Myrosin idioblast cells (MIs) are specialized cellular structures that synthesize and store myrosinase, a crucial enzyme in the glucosinolate (GSL)-myrosinase system important for the defense of cruciferous plants agains...Myrosin idioblast cells (MIs) are specialized cellular structures that synthesize and store myrosinase, a crucial enzyme in the glucosinolate (GSL)-myrosinase system important for the defense of cruciferous plants against herbivores and pathogens. Jasmonic acids (JAs) are defense-related phytohormones that increase resistance to insect feeding through promoting GSL biosynthesis, but the detailed mechanism of JAs in MI development remains to be fully explored. Here, we conducted a systematic investigation into the regulatory role of JAs in the development of MI in Arabidopsis (Arabidopsis thaliana). Phenotypic analyses revealed that MeJA exogenous application significantly increases the number and area ratio of MIs in wild-type plants. JAs promoted MI development via the core COI1-JAZ1-MYC2 module, which depends on the key regulator FAMA. Biochemical analysis suggested that at the transcription level, MYC2 directly binds to the promoter region of FAMA, thereby activating expression. Considering protein interactions, MYC2 associates with FAMA and enhances transcription of the downstream target gene bHLH090. Genetic evidence corroborated that the functionality of MYC2 depends on FAMA. Furthermore, JAZ1 functions as a competitive inhibitor of the MYC2-FAMA interaction. Increasing the JA concentration caused FAMA upregulation and JAZ1 degradation, yielding the formation of abundant FAMA-MYC2 complexes, which promote MI development.
Elevated temperatures during grain filling severely constrain rice yield and quality. Although additional nitrogen can mitigate the adverse effects of elevated temperature, both factors increase grain protein content, an...Elevated temperatures during grain filling severely constrain rice yield and quality. Although additional nitrogen can mitigate the adverse effects of elevated temperature, both factors increase grain protein content, and the underlying physiological mechanisms remain poorly understood. Here, we conducted actual field warming (2.28°C day/4.33°C night) during the grain-filling period and applied an additional 60 kg N ha-1. The grain weight of superior spikelets (SS) was 2.1% lower under elevated temperature (ET), while ET increased the grain weight by 4.23% and protein content by 2.95% in inferior spikelets (IS). ET promoted the level of free amino acids, improved the activities of glutamine synthetase and glutamate synthase, and up-regulated the expression of an amino acid transporter gene (OsLHT1), mainly 9-15 days after flowering (DAF), in the leaf, ultimately accelerating leaf senescence post 20 DAF. Regardless of the temperature, the effect of nitrogen on leaves was similar to that of ET, while delaying leaf senescence and further increasing protein content in SS and IS. Metabolomic analysis further confirmed that ET accelerated leaf senescence and amino acid depletion in the leaves. Furthermore, the higher levels of L-histidine, along with increased levels of stress-responsive metabolites (D-raffinose and gentisic acid), collectively contributed to the improved protein content in IS under ET. Overall, the study provides insight into grain protein accumulation under warming.
Cytosolic calcium ion (Ca2+) signals are important for abiotic stress tolerance in plants; however, their generation and regulation remain poorly understood. Here, we show that the rice calmodulin (CaM) protein OsCaM1.1...Cytosolic calcium ion (Ca2+) signals are important for abiotic stress tolerance in plants; however, their generation and regulation remain poorly understood. Here, we show that the rice calmodulin (CaM) protein OsCaM1.1 regulates multiple aspects of Ca2+ signals and modulates abiotic stress tolerance and growth in rice. OsCaM1.1 increases stress tolerance to heat, chilling, salt, and drought, as demonstrated by both its loss-of-function mutants and its overexpression lines. It promotes Ca2+ influx and stomatal closure in response to heat, chilling, salt, and the hormone abscisic acid. OsCaM1.1 can physically interact with multiple Ca2+ channel cyclic nucleotide-gated channel (CNGC) proteins, and overexpression of OsCNGC16 rescued defects of both stress tolerance and stomatal closure in the OsCaM1.1 mutant, indicating that OsCaM1.1 regulates CNGC proteins for its stress tolerance function. Under non-stress conditions, OsCaM1.1 promotes Ca2+ oscillations in root hairs and enhances root hair elongation. It also maintains low basal cytosolic Ca2+ levels under non-stress conditions. These findings indicate that OsCaM1.1 is a regulator of both transient Ca2+ signals and resting or basal Ca2+ signals. OsCaM1.1 likely functions in stress tolerance and regulates transient Ca2+ signals through Ca2+ channel CNGC proteins.
The genetic basis of long-term photoprotection under sustained high-light exposure in perennial trees remains poorly understood. Here, we report that a miniature inverted-repeat transposable element (MITE) located 1.2 kb...The genetic basis of long-term photoprotection under sustained high-light exposure in perennial trees remains poorly understood. Here, we report that a miniature inverted-repeat transposable element (MITE) located 1.2 kb upstream of PsiGCN2 reduces promoter activity by approximately 26% (P < 0.01) in transient heterologous reporter assays, consistent with a potential cis-regulatory contribution in Populus simonii. Genome-wide association study of 334 accessions identified PsiGCN2 (Chr07:14786025) as significantly associated with electron transport rate (ETR, P < 1 × 10-7) and non-photochemical quenching (NPQ, P < 3.72 × 10-7) under high-intensity light stress. Loss-of-function paggcn2 mutants demonstrated enhanced photoprotection during prolonged high-intensity light exposure, including 19% higher maximum PSII efficiency, 46% increased NPQ, and 10 to 20% ETR following 8-d exposure compared to wild-type plants. Transcriptomic profiling revealed 432 differentially expressed genes, with photosynthetic antenna proteins significantly upregulated (NES = 1.86, P < 0.001) and reduced enrichment of mitogen-activated protein kinase signaling components (NES = -1.56, P = 0.0078) in mutants. Nuclear-localized PsiGCN2 is positioned as an integrative regulatory hub, coordinating 514 protein interactions spanning proteostasis, photosynthesis, and RNA metabolism pathways. Genome-wide annotation identified 27,395 MITEs with 42.6% of genes harboring insertions near transcriptional boundaries. Phylogenetic analysis revealed episodic MITE amplification at 16.7 million years ago, coinciding with Mid-Miocene climatic shifts. These findings support a MITE-PsiGCN2-phenotype regulatory axis that influences the balance between photoprotection and photosynthetic efficiency under sustained high-light conditions. They also shed light on how promoter-proximal transposable element insertions contribute to transcriptional modulation of light acclimation in woody species.
Engineering a novel N2-fixing rhizobia symbiosis in cereal crops is a strategy being pursued to improve agricultural sustainability. However, if such a symbiosis were introduced it would have to be economically viable in...Engineering a novel N2-fixing rhizobia symbiosis in cereal crops is a strategy being pursued to improve agricultural sustainability. However, if such a symbiosis were introduced it would have to be economically viable in the context of plants' existing nutrient acquisition strategies, including existing symbioses with arbuscular mycorrhizal fungi (AMF) that most plants already engage in. It is important to understand how the metabolic costs and benefits from these symbioses with overlapping functions might impact plant growth when evaluating the potential benefits of this engineering strategy. To address this, we developed metabolic models describing how the relative growth rate of Zea mays is impacted by the AMF Rhizophagus irregularis and a hypothetical N2-fixing symbiosis with Bradyrhizobium diazoefficiens in isolation and in tandem. The metabolic models of the plant-AMF symbiosis and plant-AMF-rhizobia symbiosis are the first of their kind. To validate the AMF component of our model, we conducted a field evaluation comparing AMF-compatible and mutant AMF-incompatible maize hybrids. The empirically measured AMF-mediated growth benefit agreed well with model predictions. Our model of the rhizobium symbiosis predicted that the lower N content of cereal crops makes the growth penalty associated with acquiring nitrogen from rhizobia smaller than in legumes. Finally, the model of the plant-AMF-rhizobia symbiosis predicted positive synergies between rhizobia and AMF under nutrient-limited conditions but negative synergies under phosphorus-replete conditions. This indicates that these bioengineering strategies could improve cereal crop yields and may achieve greater gains in tandem, but soil nutrient levels and plant nitrogen requirements should be considered.
Cold stress constrains global crop productivity. Although raffinose accumulation confers cold tolerance, the regulatory mechanisms governing its biosynthesis remain largely unresolved. Here, we identify a cold-induced ra...Cold stress constrains global crop productivity. Although raffinose accumulation confers cold tolerance, the regulatory mechanisms governing its biosynthesis remain largely unresolved. Here, we identify a cold-induced raffinose synthase gene (CnRS1) from Campeiostachys nutans-a cold-tolerant perennial grass native to the Qinghai-Tibet Plateau and closely related to cereal crops. Overexpression of CnRS1 significantly enhanced cold tolerance in both C. nutans and transgenic rice (Oryza sativa L.). Using yeast 2-hybrid screening, biochemical interaction, and phosphorylation assays, we demonstrated that CnRS1 interacts with and is phosphorylated by the CBL-interacting protein kinase 31 (CnCIPK31) at Ser606, Thr106, and Thr764, thereby enhancing its protein stability. Furthermore, CnCIPK31 interacted with 2 calcineurin B-like proteins (CnCBL2 and CnCBL4) to form a Ca2+-dependent CBL-CIPK complex, which promotes CnRS1 phosphorylation. CnCIPK31 overexpression increased raffinose accumulation and improved cold tolerance, whereas its silencing caused hypersensitivity, which was rescued by exogenous raffinose. Genetic analysis further established that CnCIPK31 acts upstream of CnRS1 to improve plant cold tolerance. Collectively, our study reveals a Ca2+-responsive CnCBL2/4-CnCIPK31-CnRS1 signaling module that integrates calcium signaling with raffinose biosynthesis to enhance cold tolerance, providing insights into alpine plant adaptation and a strategy for improving crop cold resilience.
Root density, determined by root number, is a key trait for drought resistance and yield improvement. Here, we identified a drought-responsive root number regulator, TabHLH112-2A, in wheat (Triticum aestivum). Given its...Root density, determined by root number, is a key trait for drought resistance and yield improvement. Here, we identified a drought-responsive root number regulator, TabHLH112-2A, in wheat (Triticum aestivum). Given its interaction with the essential factor of crown root initiation, TaMOR, TabHLH112-2A was subsequently designated TaMIP1. mip1 mutants had fewer crown and lateral roots than the wild type (WT). TaMIP1 specifically bound to the E-box cis-element and induced the expression of genes involved in auxin and ABA signaling pathways, root development, and drought stress response. Two TaMIP1 haplotypes were found in the natural population. Two nonsynonymous SNPs in the active domain led to enhanced transactivation activity of TaMIP1Hap-2A-2, resulting in higher root dry weight. The TaMIP1 and TaMOR haplotypes had additive effects on RDW, and the effect of TaMOR haplotypes was epistatic to that of TaMIP1 haplotypes. Furthermore, mip1 exhibited a lower survival rate under drought stress and a higher yield under well-watered conditions. Our findings elucidate the important roles of TaMIP1 in root density and the tradeoff between yield and drought resistance. The discovery of a regulatory module and combined haplotypes bring insights and genetic resources for drought resistance and high-yield breeding.
Bud dormancy release, triggered by winter chilling, is a critical adaptive process that enables temperate and boreal perennial trees to resume growth in spring. Although recent studies have identified FLOWERING LOCUS T1...Bud dormancy release, triggered by winter chilling, is a critical adaptive process that enables temperate and boreal perennial trees to resume growth in spring. Although recent studies have identified FLOWERING LOCUS T1 (FT1) as a key regulator of dormancy release in Populus, the molecular mechanisms underlying FT1 action remain unclear. Here, through comprehensive analysis of the alternative splicing (AS) landscape during dormancy release, we identified a dormancy release-dependent AS event in FT1 that generates 2 isoforms: FT1 and an exon-skipped variant FT1SE. Functional characterization demonstrated that FT1SE acts as a dominant-negative regulator: overexpression of FT1SE delays bud break, whereas silencing accelerates it. Exon skipping introduced a premature stop codon, removing the C-terminal Phosphatidylethanolamine-binding protein (PEBP) domain. Despite this truncation, FT1SE retained interaction with FLOWERING LOCUS D-LIKE1 (FDL1) and competitively interfered with the formation of the FT1-FDL1 activation complex. We further showed that FT1SE production results from a CT-to-AG substitution at the splice acceptor site. Phylogenetic analysis indicated that this nucleotide variant is unique to species within the Populus section and is broadly distributed across cold-adapted habitats, suggesting that FT1 AS represents an evolutionary innovation enabling dormancy adaptation to freezing environments.
While the molecular mechanisms mediating submergence tolerance have been extensively studied, those underpinning age-dependent resilience remain poorly characterized. In Arabidopsis thaliana, submergence elicits a leaf-a...While the molecular mechanisms mediating submergence tolerance have been extensively studied, those underpinning age-dependent resilience remain poorly characterized. In Arabidopsis thaliana, submergence elicits a leaf-age-dependent phenotype in which senescence and death progress across an age gradient starting with older leaves. Here, we sought to investigate the mechanisms mediating the observed differential flood resilience by interrogating leaf-age-specific transcriptome and proteome changes during submergence and recovery. Following submergence, most age-dependent differences were in the magnitude or speed of transcript abundance changes, whereas qualitative leaf-age-dependent responses were most apparent during recovery. This included a strong desiccation response in old leaves despite a stronger Abscisic Acid (ABA)-signaling response. Physiological measurements suggested that faster dehydration was facilitated by submergence-mediated reduction of ABA sensitivity but was not associated with differences in stomatal conductance or cuticle integrity. We also observed a stronger induction in young shoot tissue of genes associated with endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). Mutants disabled in both UPR signaling branches were affected in new leaf formation and in the ability to restore the proteome, but not in senescence, suggesting that young tissues activate ER stress recovery to permit continuation of growth. Among mitochondrial membrane proteins differentially regulated in young leaves, loss of mitochondrial voltage-dependent anion channel function impacted submergence-recovery tolerance. Our data reveal multiple mechanisms underlying leaf-age-dependent differential submergence recovery and demonstrate how an interplay between age-related developmental traits and stress signaling pathways determines tolerance.
Pear (Pyrus) stone cells, a major determinant of fruit quality, form via cell wall lignification. The plant hormone jasmonic acid (JA) plays a pivotal role in regulating secondary cell wall formation, although the underl...Pear (Pyrus) stone cells, a major determinant of fruit quality, form via cell wall lignification. The plant hormone jasmonic acid (JA) plays a pivotal role in regulating secondary cell wall formation, although the underlying molecular mechanisms remain unclear. Here, we demonstrated that JA positively regulates stone cell lignification via respiratory burst oxidase homolog F (PuRBOHF)-dependent reactive oxygen species (ROS) production. The NAC transcription factor PuNAC43 acts as a central mediator, enhancing JA-induced ROS accumulation and lignin deposition. PuNAC43 directly binds to promoters of the PuRBOHF and lignin biosynthesis genes (cinnamoyl-CoA reductase 1, PuCCR1; cinnamyl alcohol dehydrogenase 2, PuCAD2), thereby enhancing their transcription. Additionally, PuMYC2, a central MYC transcription factor in the JA signaling pathway, mediates JA-induced upregulation of PuNAC43 function. PuMYC2 binds directly to the PuNAC43 promoter, activating its expression. Furthermore, PuMYC2 recruits the bHLH transcription factor PubHLH93 through promoter binding and protein-protein interaction, forming a heterodimeric complex that synergistically enhances the activation of PuNAC43. Notably, PuMYC2-mediated transcriptional activation depends on JA levels. Our findings reveal a JA-responsive transcriptional complex, PuMYC2-PubHLH93-PuNAC43, that elevates the transcription of PuRBOHF and lignin-related genes to drive stone cell formation, thereby revealing pivotal mechanisms underlying JA-induced lignification in pear fruit.
Infrared gas analysis (IRGA) is the primary technique for quantifying CO2 uptake in plants, but technical obstacles have limited its application to aquatic species, inhibiting exploration of their photosynthetic mechanis...Infrared gas analysis (IRGA) is the primary technique for quantifying CO2 uptake in plants, but technical obstacles have limited its application to aquatic species, inhibiting exploration of their photosynthetic mechanisms. Using an IRGA with a customised aquatic chamber, we developed a methodology for measuring CO2 responses in duckweed, a group of fast-growing, floating angiosperms. We compared three morphologically contrasting species which have commercial or environmental significance: Wolffia australiana, Spirodela polyrhiza, and Lemna minuta. Light response curves showed species variation in light saturation intensity and light saturated rates of photosynthesis, with W. australiana generally showing the highest rates of exchange per unit area under CO2 limited and CO2 saturated conditions. We estimated biochemical parameters (Vcmax, Jmax) from CO2 response curves, though direct measurement of conductance and intercellular CO2 (Ci) was not possible. Using estimated Ci, predicted Vcmax and Jmax values fell within ranges consistent with other angiosperms, and again W. australiana had the highest values. Temperature response curves suggested an optimum of 35°C. Combining gas exchange and chlorophyll fluorescence further revealed a higher photosynthetic performance in W. australiana grown at lower plant densities. The suite of methodologies presented here will enable future mechanistic studies of duckweed carbon assimilation, growth dynamics and environmental responses, advancing understanding to levels comparable with that of many other terrestrial plant species.
Drought is a major abiotic stress factor that limits tree growth. In this study, we determined the role and underlying mechanism of miR169y during the drought response. Expression analysis revealed that miR169y is notabl...Drought is a major abiotic stress factor that limits tree growth. In this study, we determined the role and underlying mechanism of miR169y during the drought response. Expression analysis revealed that miR169y is notably downregulated under abscisic acid (ABA) and drought treatments. Stable transgenic lines expressing miR169y were generated using Agrobacterium-mediated transformation techniques. Suppression of miR169y expression increased drought tolerance in poplar, whereas its overexpression led to a decrease in tolerance. Nuclear factor Y subunit A6 gene (PtrNFYA6) was identified as the primary target of miR169y, exhibiting considerable upregulation in response to drought and ABA treatments. PtrNFYA6-overexpressing lines exhibited higher relative water content, lower malondialdehyde levels, increased proline accumulation and antioxidant enzyme activities, reduced reactive oxygen species levels, and enhanced stomatal closure. Molecular experiments confirmed that PtrNFYA6 directly binds to the CCAAT-box and ABRE elements in the promoters of 9-cis-epoxycarotenoid dioxygenase genes (PtrNCED3a/3b/6), thereby activating their transcription and promoting ABA synthesis. The overexpression of PtrNCED3b further substantiated the importance of ABA in enhancing drought tolerance. This research introduces a molecular framework in which the miR169y-PtrNFYA6-PtrNCED3a/3b/6 cascade modulates ABA production and drought resilience in poplar, thereby offering crucial targets for the molecular breeding of drought-resistant forest trees.
The Arabidopsis thaliana (Arabidopsis) protein HAKAI-interacting zinc finger protein 1 (HIZ1) is both a transcriptional regulator and a component of the N6-methyladenosine (m6A) writer complex that methylates certain ade...The Arabidopsis thaliana (Arabidopsis) protein HAKAI-interacting zinc finger protein 1 (HIZ1) is both a transcriptional regulator and a component of the N6-methyladenosine (m6A) writer complex that methylates certain adenosines within mRNA. Unlike other components of this complex, HIZ1 is not expressed constitutively throughout the plant, is not required for m6A deposition, and, when ectopically overexpressed, results in reduced levels of m6A. However, the biological functions of HIZ1 remain unclear. Here, we generated additional HIZ1 constitutive overexpression (HIZ1 OX) lines and selected 8 lines, showing varying levels of expression. These plants exhibited pleiotropic developmental defects, the severity of which correlated with expression level. However, higher levels of HIZ1 overexpression did not reduce m6A levels beyond a 25% decrease, consistent with the complete titration of methylase complex interacting partner(s). Using chromatin immunoprecipitation sequencing (ChIP-seq), we showed that HIZ1 is predominantly associated with predicted gene promoter regions and is significantly enriched on genes whose transcripts in wild-type plants are m6A modified. Our results suggest that constitutive overexpression of HIZ1 promotes vegetative growth and disrupts reproductive growth in Arabidopsis. This likely occurs both through m6A inhibition and transcriptional regulation. Thus, HIZ1 may function in an analogous fashion to the mouse zinc finger protein 217 (ZFP217, human homolog ZNF217), which acts as both a transcriptional regulator and a suppressor of the m6A methylation complex.
The successful conquest of land by ancient streptophyte algae is closely related to the structural features and functional roles of their extracellular matrix and associated evolutionary innovations. Terrestrialization 5...The successful conquest of land by ancient streptophyte algae is closely related to the structural features and functional roles of their extracellular matrix and associated evolutionary innovations. Terrestrialization 500+ million years ago required adaptation to multiple and novel abiotic and biotic stressors. The establishment of a sessile habit which encompasses adhesion likely required special cell surface components like arabinogalactan protein-like macromolecules. The secretion of polysaccharide-rich extracellular polymeric substance (EPS) provided multiple services including establishment of a platform for interactions with surrounding microorganisms, i.e., biofilms. Protection against harmful UV radiation likely included production of phenolic compounds, cuticular components and sporopollenin. Innovations to the various components of the cell wall contributed to the evolution of multicellularity and large thallus sizes, water retention, defense, wound response and interactions with surrounding microorganisms. Yet we are only in an infancy stage in understanding how specific cell wall components contributed to the invasion of land by streptophyte algae. Future studies are needed that encompass much larger taxonomic screening, detailed glycomics and proteomics, mining of biosynthetic pathways and comprehensive analyses of cell wall integrity especially in response to abiotic and biotic stressors of terrestrial habitats.
Powdery mildew caused by Erysiphe quercicola is a serious threat to natural rubber (Hevea brasiliensis) production, but the molecular basis of host immunity against this pathogen remains poorly understood. Here, we ident...Powdery mildew caused by Erysiphe quercicola is a serious threat to natural rubber (Hevea brasiliensis) production, but the molecular basis of host immunity against this pathogen remains poorly understood. Here, we identify EqBPIE1 (E. quercicola Broad-spectrum Plant Immunity Elicitor 1), a fungal effector expressed during the mid-to-late colonization stage, that localizes to the cytosol, chloroplasts, and nucleus of host cells. EqBPIE1 enhances chloroplast-derived reactive oxygen species production, promotes perinuclear chloroplast clustering and stromule (stroma-filled tubule) formation, and thereby facilitates the relay of immune signals to the nucleus. In the nucleus, EqBPIE1 interacts with the NLR protein HbRG1 (H. brasiliensis Resistance Gene 1) to trigger hypersensitive cell death. Silencing EqBPIE1 increased susceptibility of H. brasiliensis to E. quercicola, whereas heterologous expression and foliar application conferred broad-spectrum resistance against fungal, bacterial, and oomycete pathogens in multiple plant species. Our findings establish an effector-mediated chloroplast-to-nucleus immune signaling pathway and highlight EqBPIE1 as a promising molecular tool for engineering durable, broad-spectrum disease resistance across crops.
Stomatal responses to changing irradiance have significant impacts on photosynthetic carbon gain as well as on plant water use efficiency. These responses can be broken down into the red or mesophyll response which links...Stomatal responses to changing irradiance have significant impacts on photosynthetic carbon gain as well as on plant water use efficiency. These responses can be broken down into the red or mesophyll response which links stomatal conductance with photosynthesis and the specific blue light response that saturates at low fluence levels and is not linked to photosynthesis. Although the two pathways are thought to operate independently, there is growing evidence that the two are more closely linked than we first appreciated. Understanding both the independence and interplay between red light (RL) and blue light (BL) responses in stomatal behaviour could reveal unexploited but critical targets for sustaining or enhancing intrinsic water-use efficiency (Wi) and crop productivity. In this update review, we synthesise the recent advances in our understanding of the RL and BL stomatal responses and propose future research directions to deepen our mechanistic understanding and inform crop improvement strategies.