Recent studies have shown that microRNA (miRNA) functions are associated with plant responses to water deficiency. Mature miRNAs are loaded onto a complex that includes the ARGONAUTE 1 (AGO1) protein; this complex then cleaves mRNAs or inhibits their translation. Here, we demonstrate that the prion-like domain of AGO1 is responsible for topological changes in AGO1 under dehydration in Arabidopsis thaliana. AGO1 undergoes liquid-liquid phase separation (LLPS), which is driven by intrinsically disordered protein domains and plays diverse roles in cellular processes. LLPS of AGO in the cytoplasm influences miRNA regulatory activity, a process related to cytoplasmic calcium levels. We found that dehydration-induced AGO1 condensation influences AGO1 activity, while contributing to dehydration tolerance in plants. Upon rehydration, the condensation-driven accumulation of AGO1 is resolved, restoring its protein levels to normal. Overall, we propose that AGO1 phase separation acts as an emergent property in response to dehydration, attenuating the energy-consuming miRNA regulatory pathway in young seedlings.

Plant receptor kinases perceive diverse peptide signals to coordinate stress responses and developmental programs. The HAESA-LIKE 3 (HSL3/NUT) receptor recognizes CTNIP/SCREW phytocytokines-disulfide-constrained peptides that regulate immune signaling and stress adaptation. However, how HSL3 distinguishes these structurally constrained peptides from linear signaling molecules remains unknown. Here we report near-atomic resolution cryo-EM structures of HSL3 in apo and CTNIP448-70-bound states at ∼2.6 Å, using Arabidopsis CTNIP4 as a representative family member, revealing the mechanism of disulfide-constrained peptide recognition. The conserved CTNIP motif occupies a negatively charged pocket in the C-terminal region of HSL3 through a combination of polar contacts, hydrogen bonds, salt bridges, and van der Waals interactions. The receptor employs a two-step recognition mechanism-electrostatic steering followed by motif anchoring-that enables rapid ligand capture, consistent with the dynamic nature of stress signaling. Notably, an N-glycan at Asn449 directly contacts the CTNIP4 peptide, establishing glycosylation as an active participant in ligand recognition. Structure-guided mutagenesis combined with reactive oxygen species (ROS) burst assays confirmed the functional importance of key binding interfaces. N-terminal truncation experiments revealed a minimal active fragment: CTNIP451-70 supported both rapid ROS production and sustained seedling growth inhibition, whereas the shorter CTNIP454-70 variant retained ROS activity but failed to trigger long-term seedling growth inhibition. Structure-guided coevolutionary analysis across plant lineages reveals patterns of both conserved and variable receptor-ligand interfaces, highlighting evolutionary flexibility while preserving core features of recognition. These conserved recognition principles, mediated by receptor glycosylation and evolutionary plasticity, enable specificity in peptide signaling, with implications for engineering stress-resilient crops.

Abstract Elevated temperature has been shown to compromise salicylic acid (SA)-mediated immunity in plants. The Arabidopsis thaliana accession C24 retains constitutively elevated SA and resistance to the hemibiotrophic pathogen Pseudomonas syringae pv. tomato DC3000 ( Pst DC3000) at elevated temperature. C24 exhibits reduced biomass compared to that of a commonly studied accession, Col-0, in which SA-mediated immunity is compromised at elevated temperature. Neither the genetic basis of temperature-resilient immunity (TRI) nor the apparent growth-defense tradeoff in C24 is known. Here, we show that a Col-0 x C24 recombinant inbred line (RIL) population resolves TRI to a chromosome 5 locus accounting for most of the mapped genetic variance. This locus (named TRI hereinafter) coincides with a hotspot of structural rearrangement between the two accessions and includes a calcium-sensor gene ( CBL9 ) and several NLR-type paralogs found only in C24. Consistent with a calcium-dependent signaling component, C24 mounts an elevated cytosolic Ca²⁺ response to Pst DC3000. Surprisingly, across the RIL population, disease resistance and biomass are only weakly correlated, with some lines exhibiting both large biomass and high pathogen resistance. These results show that temperature-resilient disease resistance is not only genetically tractable in C24 but also can be uncoupled from biomass cost. The TRI locus in C24 therefore encodes a natural mechanism(s) of temperature-resilient immunity with the growth-defense tradeoff resolved.

Plant receptor kinases perceive diverse peptide signals to coordinate stress responses and developmental programs. The HAESA-LIKE 3 (HSL3/NUT) receptor recognizes CTNIP/SCREW phytocytokines—disulfide-stabilized cyclic peptides that regulate immune signaling and stress adaptation. However, how HSL3 distinguishes these structurally constrained cyclic peptides from linear signaling molecules remains unknown. Here we report near-atomic resolution cryo-EM structures of HSL3 in apo and CTNIP448–70-bound states at ~2.6 Å, using Arabidopsis CTNIP4 as a representative family member, revealing distinct mechanisms for cyclic peptide recognition. The conserved CTNIP motif occupies a negatively charged pocket in HSL3's C-terminal region through a combination of polar contacts, hydrogen bonds, salt bridges, and van der Waals interactions. The receptor employs a two-step recognition mechanism—electrostatic steering followed by motif anchoring—that enables rapid ligand capture and release, consistent with the transient nature of stress signaling. Notably, an N-glycan at Asn449 directly contacts the CTNIP4 peptide, establishing glycosylation as an active participant in ligand recognition. Structure-guided mutagenesis combined with reactive oxygen species (ROS) burst assays confirmed the functional importance of key binding interfaces. N-terminal truncation experiments revealed a minimal active fragment: CTNIP451–70 supported both rapid ROS production and sustained seedling growth inhibition, whereas the shorter CTNIP454–70 variant retained ROS activity but failed to trigger long-term seedling growth inhibition. Structure-guided coevolutionary analysis across plant lineages reveals patterns of both conserved and variable receptor–ligand interfaces, highlighting evolutionary flexibility while preserving core features of recognition. These conserved recognition principles, mediated by receptor glycosylation and evolutionary plasticity, enable specificity in peptide signaling with implications for engineering stress-resilient crops. ### Competing Interest Statement The authors have declared no competing interest. Institute for Basic Science, https://ror.org/00y0zf565, IBS-R021-D1-2025-a00 National Research Foundation of Korea, RS-2024–00338015, RS-2025-25423521

MicroRNAs (miRNAs) are noncoding RNAs, ~21-24 nucleotides in length, that play a pivotal role in post-transcriptional gene regulation by inducing cleavage or translational repression of target mRNAs with complementary sequences. In this study, we identified miRNAs expressed during the early developmental stage of mung bean (Vigna radiata), a major legume crop, using small RNA sequencing (sRNA-seq), and analyzed their expression profiles across various mung bean tissues. Mung bean-specific miRNAs were found to be highly expressed in the aerial parts of seedlings, particularly in the leaves. Furthermore, the expression of these miRNAs was effectively validated using Tailed-Hoogsteen triplex DNA-encapsulated silver nanocluster (DNA/AgNC) sensors. The nanosensor enables rapid detection of target miRNAs within 30 min and is easy to apply for field-based assessments. The predicted target mRNAs of the identified miRNAs were associated with a range of biological processes relevant to early-stage development. This study highlights the potential of nanosensor-based approaches for the efficient identification of novel miRNAs in staple crops, offering a promising strategy to reduce the cost, time, and labor required during the transition from laboratory research to field applications.

Climate warming influences disease development by targeting critical components of the plant immune system, including pattern-triggered immunity (PTI), effector-triggered immunity (ETI) and production of the central defense hormone salicylic acid (SA) at the primary pathogen infection site. However, it is not clear if and/or how temperature impacts systemic immunity. Here we show that pathogen-triggered systemic acquired resistance (SAR) in Arabidopsis thaliana is suppressed at elevated temperature. This was accompanied by global downregulation of SAR-induced genes at elevated temperature. Abolished SAR under warmer conditions was associated with reduced biosynthesis of the SAR metabolite N-hydroxypipecolic acid (NHP) in Arabidopsis and other plant species (such as tomato and rapeseed), as demonstrated by downregulation of NHP biosynthetic genes (ALD1 and FMO1) and reduced NHP and pipecolic acid (Pip) levels. Although multiple SAR signals have been shown previously, exogenous NHP or Pip was sufficient to restore disease protection at elevated temperature, indicating that heat-mediated SAR suppression is due to downregulation of the NHP biosynthetic pathway. Along with ALD1 and FMO1, local and systemic expression of the SA biosynthetic gene ICS1 was also suppressed at warm temperature. Finally, we defined a transcriptional network controlling thermosensitive NHP biosynthesis via the master transcription factors CBP60g and SARD1. Our findings demonstrate that warm temperatures impact not only local but also systemic immunity by impinging on NHP biosynthesis, providing a roadmap toward engineering climate-resilient plant immune systems.

As the climate crisis intensifies, finding strategies to mitigate its cascading effects is now a pressing global priority for both scientists and policymakers. In agriculture and ecology, a key first step is to understand how changing environmental conditions affect plant-microbe interactions, especially given the knowledge gap between findings from controlled experiments and those from field studies. In this review, we highlight known fluctuations in host factors that mediate interactions with surrounding microorganisms under changing climate conditions and discuss potential future directions to alleviate the impacts of climate changes.

Successful resistance to disease-causing pathogens is underpinned by properly regulated immune signalling and defence responses in plants. The plant immune system is controlled at multiple levels of gene and protein regulation-from chromatin-associated epigenetic processes to protein post-translational modifications. Optimal fine-tuning of plant immune signalling and responses is important to prevent plant disease development, which is being exacerbated by a globally changing climate. In this review, we focus on how changing climatic factors mechanistically intercept plant immunity at different levels of regulation (chromatin, transcriptional, post-transcriptional, translational, and post-translational). We specifically highlight recent studies that have provided molecular insights into critically important climate-sensitive nodes and mechanisms of the plant immune system. We then propose several potential future directions to build climate-resilient plant disease resistance using cutting-edge biotechnology. Overall, this conceptual understanding and promising biotechnological advances provide a foundational platform towards novel approaches to engineer plant immune resilience.

Reactive oxygen species (ROS) are crucial in plant growth, defense, and stress responses, making them vital for improving crop resilience. Various ROS sensing methods for plants have been developed to detect ROS in vitro and in vivo. However, each method comes its own advantages and disadvantages, leading to an increasing demand for a simple and effective sensory system for ROS detection in plants. Here, we introduce novel DNA silver nanoclusters (DNA/AgNCs) sensors for visualizing ROS in plants. Two sensors, C20/AgNCs and FAM-C20/AgNCs-Cy5, detect intracellular ROS signaling in response to stimuli such as abscisic acid, salicylic acid, ethylene, and bacterial peptide elicitor flg22. Notably, FAM-C20/AgNCs-Cy5 exceeds the sensing capabilities of HyPer7, a widely recognized ROS sensor. Taken together, we suggest that fluorescent i-motif DNA/AgNCs system is an effective tool for visualizing ROS signals in plant cells. This advancement is important to advancing our understanding of ROS-mediated processes in plant biology.

Calcium ions act as secondary messengers in diverse signaling pathways in plants throughout their life cycle. Studies have revealed that calcium is involved in developmental events and in responses to external stimuli, such as biotic and abiotic stresses. Cellular calcium ion levels are tightly controlled by intricate molecular machinery such as calcium channels and pumps. Transient and spatial fluctuations in calcium levels are subsequently recognized by diverse calcium-decoding molecules, resulting in signal transduction. In this review, we highlight recent findings on natural variations in genes controlling calcium signaling in diverse plant biological processes. We then show how the calcium ion context is utilized by fine-tuning the natural variation in centrally important genes.

A novel fluorescent i-motif DNA silver nanoclusters system has been developed for visualization of reactive oxygen species in plants, enabling the detection of intracellular signaling in plant cells. Reactive oxygen species (ROS) are crucial in plant growth, defense, and stress responses, making them vital for improving crop resilience. Various ROS sensing methods for plants have been developed to detect ROS in vitro and in vivo. However, each method comes its own advantages and disadvantages, leading to an increasing demand for a simple and effective sensory system for ROS detection in plants. Here, we introduce novel DNA silver nanoclusters (DNA/AgNCs) sensors for visualizing ROS in plants. Two sensors, C20/AgNCs and FAM-C20/AgNCs-Cy5, detect intracellular ROS signaling in response to stimuli, such as abscisic acid, salicylic acid, ethylene, and bacterial peptide elicitor flg22. Notably, FAM-C20/AgNCs-Cy5 exceeds the sensing capabilities of HyPer7, a widely recognized ROS sensor. Taken together, we suggest that fluorescent i-motif DNA/AgNCs system is an effective tool for visualizing ROS signals in plant cells. This advancement is important to advancing our understanding of ROS-mediated processes in plant biology.

SUMMARYACCELERATED CELL DEATH 6 (ACD6) mediates a trade-off between growth and defense inArabidopsis thaliana. However, the precise biochemical mechanism by which ACD6 and related proteins in plants act remains enigmatic. Here, we identified two loci,MODULATOR OF HYPERACTIVE ACD6 1(MHA1) and its paralogMHA1-LIKE(MHA1L), that code for ∼7 kDa proteins that differentially interact with specific ACD6 variants. MHA1L enhances accumulation of an ACD6 complex, thereby increasing activity of theACD6standard allele for regulating plant growth and defenses. ACD6 is a multipass transmembrane protein with intracellular ankyrin repeats that are structurally similar to those found in mammalian ion channels. Several lines of evidence link increased ACD6 activity to enhanced calcium influx, likely mediated by ACD6 itself and with MHA1L as a direct regulator of ACD6.

Water-deficit affects nearly every biological event in plants, and recent studies have shown that microRNA-functionality is associated with plant responses to water-deficiency. Liquid-liquid phase separation facilitates the condensation of biomolecules, which is driven by intrinsically disordered proteins and plays diverse roles in cellular processes. Here, we show that the prion-like domain (PrLD) of ARGONAUTE 1 is responsible for topological changes from liquid droplets to solid-condensations of AGO1 under dehydration. Unlike SERRATE, which forms functional RNP granules for miRNA biogenesis, AGO1 RNP granules are non-functional condensates, which is particularly facilitated by cytoplasmic calcium ions. We found that dehydration-induced AGO1 condensation inhibits RNA-induced Silencing Complex (RISC) activity. Following rewatering, the condensed AGO1 is degraded through three consecutive proteolytic processes, indicating that the liquid-to-solid phase transition of AGO1 is a reversible process. Overall, we propose that AGO1 phase transition may serve as a sensor for intense dehydration and attenuates the energy-consuming miRNA-regulatory pathway.

Salicylic acid (SA) is a central plant hormone mediating immunity, growth, and development. Recently, studies have highlighted the sensitivity of the SA pathway to changing climatic factors and the plant microbiome. Here we summarize organizing principles and themes in the regulation of SA biosynthesis, signaling, and metabolism by changing abiotic/biotic environments, focusing on molecular nodes governing SA pathway vulnerability or resilience. We especially highlight advances in the thermosensitive mechanisms underpinning SA-mediated immunity, including differential regulation of key transcription factors (e.g., CAMTAs, CBP60g, SARD1, bHLH059), selective protein-protein interactions of the SA receptor NPR1, and dynamic phase separation of the recently identified GBPL3 biomolecular condensates. Together, these nodes form a biochemical paradigm for how the external environment impinges on the SA pathway.

Extreme weather conditions associated with climate change affect many aspects of plant and animal life, including the response to infectious diseases. Production of salicylic acid (SA), a central plant defence hormone 1–3 , is particularly vulnerable to suppression by short periods of hot weather above the normal plant growth temperature range via an unknown mechanism 4–7 . Here we show that suppression of SA production in Arabidopsis thaliana at 28 °C is independent of PHYTOCHROME B 8,9 (phyB) and EARLY FLOWERING 3 10 (ELF3), which regulate thermo-responsive plant growth and development. Instead, we found that formation of GUANYLATE BINDING PROTEIN-LIKE 3 (GBPL3) defence-activated biomolecular condensates 11 (GDACs) was reduced at the higher growth temperature. The altered GDAC formation in vivo is linked to impaired recruitment of GBPL3 and SA-associated Mediator subunits to the promoters of CBP60g and SARD1 , which encode master immune transcription factors. Unlike many other SA signalling components, including the SA receptor and biosynthetic genes, optimized CBP60g expression was sufficient to broadly restore SA production, basal immunity and effector-triggered immunity at the elevated growth temperature without significant growth trade-offs. CBP60g family transcription factors are widely conserved in plants 12 . These results have implications for safeguarding the plant immune system as well as understanding the concept of the plant–pathogen–environment disease triangle and the emergence of new disease epidemics in a warming climate.

Drought stress has detrimental effects on plants. Although the abscisic acid (ABA)-mediated drought response is well established, defensive mechanisms to cope with dehydration-induced proteotoxicity have been rarely studied. DRR1 was identified as an Arabidopsis drought-induced gene encoding an ER-localized RING-type E3 Ub ligase. Suppression of DRR1 markedly reduced tolerance to drought and proteotoxic stress without altering ABA-mediated germination and stomatal movement. Proteotoxicity- and dehydration-induced insoluble ubiquitinated protein accumulation was more obvious in DRR1 loss-of-function plants than in wild-type plants. These results suggest that DRR1 is involved in an ABA-independent drought stress response possibly through the mitigation of dehydration-induced proteotoxic stress.

A grand challenge facing plant scientists today is to find innovative solutions to increase global crop production in the context of an increasingly warming climate. A major roadblock to global food sufficiency is persistent loss of crops to plant diseases and insect infestations. The United Nations has declared 2020 as the International Year of Plant Health. For historical reasons, molecular studies of plant-biotic interactions in the past several decades have not paid enough attention to how variable climate conditions affect plant-biotic interactions. Here, we highlight a few recent studies that begin to reveal how major climatic drivers impact the plant immune system, particularly secondary messenger and defense hormone signaling, and discuss possible approaches toward engineering climate-resilient plant immunity as part of an ongoing global effort to design 'dream' crops of the future.

Ubiquitination is a critical post-translational protein modification that has been implicated in diverse cellular processes, including abiotic stress responses, in plants. In the present study, we identified and characterized a T-DNA insertion mutant in the At5g10650 locus. Compared to wild-type Arabidopsis plants, at5g10650 progeny were hyposensitive to ABA at the germination stage. At5g10650 possessed a single C-terminal C3HC4-type Really Interesting New Gene (RING) motif, which was essential for ABA-mediated germination and E3 ligase activity in vitro. At5g10650 was closely associated with microtubules and microtubule-associated proteins in Arabidopsis and tobacco leaf cells. Localization of At5g10650 to the nucleus was frequently observed. Unexpectedly, At5g10650 was identified as JAV1-ASSOCIATED UBIQUITIN LIGASE1 (JUL1), which was recently reported to participate in the jasmonate signaling pathway. The jul1 knockout plants exhibited impaired ABA-promoted stomatal closure. In addition, stomatal closure could not be induced by hydrogen peroxide and calcium in jul1 plants. jul1 guard cells accumulated wild-type levels of H2 O2 after ABA treatment. These findings indicated that JUL1 acts downstream of H2 O2 and calcium in the ABA-mediated stomatal closure pathway. Typical radial arrays of microtubules were maintained in jul1 guard cells after exposure to ABA, H2 O2 , and calcium, which in turn resulted in ABA-hyposensitive stomatal movements. Finally, jul1 plants were markedly more susceptible to drought stress than wild-type plants. Overall, our results suggest that the Arabidopsis RING E3 ligase JUL1 plays a critical role in ABA-mediated microtubule disorganization, stomatal closure, and tolerance to drought stress.

The first layer of the plant immune system comprises plasma membrane-localized receptor proteins and intracellular receptors of the nucleotide-binding leucine-rich repeat protein superfamily. Together, these immune receptors act as a network of surveillance machines in recognizing extracellular and intracellular pathogen invasion-derived molecules, ranging from conserved structural epitopes to virulence-promoting effectors. Successful pathogen recognition leads to physiological and molecular changes in the host plants, which are critical for counteracting and defending against biotic attack. A breadth of significant insights and conceptual advances have been derived from decades of research in various model plant species regarding the structural complexity, functional diversity, and regulatory mechanisms of these plant immune receptors. In this article, we review the current state-of-the-art of how these host surveillance proteins function and how they are regulated. We will focus on the latest progress made in plant species belonging to the Solanaceae family, because of their tremendous importance as model organisms and agriculturally valuable crops.

MISFOLDED PROTEIN SENSING RING1 (MPSR1) is a chaperone-independent E3 ubiquitin ligase that participates in protein quality control by eliminating misfolded proteins in Arabidopsis (Arabidopsis thaliana). Here, we report that in the early stages of proteotoxic stress, cellular levels of MPSR1 increased immediately, whereas levels of HEAT SHOCK PROTEIN90.1 (AtHSP90.1) were unaltered despite massively upregulated transcription. At this stage, the gene-silencing pathway mediated by microRNA 414 (miR414) suppressed AtHSP90.1 translation. By contrast, under prolonged stress, AtHSP90.1 was not suppressed, and instead competed with MPSR1 to act on misfolded proteins, promoting the destruction of MPSR1. Deficiency or excess of MPSR1 significantly abolished or intensified the suppression of AtHSP90.1, respectively. Similar to the MPSR1-overexpressing transgenic plants, the miR414-overexpressing plants showed an increased tolerance to proteotoxic stress as compared to the wild-type plants. Although the functional relationship between MPSR1 and miR414 remains unclear, both MPSR1 and miR414 demonstrated negative modulation of the expression of AtHSP90.1. The inverse correlation between MPSR1 and AtHSP90.1 via miR414 may adjust the set-point of the HSP90-mediated protein quality control process in response to increasing stress intensity in Arabidopsis.