 96 isolates. Races 1 and 3 were predominant races, comprising 58 and 14 isolates, and representing 60 and 7% of the pathogen population, respectively. Races 4, 5, 44, and 54 were identified for the first time in Heilongjiang, and each was represented by two to three isolates only. Sixty-two soybean cultivars commonly grown in Heilongjiang Province were evaluated for their resistance to the eight P. sojae races identified using the hypocotyl inoculation technique. Based on the percentage of plant mortality rated 5 days after inoculation, 44 cultivars were resistant (<30% mortality) to at least one race. These cultivars may be used as sources of resistance in soybean breeding programs.

Overexpression of a soybean 4-coumaric acid: coenzyme A ligase (GmPI4L) enhances resistance to Phytophthora sojae in soybean.

Phytophthora root and stem rot of soybean (Glycine max (L.) Merr.) caused by Phytophthora sojae is a destructive disease worldwide. The enzyme 4-coumarate: CoA ligase (4CL) has been extensively studied with regard to plant responses to pathogens. However, the molecular mechanism of the response of soybean 4CL to P. sojae remains unclear. In a previous study, a highly upregulated 4CL homologue was characterised through suppressive subtractive hybridisation library and cDNA microarrays, in the resistant soybean cultivar 'Suinong 10' after infection with P. sojae race 1. Here, we isolated the full-length EST, and designated as GmPI4L (P. sojae-inducible 4CL gene) in this study, which is a novel member of the soybean 4CL gene family. GmPI4L has 34-43% over all amino acid sequence identity with other plant 4CLs. Overexpression of GmPI4L enhances resistance to P. sojae in transgenic soybean plants. The GmPI4L is located in the cell membrane when transiently expressed in Arabidopsis protoplasts. Further analyses showed that the contents of daidzein, genistein, and the relative content of glyceollins are significantly increased in overexpression GmPI4L soybeans. Taken together, these results suggested that GmPI4L plays an important role in response to P. sojae infection, possibly by enhancing the content of glyceollins, daidzein, and genistein in soybean.

GmBTB/POZ, a novel BTB/POZ domain-containing nuclear protein, positively regulates the response of soybean to Phytophthora sojae infection.

Phytophthora sojae is a destructive pathogen of soybean [Glycine max (L.) Merr.] which causes stem and root rot on soybean plants worldwide. However, the pathogenesis and molecular mechanism of plant defence responses against P. sojae are largely unclear. Herein, we document the underlying mechanisms and function of a novel BTB/POZ protein, GmBTB/POZ, which contains a BTB/POZ domain found in certain animal transcriptional regulators, in host soybean plants in response to P. sojae. It is located in the cell nucleus and is transcriptionally up-regulated by P. sojae. Overexpression of GmBTB/POZ in soybean resulted in enhanced resistance to P. sojae. The activities and expression levels of enzymatic superoxide dismutase (SOD) and peroxidase (POD) antioxidants were significantly higher in GmBTB/POZ-overexpressing (GmBTB/POZ-OE) transgenic soybean plants than in wild-type (WT) plants treated with sterile water or infected with P. sojae. The transcript levels of defence-associated genes were also higher in overexpressing plants than in WT on infection. Moreover, salicylic acid (SA) levels and the transcript levels of SA biosynthesis-related genes were markedly higher in GmBTB/POZ-OE transgenic soybean than in WT, but there were almost no differences in jasmonic acid (JA) levels or JA biosynthesis-related gene expression between GmBTB/POZ-OE and WT soybean lines. Furthermore, exogenous SA application induced the expression of GmBTB/POZ and inhibited the increase in P. sojae biomass in both WT and GmBTB/POZ-OE transgenic soybean plants. Taken together, these results suggest that GmBTB/POZ plays a positive role in P. sojae resistance and the defence response in soybean via a process that might be dependent on SA.

GmSnRK1.1, a Sucrose Non-fermenting-1(SNF1)-Related Protein Kinase, Promotes Soybean Resistance to Phytophthora sojae.

Phytophthora root and stem rot, a destructive disease of soybean [Glycine max (L.) Merr.], is caused by the oomycete Phytophthora sojae. However, how the disease resistance mechanisms of soybean respond to P. sojae infection remains unclear. Previously, we showed that GmWRKY31, which interacts with a sucrose non-fermenting-1(SNF1)-related protein kinase (SnRK), enhances resistance to P. sojae in soybean. Here, we report that the membrane-localized SnRK GmSnRK1.1 is involved in the soybean host response to P. sojae. The overexpression of GmSnRK1.1 (GmSnRK1.1-OE) increased soybean resistance to P. sojae, and the RNA interference (RNAi)-mediated silencing of GmSnRK1.1 (GmSnRK1.1-R) reduced resistance to P. sojae. Moreover, the activities and transcript levels of the antioxidant enzymes superoxide dismutase and peroxidase were markedly higher in the GmSnRK1.1-OE transgenic soybean plants than in the wild type (WT), but were reduced in the GmSnRK1.1-R plants. Several isoflavonoid phytoalexins related genes GmPAL, GmIFR, Gm4CL and GmCHS were significantly higher in "Suinong 10" and GmSnRK1.1-OE lines than these in "Dongnong 50," and were significantly lower in GmSnRK1.1-R lines. In addition, the accumulation of salicylic acid (SA) and the expression level of the SA biosynthesis-related gene were significantly higher in the GmSnRK1.1-OE plants than in the WT and GmSnRK1.1-R plants, moreover, SA biosynthesis inhibitor treated GmSnRK1.1-R lines plants displayed clearly increased pathogen biomass compared with H2O-treated plants after 24 h post-inoculation. These results showed that GmSnRK1.1 positively regulates soybean resistance to P. sojae, potentially functioning via effects on the expression of SA-related genes and increased accumulation of SA.

Impact of Elevated CO2 on Seed Quality of Soybean at the Fresh Edible and Mature Stages.

Although the effect of elevated CO2 (eCO2) on soybean yield has been well documented, few studies have addressed seed quality, particularly at the fresh edible (R6) and mature stages (R8). Under the current global scenario of increasing CO2 levels, this potentially threatens the nutritional content and quality of food crops. Using four soybean cultivars, we assessed the effects of eCO2 on the concentrations of crude protein, crude oil, and isoflavones and analyzed the changes in free amino acids, fatty acids, and mineral elements in seeds. At R6, eCO2 had no influence on soybean seed protein and oil concentrations. At R8, eCO2 significantly decreased seed protein concentration but increased seed oil concentration; it also significantly decreased total free amino acid concentration. However, at the same stage, the proportion of oleic acid (18:1) among fatty acids increased in response to eCO2 in the cultivars of Zhongke-maodou 2 (ZK-2) and Zhongke-maodou 3 (ZK-3), and a similar trend was found for linoleic acid (18:2) in Zhongke-maodou 1 (ZK-1) and Hei-maodou (HD). Total isoflavone concentrations increased significantly at both the R6 and R8 stages in response to eCO2. Compared with ambient CO2, the concentrations of K, Ca, Mg, P, and S increased significantly under eCO2 at R6, while the Fe concentration decreased significantly. The response of Zn and Mn concentrations to eCO2 varied among cultivars. At R8 and under eCO2, Mg, S, and Ca concentrations increased significantly, while Zn and Fe concentrations decreased significantly. These findings suggest that eCO2 is likely to benefit from the accumulation of seed fat and isoflavone but not from that of protein. In this study, the response of seed mineral nutrients to eCO2 varied between cultivars.