Rhg1 has been the most effective QTL deployed in soybeans (Glycine max) to control soybean cyst nematode (Heterodera glycines; SCN). However, the resistance mechanisms and specificity of Rhg1 towards SCN as opposed to other plant parasitic nematodes are not well known. In this study we report that Rhg1 can hinder the parasitism of the root lesion nematode Pratylenchus penetrans (Pp), which is commonly found along with SCN in northern U.S. soybean fields. Two elite public soybean varieties, carrying either rhg1-a + Rhg4 or rhg1-b, negatively impacted Pp populations measured at 30 dpi. This contribution of Rhg1 was more rigorously demonstrated using near isogenic lines for rhg1-b. Additionally, in three of the four soybean genetic backgrounds tested we observed significantly more Pp at 30 dpi when Pp were co-inoculated with SCN, consistent with a previous study. No significant changes in SCN cyst numbers upon co-inoculation with Pp were observed for any of the SCN-resistant rhg1 lines. No nematode-induced changes in Rhg1 transcript abundances were observed when sampling infected root sections in any of the plant genotypes, and no changes in a jasmonate response indicator were observed. However, the 3 dpi salicylate-mediated response to SCN or Pp became more robust during SCN + Pp co-infection. Multiple hypotheses for further dissection and manipulation of soybean resistance to Pratylenchus root lesion nematodes are suggested based on the present findings.

Soybean cyst nematode (SCN, Heterodera glycines) causes major yield losses, and Rhg1 locus-mediated genetic resistance is becoming less effective. Hormonal signaling pathways, particularly salicylic acid (SA) and gibberellic acid (GA), are increasingly implicated in nematode resistance, but their use in plant protection remains underexplored. Here, we tested exogenous application of chemical modulators of these pathways for effects on SCN resistance and dependence on the presence or absence of Rhg1-b. We found that foliar application of the SA mimic acibenzolar-S-methyl (ASM) or the systemic acquired resistance mediator pipecolic acid (Pip) conferred strong, genotype-independent resistance, comparable to genetic resistance at Rhg1. Combining ASM with the GA biosynthesis inhibitor paclobutrazol (PBZ) synergistically enhanced protection, reducing cyst formation by 39.2% and effectively making plants moderately resistant to SCN. These effects were observed in the susceptible cultivar Williams 82, as well as across near-isogenic lines with varying Rhg1 copy numbers. ASM alone boosted resistance in resistant Rhg1-b and susceptible Rhg1-c backgrounds, whereas PBZ provided additional benefits only in Rhg1-c, revealing genotype-specific interactions between hormonal signaling and host resistance. Targeted modulation of SA and GA pathways thus provides an effective and sustainable strategy to suppress SCN, complementing and extending the durability of genetic resistance. [Formula: see text] Copyright © 2026 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Soybean cyst nematode (SCN, Heterodera glycines) causes major yield losses, and Rhg1 locus-mediated genetic resistance is becoming less effective. Hormonal signaling pathways, particularly salicylic acid (SA) and gibberellic acid (GA), are increasingly implicated in nematode resistance, but their use in plant protection remains underexplored. Here, we tested exogenous application of chemical modulators of these pathways for effects on SCN resistance and dependence on the presence or absence of Rhg1-b. We found that foliar application of the SA mimic acibenzolar-S-methyl (ASM) or the systemic acquired resistance mediator pipecolic acid (Pip) conferred strong, genotype-independent resistance, comparable to genetic resistance at Rhg1. Combining ASM with the GA biosynthesis inhibitor paclobutrazol (PBZ) synergistically enhanced protection, reducing cyst formation by 39.2% and effectively making plants moderately resistant to SCN. These effects were observed in the susceptible cultivar Williams 82, as well as across near-isogenic lines with varying Rhg1 copy numbers. ASM alone boosted resistance in resistant Rhg1-b and susceptible Rhg1-c backgrounds, while PBZ provided additional benefits only in Rhg1-c, revealing genotype-specific interactions between hormonal signaling and host resistance. Targeted modulation of SA and GA pathways thus provides an effective and sustainable strategy to suppress SCN, complementing and extending the durability of genetic resistance.

The rhg1-a and rhg1-b haplotypes of the soybean Rhg1 locus are economically effective tools for the control of soybean cyst nematode (SCN; Heterodera glycines), but ongoing SCN evolution requires improved sources of resistance. Both Rhg1 haplotypes carry multiple tandem repeat copies of a four-gene block encoding four disparate proteins, and resistance efficacy scales with copy number. The haplotypes encode different variants of an unusual α-SNAP protein whose abundance increases in the nematode-reprogrammed plant cells that form the syncytium (nematode feeding site), which subsequently collapses. The present study explored the simultaneous presence of two α-SNAPRhg1 protein types and elevation of α-SNAPRhg1 abundance, which were hypothesized to improve SCN resistance but are not achievable by conventional soybean breeding. We accomplished both via transgenic additions to an rhg1-a Rhg4 soybean line. Existing resistance to the HG type 0 and HG type 2.5.7 SCN was strengthened, measured as cyst production and final SCN egg counts, but resistance was not improved against an HG type 1.3.6.7 SCN population that was already highly virulent on the parent line. Greenhouse and field studies with a subset of lines showed no significant yield penalties. We conclude that cisgenic combination or elevated expression of the different α-SNAP genes may extend the efficacy and/or durability of Rhg1-mediated resistance. We also observed differential protein abundances for some α-SNAPRhg1 isoforms after inoculation with different HG type populations of SCN. The studies provide a foundation for further refinement of Rhg1-based SCN control. [Formula: see text] Copyright © 2025 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The soybean complex resistance locus Rhg1 is a primary contributor to the control of soybean cyst nematode (Heterodera glycines; SCN), a major threat to soybean production worldwide. Two genes within this locus, GmAATRhg1 and GmSNAP18, are upregulated during SCN infection. In this study, we found that SCNresistant soybean varieties exhibit elevated ethylene (ETH) production upon infection. Exogenous ETH application further increased GmAATRhg1 and GmSNAP18 transcript levels and enhanced the SCN-resistance response. These genes are divergently transcribed from a shared promoter containing three ERELEE4 ethylene-responsive elements (AATTCAAA). Using yeast one-hybrid screening, we identified the AP2/ERF transcription factor GmTINY (Glyma.01G147600) as an ERELEE4-binding protein. GmTINY is localized to the nucleus, and its expression is induced by both ETH treatment and SCN infection. Molecular, biochemical, and genetic analyses showed that GmTINY binds to the Rhg1 ERELEE4 elements and regulates the expression of GmAATRhg1 and GmSNAP18, thereby enhancing SCN resistance. Additionally, over-expression of GmTINY induced several xyloglucan endotransglycosylase/hydrolase (XTH) genes, particularly GmXTH2, which is implicated in cell wall remodeling and restricts nematode development beyond the J2 stage. Together, these findings reveal an ETH-GmTINY signaling axis that regulates both Rhg1-medi-ated and cell wall-related resistance pathways, providing new insights for engineering durable SCN resistance in soybean.

Rhg1 is the most important locus conferring resistance to soybean cyst nematode (SCN; Heterodera glycine Ichinohe) in soybean [ Glycine max (L.) Merr.]. Previous research has shown that to obtain viable plants, the SCN resistance allele at Rhg1 on chromosome 18 needs to be paired with NSF RAN07 , an atypical r esistance‐ a ssociated N SF allele of the N‐ethylmaleimide sensitive factor ( NSF ) gene on chromosome 07. This causes segregation distortion in populations developed from crosses between resistant and susceptible plants. Our study aimed to improve our understanding of this segregation distortion and determine the developmental stage at which it occurs. DNA from developing F 2 seeds and F 2 plants originating from crosses between resistant and susceptible parents was genotyped with markers for the rhg1 and NSF loci using TaqMan assays. Chi‐square tests revealed significant deviations from the expected Mendelian segregation ratio (1:2:1:2:4:2:1:2:1) in both F 2 seeds and plants, indicating segregation distortion at these loci. The absence of the rhg1‐b_rhg1‐b_NSF Ch07 _NSF Ch07 genotype supports the previous finding that the combination of the resistance allele rhg1‐b and the commonly occurring NSF Ch07 allele is lethal, apparently because the α‐SNAP (where SNAP is soluble NSF attachment protein) encoded by rhg1‐b or rhg1‐a interacts well with the NSF RAN07 protein but not the more common NSF Ch07 protein. The findings indicate that segregation distortion occurs prior to seed maturation and is primarily due to zygotic selection during early seed development. The results emphasize the need to consider this genetic interaction in breeding efforts to improve soybean since segregation distortion may affect the inheritance of SCN resistance and other traits linked to Rhg1 or NSF Ch07 .

Amino acid transporters play crucial roles in plant nitrogen metabolism but also in defense responses. AATRhg1, an apparent amino acid transporter encoded by Glyma.18g022400 (Rhg1-GmAAT) at the soybean Rhg1 locus, contributes to resistance to soybean cyst nematode (SCN), although the in planta function of AATRhg1 remains elusive. In this study, we discovered that overexpression of Rhg1-GmAAT in soybean roots enhances the betalain pigment synthesis driven by a RUBY transgene cassette, potentially through its transporter activity affecting tyrosine levels and amino acid homeostasis. Silencing Rhg1-GmAAT also moderately increased betalain accumulation, while co-overexpression of Rhg1-GmAAT and GmRBOHG (encoding an AATRhg1-interacting NADPH oxidase) blocked the betalain phenotype, indicating a complex role of AATRhg1 in regulating cellular metabolism. Soybean AATRhg1 did not show a betalain accumulation phenotype when co-overexpressed with RUBY in Nicotiana benthamiana leaves, suggesting that soybean AATRhg1 functions differently in N. benthamiana. In soybean, expression of AATRhg1 proteins with mutations at conserved residues D122A or Y268L mitigated or enhanced the betalain phenotypes, respectively, suggesting that these residues are important for AATRhg1 function. This study advances our understanding of AATRhg1 while presenting a novel strategy for enhancing betalain biosynthesis by modulating the transport and homeostasis of amino acids.

Research in Arabidopsis thaliana has a powerful influence on our understanding of gene functions and pathways. However, not everything translates from Arabidopsis to crops and other plants. Here, a group of experts consider instances where translation has been lost and why such translation is not possible or is challenging. First, despite great efforts, floral dip transformation has not succeeded in other species outside Brassicaceae. Second, due to gene duplications and losses throughout evolution, it can be complex to establish which genes are orthologs of Arabidopsis genes. Third, during evolution Arabidopsis has lost arbuscular mycorrhizal symbiosis. Fourth, other plants have evolved specialized cell types that are not present in Arabidopsis. Fifth, similarly, C4 photosynthesis cannot be studied in Arabidopsis, which is a C3 plant. Sixth, many other plant species have larger genomes, which has given rise to innovations in transcriptional regulation that are not present in Arabidopsis. Seventh, phenotypes such as acclimation to water stress can be challenging to translate due to different measurement strategies. And eighth, while the circadian oscillator is conserved, there are important nuances in the roles of circadian regulators in crop plants. A key theme emerging across these vignettes is that even when translation is lost, insights can still be gained through comparison with Arabidopsis.

Soybean cyst nematode (SCN) is the most yield-reducing pathogen of soybean. Resistance based on the complex rhg1-b haplotype has been the primary control measure, but gradual SCN evolution is incrementally eroding rhg1-b efficacy. rhg1-b carries ten copies of a ~31 kb chromosomal segment with three different genes that contribute to resistance. We are functionally dissecting the rhg1-b α-SNAP to understand and possibly improve this novel defense mechanism. α-SNAP is a housekeeping protein with C-terminal amino acids that are conserved across multicellular eukaryotes. a-SNAP interacts with NSF to recycle SNARE protein bundles that mediate vesicle fusion to target membranes. The rhg1-b a-SNAPRhg1HC protein differs from canonical α-SNAPs at its C-terminus and apparently poisons the nematode-plant biotrophy when its level increases 10-20 fold at SCN feeding sites (syncytia). We will describe our site-directed mutagenesis to alter and test functional C-terminal residues and potentially block SCN effector action against rhg1-b α-SNAP. We are also investigating the mechanisms by which a-SNAPRhg1HC abundance is elevated at syncytia, and will report an in situ assay system to obtain spatially resolved transcript abundance data at syncytia via confocal microscopy. This system is now being used to dissect rhg1-b promoter elements as one logical first step to understanding mechanisms of infection-site upregulation and potentially allow promoter modification to enhance SCN resistance.

Whole genome resequencing (WGRS) platforms provide exceptional fingerprinting of the entire genome but are expensive and less flexible to use as a routine genotyping tool for targeting causal polymorphisms within a germplasm collection or breeding program. Therefore, there has been a continuous effort to develop small-scale genotyping platforms that facilitate robust and quick assessments of the allelic status of causal variants for important traits within soybean breeding programs. The objective was to develop a comprehensive panel of soybean cyst nematode (SCN) resistance TaqMan® assays via selecting the causative genes and analyzing their associated alleles. The Soybean Allele Catalog was utilized to investigate WGRS-derived variants which are predicted to cause a change in the amino acid sequence of a gene product. This panel of TaqMan® assays reflects current knowledge about known SCN resistance-causing genes and their associated alleles: GmSNAP18-a and -b, GmSNAP11, GmSHMT08, GmSNAP15, GmNSFRAN07, and GmSNAP02-ins and -del. Developed assays were tested using elite breeding lines and segregating populations. TaqMan assays were compared to other currently available KASP and CAPS assays. All assays showed excellent allele determination efficiencies. This SCN genotyping assay panel can be utilized as a simplified, accurate and reliable genotyping platform further equipping the updated soybean breeding toolbox.

Rhg1 (Resistance to Heterodera glycines 1) mediates soybean (Glycine max) resistance to soybean cyst nematode (SCN; H. glycines). Rhg1 is a 4-gene, ∼30-kb block that exhibits copy number variation, and the common PI 88788-type rhg1-b haplotype carries 9 to 10 tandem Rhg1 repeats. Glyma.18G022400 (Rhg1-GmAAT), 1 of 3 resistance-conferring genes at the complex Rhg1 locus, encodes the putative amino acid transporter AATRhg1 whose mode of action is largely unknown. We discovered that AATRhg1 protein abundance increases 7- to 15-fold throughout root cells along the migration path of SCN. These root cells develop an increased abundance of vesicles and large vesicle-like bodies (VLB) as well as multivesicular and paramural bodies. AATRhg1 protein is often present in these structures. AATRhg1 abundance remained low in syncytia (plant cells reprogrammed by SCN for feeding), unlike the Rhg1 α-SNAP protein, whose abundance has previously been shown to increase in syncytia. In Nicotiana benthamiana, if soybean AATRhg1 was present, oxidative stress promoted the formation of large VLB, many of which contained AATRhg1. AATRhg1 interacted with the soybean NADPH oxidase GmRBOHG, the ortholog of Arabidopsis thaliana RBOHD previously found to exhibit upregulated expression upon SCN infection. AATRhg1 stimulated reactive oxygen species (ROS) generation when AATRhg1 and GmRBOHG were co-expressed. These findings suggest that AATRhg1 contributes to SCN resistance along the migration path as SCN invades the plant and does so, at least in part, by increasing ROS production. In light of previous findings about α-SNAPRhg1, this study also shows that different Rhg1 resistance proteins function via at least 2 spatially and temporally separate modes of action.

Plant-parasitic nematodes are a major threat to crop production in all agricultural systems. The scarcity of classical resistance genes highlights a pressing need to find new ways to develop nematode-resistant germplasm. Here, we sequence and assemble a high-quality phased genome of the model cyst nematode Heterodera schachtii to provide a platform for the first system-wide dual analysis of host and parasite gene expression over time, covering all major parasitism stages. Analysis of the hologenome of the plant-nematode infection site identified metabolic pathways that were incomplete in the parasite but complemented by the host. Using a combination of bioinformatic, genetic, and biochemical approaches, we show that a highly atypical completion of vitamin B5 biosynthesis by the parasitic animal, putatively enabled by a horizontal gene transfer from a bacterium, is required for full pathogenicity. Knockout of either plant-encoded or now nematode-encoded steps in the pathway significantly reduces parasitic success. Our experiments establish a reference for cyst nematodes, further our understanding of the evolution of plant-parasitism by nematodes, and show that congruent differential expression of metabolic pathways in the infection hologenome represents a new way to find nematode susceptibility genes. The approach identifies genome-editing-amenable targets for future development of nematode-resistant crops.

Resistance to the soybean cyst nematode (SCN) is a topic incorporating multiple mechanisms and multiple types of science. It is also a topic of substantial agricultural importance, as SCN is estimated to cause more yield damage than any other pathogen of soybean, one of the world's main food crops. Both soybean and SCN have experienced jumps in experimental tractability in the past decade, and significant advances have been made. The rhg1-b locus, deployed on millions of farm acres, has been durable and will remain important, but local SCN populations are gradually evolving to overcome rhg1-b. Multiple other SCN resistance quantitative trait loci (QTL) of proven value are now in play with soybean breeders. QTL causal gene discovery and mechanistic insights into SCN resistance are contributing to both basic and applied disciplines. Additional understanding of SCN and other cyst nematodes will also grow in importance and lead to novel disease control strategies.

Poly(ADP-ribosyl)ation (PARylation) is a posttranslational modification reversibly catalyzed by poly(ADP-ribose) polymerases (PARPs) and poly(ADP-ribose) glycohydrolases (PARGs) and plays a key role in multiple cellular processes. The molecular mechanisms by which PARylation regulates innate immunity remain largely unknown in eukaryotes. Here we show that Arabidopsis UBC13A and UBC13B, the major drivers of lysine 63 (K63)-linked polyubiquitination, directly interact with PARPs/PARGs. Activation of pathogen-associated molecular pattern (PAMP)-triggered immunity promotes these interactions and enhances PARylation of UBC13. Both parp1 parp2 and ubc13a ubc13b mutants are compromised in immune responses with increased accumulation of total pathogenesis-related (PR) proteins but decreased accumulation of secreted PR proteins. Protein disulfide-isomerases (PDIs), essential components of endoplasmic reticulum quality control (ERQC) that ensure proper folding and maturation of proteins destined for secretion, complex with PARPs/PARGs and are PARylated upon PAMP perception. Significantly, PARylation of UBC13 regulates K63-linked ubiquitination of PDIs, which may further promote their disulfide isomerase activities for correct protein folding and subsequent secretion. Taken together, these results indicate that plant immunity is coordinately regulated by PARylation and K63-linked ubiquitination.