Aleksandra Skirycz

Small molecules can control plant disease either by disarming the pathogen or by priming host immunity, but single compounds that do both are rare and mechanistically unexplained. Here we show that hopeaphenol (HP), a resveratrol tetramer from stilbene-producing plants, acts on histidine kinase (HK) signaling on both sides of the Arabidopsis– Pseudomonas interaction. In Pseudomonas syringae pv. tomato DC3000, HP represses the type III secretion regulon and motility genes, restricts surface motility independently of effector delivery, and binds a defined subset of virulence-associated sensor kinases while reducing their autophosphorylation; comparable engagement occurs in Pectobacterium atrosepticum . Binding and inhibition depend on the tetrameric scaffold rather than the resveratrol monomer. In the host, HP binds the CHASE domains of the cytokinin receptors AHK2, AHK3 and AHK4 and elicits an immune-associated, rather than canonical cytokinin, transcriptional output. HP-dependent potentiation of PTI and ETI responses and early restriction of bacterial growth require AHK3 and AHK4. HP thus coordinates opposing outputs from divergent HK systems across kingdoms.

Quinoa ( Chenopodium quinoa ) is a nutrient‐rich pseudocereal with diverse specialized metabolites, yet the genetic basis of this metabolic diversity is poorly understood. Here we integrate whole‐genome sequencing and multi‐tissue metabolic profiling of 603 quinoa accessions. We detected 4688 metabolic features and identified over 1000 metabolites in seeds, leaves, and roots. Using multi‐tissue genome‐wide association, we mapped the genetic architecture of quinoa metabolome by identifying 584 quantitative trait loci (QTL) and prioritized 219 candidate genes across 58 major QTL governing saponin, betalain, and flavonoid biosynthesis. Moreover, we constructed a drought‐responsive multi‐omics regulatory network and uncovered additional key genes involved in quinoa stress signaling and metabolic pathways. Finally, we cloned and functional validated the roles of cytochrome P450 76AD1 (CYP76AD1) in betalamate accumulation, UDP‐glycosyltransferase (UGT91C1) in flavonoid glycosylation, and CYP72A154 and soyasapogenol B glucuronide galactosyltransferase in saponin biosynthesis. This multi‐omic framework provides a high‐resolution map of the quinoa metabolome and a foundation for breeding nutrient‐rich and stress‐resilient quinoa cultivars.

RNA covalent modifications (RCMs) influence RNA stability and translation efficiency, and thus play critical roles in eukaryotic growth and development. However, their role in regulating plant performance under abiotic stress remains largely unexplored. Here, we integrated multi-omics data in six Sorghum bicolor accessions under water-limiting conditions in the field to explore the relationship between RCMs and drought response. Within a stress and photosynthesis-associated gene co-expression module, we identified SbDUS2, a member of a family of enzymes conserved across eukaryotes, that catalyzes the reduction of uracil to dihydrouridine (DHU) on RNA molecules. DHU-modified transcripts in this module were enriched for photosynthetic functions and showed strong correlation with photosynthetic traits. To elucidate the function of this RCM, we characterized loss of function dus2 mutants in Arabidopsis thaliana. Under control conditions, these DHU-deficient mutants exhibited impaired germination and delayed development. Furthermore, under heat or water-limiting conditions, these mutants showed significantly reduced net CO2 assimilation and survival. Using multiple transcriptome-wide RNA stability assays, we demonstrated that transcripts associated with lower DHU levels in a dus2 background generally exhibited increased stability compared to Col-0 controls. Particularly, lack of DUS2 led to the hyperstability of photosynthesis-related transcripts, impeding their turnover and likely preventing proper photosynthetic acclimation during stress. We propose a model where DHU acts as a critical post-transcriptional regulator marking mRNAs for rapid turnover under stress, highlighting an overlooked regulatory layer contributing to plant resilience.

Protein–metabolite interactions (PMIs) are critical regulators of cellular processes, yet their roles in plant–pathogen interactions remain poorly understood. Here, we applied PROMIS (Protein–Metabolite Interactions using Size separation) to map the Arabidopsis protein–metabolite interactome during infection with Pseudomonas syringae pv. tomato DC3000 (Pto DC3000). PROMIS resolved elution profiles for >5,700 proteins and 192 annotated metabolites across infected, systemic, and control tissues, revealing infection-associated remodeling of protein assemblies and metabolite co-fractionation patterns. Integrating co-fractionation with coronatine-induced transcriptional responses, we identified NATA1, a polyamine N-acetyltransferase, as a coronatine-associated host protein binder. Biochemical assays confirmed direct coronatine–NATA1 interaction (Kd = 586 nm), whereas the homolog NATA2 did not bind coronatine. Coronatine did not strongly inhibit NATA1 catalytic turnover under our assay conditions but consistently depressed NATA1 dimer formation in vitro and in planta. In silico docking together with targeted mutational analyses identified a coronatine-sensitive interface on NATA1, revealing residue-specific coupling between ligand binding, assembly state, and acetyl-CoA engagement. Collectively, these findings establish PROMIS as a robust platform for infection-state PMI discovery and uncover a coronatine-responsive regulatory node that links small-molecule signaling to host polyamine acetylation.

Cyclic adenosine monophosphate (cAMP) is a fundamental second messenger involved in diverse signaling pathways across both animals and plants. While the role of 3′,5′-cAMP has been extensively characterized, the biological significance of its structural isomer, 2′,3′-cAMP, remains largely unexplored, particularly in plants. Here, we show that 2′,3′-cAMP and 3′,5′-cAMP represent parallel signaling systems in Arabidopsis thaliana , with different enzymatic origins and largely distinct downstream effects. In vitro enzymatic assays show that plant adenylate cyclases (ACs), including AFB5 and HpAC1, produce specifically 3′,5′-cAMP from ATP, whereas the TIR domain of protein L7 also catalyzes the formation of 2′,3′-cAMP from RNA. Comprehensive multiomics analyses reveal that two isomers elicit distinct yet partially overlapping metabolic, proteomic, and transcriptional response: 2′,3′-cAMP activates broad, stress-adaptive gene expression reprogramming, while 3′,5′-cAMP fine-tunes responses related to nutrient status and cellular homeostasis. Our findings establish the existence of dual cAMP signaling systems in plants, each with specialized functions and provide insights into the complex regulatory networks governing plant physiology.

Plastids are dynamic organelles that remodel their composition, ultrastructure, and function according to developmental and environmental demands. The synthetic molecule X57 induces the conversion of leaf chloroplasts into tocopherol-rich plastids lacking thylakoids and containing proliferating plastoglobules. Removal of X57 triggers chloroplast re-differentiation, enabling precise spatial-temporal dissection of these transitions. X57 directly binds and inhibits the phosphatase SAL1, causing accumulation of its substrate PAP, a retrograde signal that modulates nuclear gene expression. SAL1 inhibition and subsequent PAP accumulation activate a cascade that depletes cytokinins and down-regulates GOLDEN2-LIKE1 (GLK1) and other transcription factors involved in chloroplast biogenesis. SAL1-defective mutants fail to undergo this signaling pathway. The SAL1-PAP-mediated weakening of chloroplast identity preconditions plastids for their eventual conversion into storage-type organelles upon X57-promoted accumulation of tocopherols. After X57 withdrawal, photosynthetic gene expression and chloroplast functions are restored. This framework identifies key molecular mechanisms underlying chloroplast plasticity, a central process in biology.

Climate change and especially the concomitant increasing frequency of drought and salinity induced water-deficit stresses, is a major limiting factor of plant growth and crop productivity worldwide. Understanding the adaptive responses of plants to water-deficit stress has therefore become a central challenge in plant biotechnology. Considerable evidence indicates that late embryogenesis abundant (LEA) proteins contribute to water-deficit stress tolerance and the stabilization of metabolic enzymes, yet the molecular basis of these responses remains unknown. To date, limited direct evidence for specific in vivo interactions between LEA proteins and their molecular targets has been reported. Here, we identify for the first time the native in vivo interactome of three Arabidopsis dehydrin LEA proteins - LEA4 (COR47), LEA5 (ERD10), and LEA10 (ERD14) - expressed under their own promoters and in response to salt stress. Our results show that LEA4, LEA5, and LEA10 dehydrins interact with each other, and share six common candidate protein interactors, suggesting that they form a core protein complex in salt stress conditions. We confirmed the direct protein-protein interaction between LEA5 with LEA4, LEA10, OSCA3, PLD α 1, PIP2B and OST1/AHA1 at the biochemical level. Phenotypic analyses of loss-of-function genetic mutants revealed that the interactions of LEA5 with OSCA3 and PIP2B promote seed germination under salt stress. LEA5 interaction with PLDα1 promotes root growth under salt stress, while its interaction with PIP2B enhances root growth under osmotic stress, indicating distinct stress-specific functional roles for each interaction. In conclusion, this study identifies the native in vivo interactions of three dehydrin proteins, while uncovering the functional relevance of these interactions under salt and osmotic stress conditions, thus providing novel mechanistic insights into the role of LEA proteins in water-deficit stress adaptation.

Mitochondrial disease encompasses a group of genetically inherited disorders hallmarked by an inability of the respiratory chain to produce sufficient ATP. These disorders present with multisystemic pathologies that predominantly impact highly energetic tissues such as skeletal muscle. There is no cure or effective treatment for mitochondrial disease. We have discovered a small molecule known as oxybutynin that can bypass Complex III mitochondrial dysfunction in primary murine and human skeletal muscle progenitor cells (MPCs). Oxybutynin administration improves MPC proliferative capacity, enhances cellular glycolytic function, and improves myotube formation. Mechanistically, results from our isothermal shift assay indicates that oxybutynin interacts with a suite of proteins involved in mRNA processing which then trigger the upregulation biological pathways to circumvent CIII mitochondrial dysfunction. Taken together, we provide evidence for the small molecule oxybutynin as a potential therapeutic candidate for the future treatment of CIII mitochondrial dysfunction.

Dipeptides are structurally diverse small molecules with varying modes of biogenesis and function. In plants, dipeptides were historically associated with nitrogen storage and mobilization; however, they are also reported to act as antioxidants, signaling molecules, protein regulators, and modulators of microbial communities. In this review, we discuss the structural diversity and biogenesis of dipeptides, with respect to (i) proteinogenic dipeptides that are products of protein degradation, (ii) non-proteinogenic amino acid dipeptides, such as those containing a γ-glutamyl group, and (iii) cyclic dipeptides, largely derived from microorganisms. Highlighted are recent examples of dipeptides that were shown to display plant health-promoting activities, including boosting growth and improving stress resilience against pathogens, salinity, chilling, and heat, making the case that these overlooked compounds are more than what meets the eye.

SUMMARY: Mineral nutrient uptake and deposition profoundly influence plant development, stress resilience, and productivity. Silicon (Si), though classified as a non‐essential element, significantly influences a plant's physiology, particularly in fortifying defense responses and mitigating stress. While the genetic and molecular mechanisms of Si uptake and transport are well studied in monocots, particularly rice, their role in dicot species, such as soybean, remains unclear at the cellular and molecular levels. In this study, we utilized single‐nucleus RNA sequencing (snRNA‐seq) to dissect cellular responses to Si accumulation in soybean leaves. We identified distinct cellular populations, including a unique Si‐induced or Si‐associated cell cluster within vascular cells, suggesting a specialized mechanism of Si distribution. Si treatment notably induced the expression of defense‐related genes, with a pronounced enrichment in vascular cells, underscoring their pivotal role in activating plant defense mechanisms. Moreover, Si modulated the expression of genes involved in phytoalexin biosynthesis, salicylic acid, and immune receptor signaling, suggesting transcriptional priming of genes involved in defense responses. Further investigation of Si transporters revealed precise expression of an Si efflux gene in epidermal cells in response to Si treatment. We also validated the role of efflux Si transporters using a Xenopus oocyte assay and CRISPR/Cas9 genome editing of composite soybean plant roots. This study provides critical insights into the biotic stress regulatory networks influenced by Si treatment in soybean leaves at the single‐cell level, thus laying the foundation for enhancing stress tolerance through optimized mineral nutrient uptake.