Toll/interleukin-1 receptor (TIR) enzymes are prominent immune components in diverse organisms across the tree of life. In flowering plants, TIRs are often integrated into nucleotide binding leucine-rich repeat (NLR) receptors whose oligomerization-dependent biochemical activities create second messengers perceived by ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1)-family signaling complexes. TIR-NLRs and TIR proteins are present across the full spectrum of plant evolution, yet EDS1 signaling is a derived trait in seed plants. Here, we examined the functional dependency of diverse plant TIRs on the EDS1 pathway in the angiosperms Nicotiana benthamiana and Nicotiana tabacum. While the isolated TIR domains from non-seed plants generally required EDS1 for immune cell death activation, we also identified TIRs that functioned independent of EDS1. However, chimeric TIR-NLRs incorporating these diverse TIR domains onto the AtWRR4a receptor chassis showed a full reversion to EDS1-dependency. Extending this phenomenon further, we demonstrated that the AtWRR4a architecture enforces EDS1-dependence onto a bacterial TIR domain that is otherwise EDS1-independent. Collectively, our work demonstrates that NLR immune receptor architecture influences TIR-related immunity and provides further context to their ancient acquisition into plant immune systems.

Plant diseases remain a major threat to food security, reducing crop yield worldwide. A central challenge in crop protection is that effective immunity often comes at a cost. Plants must continuously balance growth with defense, and this balance complicates molecular breeding strategies aimed at improving disease resistance. When immune pathways are constitutively activated, plants are often better protected against pathogens, but the price for this protection is reduced growth or fitness. This defense-growth trade-off has therefore become a major obstacle to engineering disease resistance in crops.

When organisms encounter pathogens, they rapidly activate complex defense programs to ensure survival. While these immune responses are vital, they often also incur trade-offs, such as reduced growth and development and must therefore be tightly controlled. In this study, we reveal that the steroid hormones brassinosteroids (BRs) contribute to this control in Arabidopsis thaliana by repressing immunity-related genes. We provide evidence that the BR-regulated basic helix-loop-helix (bHLH) transcription factor CESTA (CES), along with its homologs BR ENHANCED EXPRESSION (BEE)1-3, mediate DNA methylation changes at transposable element (TE)-rich loci containing nucleotide-binding leucine-rich-repeat (NLR)-type receptor genes, including SUPPRESSOR OF NPR1-1 CONSTITUTIVE 1 (SNC1). These CES-induced methylation changes correlate with altered splicing of SNC1 pre-mRNA, a process that requires the BR receptor BRASSINOSTEROID INSENSITIVE 1 (BRI1). In support, we show that CES associates with components of the chromatin remodeling and splicing machinery. Together, our findings reveal a previously unrecognized BR-induced mechanism that modulates the epigenetic and post transcriptional regulation of immune genes, enabling plants to prioritize growth over defense.

Effective management of Phytophthora root rot in soybean is compromised by the rapid loss of efficacy of the most widely deployed resistance genes to Phytophthora sojae (Rps). However, some genes such as Rps3a and Rps6 are still offering a strong protection but are nonetheless rare in elite material, probably owing to the fact that they have never been properly characterized. In this study, we have employed RenSeq, and whole genome sequencing to unravel the nature of Rps6 as a complex locus composed of 10 NLR genes. In a cell death assay using soybean protoplasts, we show that one of the candidates from the cluster interacts robustly with Avr6. Transfer of the candidate gene into a susceptible root system confirmed its function and status as the bona fide Rps6. Through sequence comparison with other Rps differentials, we further discovered that Rps3c and Rps4, originally thought to be distinct genes on different chromosomes, are, in fact, the exact same resistance gene as Rps6, mediating recognition to the same effector, Avr6. These results clarify a long-standing confusion regarding the identity of some elusive Rps genes and offer the precise sequence and position of Rps6. At the same time, along with the recently exposed homology between Rps3b and Rps11, these findings raise some concerns about the apparent multiplicity of Rps genes. Indeed, there may be fewer sources of resistance than assumed, which should instill caution in using current Rps genes to ensure durable management of Phytophthora root rot.

Artificial intelligence (AI) systems such as AlphaFold have transformed structural biology by enabling accurate prediction of protein structures. However, their capacity to uncover new classes of macromolecular assemblies remains largely untapped. We developed the Structural Novelty Index (SNI), a quantitative framework for identifying protein complexes that diverge from canonical architectures. As one implementation of SNI, we developed SNINRC-Hexa, to identify unconventional resistosomes formed by nucleotide-binding, leucine-rich repeat immune receptors (NLRs). We used it to analyze AlphaFold 3 models of 637 non-redundant NRC proteins from 346 genomes representing 85 plant species. This analysis identified candidates with predicted architectures distinct from the canonical hexameric resistosomes of NRC proteins. Biochemical purification and negative-stain transmission electron microscopy of NRC7 orthologs from multiple species supported the SNI prediction and revealed an unexpected undecameric (11-mer) assembly. Our results establish SNI as a scalable approach for discovering atypical protein complexes.

Dioecious plants, which have distinct male and female individuals, constitute ~5% of angiosperm species and have emerged frequently and independently from hermaphroditic ancestors. Although recent molecular studies of sex determination have started to reveal the diversity of the genetic systems underlying dioecy, research on the evolution of dioecy is limited, especially in monocots. Here, we explore the molecular basis of sex determination in the monocot Dioscorea tokoro, a dioecious wild yam endemic to East Asia. Chromosome-scale and haplotype-resolved genome assemblies and linkage analysis suggested that this plant has a male heterogametic sex-determination (XY) system, with sex-determination regions located on chromosome 3. Sequence comparison between the X- and Y-chromosomes and read coverage analysis revealed X- and Y-specific regions in putative pericentromeric chromosome regions. Within the Y-specific region, we propose two candidate genes that are likely involved in sex determination: BLH9, encoding a homeobox protein, and HSP90, encoding a molecular chaperone. BLH9 functions in a similar way as AtBLH9 in Arabidopsis thaliana. BLH9 could be involved in suppression of female organ development, whereas HSP90 might be required for pollen development. These results shed light on the complex evolution of dioecy in plants.

Plants secrete a variety of proteases as a defense response during infection by microbial pathogens. However, the relationship between their catalytic activities and antimicrobial functions remains largely unknown. Particularly, few biologically relevant substrates of these proteases have been identified. Huanglongbing (HLB) has been a major threat to the citrus industry worldwide. The HLB-associated bacterium, "Candidatus Liberibacter asiaticus" (Las), was previously shown to deploy an inhibitor of papain-like cysteine proteases (PLCPs) to promote disease in citrus. In this study, we identified an outer membrane protein (OMP) of Las, LasOMP1, as a substrate of the citrus PLCP CsRD21a. LasOMP1 is one of the most highly expressed genes in Las. CsRD21a cleaves LasOMP1 and produces cleaved peptide products, which could be detected in vitro and in HLB-diseased citrus plants. We found that CsRD21a targets the N-terminal portion of LasOMP1, potentially at an extracellular loop region. Importantly, transgenic sweet orange overexpressing CsRD21a showed reduced Las populations and improved plant growth, highlighting that engineering this protease is a promising strategy to enhance HLB resistance in citrus. Together, our work reveals a pathogen-derived substrate of plant PLCPs and suggests bacterial OMPs may be direct targets of plant defense.