polyglutamic acid-functionalized carbon dots improve Cadmium tolerance in Houttuynia cordata by reducing Cadmium accumulation, restoring physiological functions, and reshaping rhizosphere microbial communities. Cadmium (Cd) pollution significantly inhibits the growth and development of H. cordata and poses a serious threat to the safe production of this medicinal plant. In this study, polyglutamic acid-functionalized carbon dots (PGA-CDs) were synthesized by the hydrothermal method, and the mechanism of their role in alleviating Cd stress in H. cordata was systematically investigated. The results showed that compared with the Cd group, the biomass of H. cordata in the Cd+PGA-CDs group significantly increased, and the Cd2+ concentration in the plant decreased. At the same time, PGA-CDs effectively removed reactive oxygen species and regulated the activity of related antioxidant enzymes to alleviate oxidative damage. Moreover, PGA-CDs significantly alleviated the damage to the ultrastructure of chloroplasts caused by Cd stress and enhanced the photosynthetic capacity of the plants. Transcriptome analysis indicated that PGA-CDs treatment significantly changed the expression patterns of genes related to photosynthesis, secondary metabolism, lipid metabolism, and signal response, and regulated the expression of multiple transcription factors and genes related to metal ion homeostasis and transport. Additionally, PGA-CDs increased the α diversity of the rhizosphere microbial community and promoted the enrichment of microbial groups related to plant symbiosis or environmental adaptation, such as the Pseudomonadota, Bacteroidota and Verrucomicrobiota. Through integrated analysis, it further revealed the potential synergistic relationship between gene expression changes, rhizosphere microbial composition, and plant physiological indicators. This study provides new insights into the use of nanomaterials to enhance the adaptability of plants to heavy metals stress.
DNA aptamers selected against PSTVd inhibited or promoted PSTVd infection in co-inoculated assays, correlating with their specific binding sites predicted in silico. Viroids are noncoding plant pathogens consisting of highly structured, single-stranded, circular RNA approximately 230–430 nucleotides. To develop DNA aptamers that specifically bind to potato spindle tuber viroid (PSTVd), a systematic evolution of ligands by exponential enrichment (SELEX) was performed from a random population of 30-nucleotide single-stranded DNAs. Amplicon analysis by next-generation sequencing of every five rounds up to 15 SELEX rounds revealed that the random DNA population became progressively enriched and less diverse at each sampling point; that is, the number of unique sequences detected in the 15th round was 1/18th of that in the 5th round. Sixteen sequences ranked high in 5th, 10th, and 15th rounds, as well as two low abundance sequences in 5th round, were selected to analyze the affinities to PSTVd by pull-down assay. Among them, seven bound either in vitro-transcribed or native PSTVd. The strongest binder was the one from low abundance in 5th round, while five were those ranked high in 15th round. Four sequences, including the one previously reported, were selected and further analyzed for their affinities to PSTVd by co-inoculation assay on tomato plants with infectious PSTVd transcripts. Some significantly inhibited PSTVd infection, whereas others promoted it, suggesting that aptamers may affect viroid infectivity.
A major chromosome 2 region is associated with the genotype-dependent transient GFP expression after Agrobacterium infiltration of rose petals and provides markers for selecting permissive genotypes. Transient expression assays are valuable for testing gene function in rose, where stable transformation remains laborious and genotype-dependent. We evaluated Agrobacterium-mediated transient green fluorescent protein (GFP) expression in the petals of 96 rose genotypes and used genome-wide association analysis to identify associated host genomic regions. GFP expression was scored semiquantitatively on a scale from 0 to 4 at 3 and 5 days post-infiltration (dpi). The genotypes differed strongly in expression at both time points, and the 3- and 5-dpi scores were strongly correlated (r = 0.90, p < 0.001). At 5 dpi, the mean scores ranged from 0 to 3.98, with the cultivars Sebastian Kneipp, Friesia and Comtessa AL among the most permissive genotypes. A genome-wide association study (GWAS) of 37,161 high-quality single-nucleotide polymorphisms (SNPs) revealed a major association on chromosome 2 at 69–73 Mbp and a second peak on unanchored chromosome 0 contigs that may correspond to the same region. The top SNPs reached -log10(P) = 7.85. This identified interval represents a high-priority candidate region containing multiple potential host factors. Candidate annotations within the region include genes related to auxin transport, ubiquitin-mediated protein turnover, ribonucleic acid (RNA) metabolism, membrane-associated defence responses and nuclear transport. The results identify useful rose genotypes for petal transient assays and provide a genetic entry point for dissecting host control of Agrobacterium-mediated transient expression.
The development of CRISPR multiplex genome-editing (MGE) tools is rapidly transforming plant functional genomics and accelerating crop improvements. By simultaneously targeting two or more DNA loci, it allows scientists to precisely edit multiple genes at the single-nucleotide level, within the target genome. Simultaneous manipulation of multiple targets has revolutionized the functional elucidation studies, particularly the dissection of complex genetic pathways. Due to its superior precision and feasibility, CRISPR-MGE is widely accepted and has largely replaced alternative editing tools such as TALENs and ZFNs. Several CRISPR-MGE strategies, including the use of individual expression cassettes, tRNA-processing enzymes, Csy4 or ribozymes, have been successfully deployed in plants. Recent advancements, such as Cpf1, transgene-free methods, or ultra-multiplexing approaches, have further refined the technology into a powerful, efficient, and robust toolkit. MGE enables complex genome editing, including multiple-gene knockouts, base alterations, transcriptional regulation, metabolic engineering, or their combinations. Consequently, it is ideal for elucidating the function of transcription factors that are key molecular players in regulating diverse plant responses, especially in stress pathways. Several stress-responsive TFs have been functionally characterized via CRISPR-MGE, and more advanced tools are being employed. This review evaluates multiplexing tools, their diverse applications, and the current progress toward developing advanced MGE tools. Ultimately, we provide evidence to encourage the use of advanced MGE tools for functional characterization studies of stress-responsive TFs, thereby highlighting their potential to accelerate crop improvement.
CgMYC2 links jasmonate signaling to naringin biosynthesis by binding G-box motifs and activating flavonoid-pathway promoters in Citrus grandis ‘Tomentosa’ Naringin, the predominant bitter-flavanone glycoside in Citrus grandis 'Tomentosa', has well-characterized biosynthetic enzymes, yet the transcriptional regulators coupling hormonal signals to pathway activation remain poorly understood. We demonstrate that CgMYC2, a jasmonate-responsive bHLH transcription factor, functions as a central activator of naringin biosynthesis. Exogenous methyl jasmonate (MeJA) treatment increased naringin content 3.45-fold in seedlings, coinciding with a rapid 6.6-fold induction of CgMYC2 that preceded the peak transcription of five core biosynthetic genes (CgPAL5, CgCHS, CgFNS, Cg7GlcT, and Cg1,2RhaT). Physical interaction between CgMYC2 and the JAZ protein CgJAZ3 was confirmed by pull-down and Co-IP assays, placing CgMYC2 within the canonical jasmonate signaling cascade. Y1H confirmed CgMYC2 binding to the Cg1,2RhaT promoter, EMSA demonstrated direct G-box-dependent binding to all five pathway promoters, and dual-luciferase assays showed transactivation of all five promoters, with the strongest activation for CgCHS. As complementary chromatin-level support, a single-sample CUT Tag profile revealed G-box-enriched CgMYC2-associated chromatin regions across jasmonate-responsive and secondary-metabolic loci. Virus-induced gene silencing (VIGS) of CgMYC2 reduced naringin content by 21
An efficient genetic transformation platform enables functional validation of PgMYB10, identifying it as a master regulator governing anthocyanin biosynthesis in pomegranate. Limited availability of stable genetic transformation systems restricts functional genomics research in pomegranate. Here, we established efficient in vitro regeneration and Agrobacterium tumefaciens-mediated transformation systems for ‘Taishanhong’ pomegranate using stem segment explants. Optimized medium combinations produced high-frequency regeneration: a 93.3
Hormone signaling and MADS-box genes regulate grapevine tendril and inflorescence growth divergence, offering molecular insights for managing tendril growth. Grapevine (Vitis vinifera L.) tendrils and inflorescences are homologous organs; however, their divergent development has important agronomic consequences because excessive tendril growth increases vineyard management costs. To explore the regulatory mechanisms, we compared the inflorescence-prone cultivar 'Einset Seedless' (ENT) with the tendril-prone cultivar 'Pinot Noir' (PN) using anatomical observation, transcriptome analysis of specific tendril nodes, and functional characterization of MADS-box genes. ENT exhibited a higher flowering rate at tendril nodes 1–4 than PN. Transcriptome profiling of specific tendril nodes uncovered 549 differentially expressed genes (DEGs) through an intersection/exclusion strategy, with Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment indicating that hormone and mitogen-activated protein kinase (MAPK) signaling were the primary candidates driving the divergence. To assess the spatiotemporal dynamics of these DEGs, we performed Mfuzz clustering, which revealed that multiple expression trajectories were highly consistent with the flowering gradient across different ENT and PN nodes. Plant hormone signal transduction was the predominantly enriched pathway across all dynamic clusters, highlighting the centrality of phytohormones in this process. Guided by this transcriptional evidence, we measured endogenous zeatin and gibberellin (GA₃) contents in the nodal tissues. Remarkably, the zeatin-to-GA₃ ratio not only paralleled the flowering gradient but also correlated with the cluster expression trajectories, providing physiological evidence for a cytokinin–gibberellin interaction model governing organ divergence. Additionally, we analyzed the differentially expressed transcription factors among the DEGs and identified a MADS-box gene, FRUITFULL-LIKE (VvFUL-L), which was markedly upregulated in PN tendrils. Heterologous overexpression of VvFUL-L in arabidopsis promoted early flowering and reduced inflorescence branching, suggesting its potential role in regulating lateral meristem development and affecting tendril formation. Collectively, these findings establish that Hormone Signaling, particularly cytokinin–GA crosstalk, and MADS-box regulators, such as VvFUL-L, are key regulators of inflorescence versus tendril growth in grapevines, providing a basis for future molecular and breeding studies.
The first T2T reference genome of Guazuma ulmifolia is reported, which serves as a core genomic resource for stress adaptation research and stress-tolerant breeding in cacao wild relatives. Climate change, particularly increased incidence of drought, poses a major threat to food security. Understanding the genomic basis of environmental adaptation in crop wild relatives can provide valuable resources for improving stress resilience. Guazuma ulmifolia, a wild relative of Theobroma cacao with important ecological and medicinal value, lacks high-quality reference genomic resources. Here, we report the first telomere-to-telomere (T2T) chromosome-level genome assembly of G. ulmifolia, with a genome size of 311.31 Mb, contig N50 of 35.19 Mb, and 98.70
Agriculture must achieve substantial yield gains while drastically reducing its environmental footprint to meet the food demands of a growing global population by 2050. Current crop losses to pests and pathogens account for 20–40
The ARM repeat only gene VlARO positively regulates grape resistance to gray mold by promoting the guaijaverin accumulation, which directly suppresses the mycelial growth of Botrytis cinerea. Gray mold, caused by the necrotrophic fungus Botrytis cinerea, severely threatens global grapevine production. Here, we identified VlARO, a nuclear-localized gene from the highly resistant grapevine variety ‘Beta’, as a positive regulator of gray mold resistance. Although VlARO contains Armadillo repeats, it lacks a canonical U-box domain, thus distinguishing it from typical plant U-box E3 ubiquitin ligases. Overexpression of VlARO in grape leaves significantly enhanced resistance to B. cinerea, whereas its silencing weakened this resistance. Stable overexpression in grape calli and transgenic Arabidopsis consistently conferred enhanced resistance, as evidenced by markedly reduced lesion formation. Mechanistically, VlARO significantly activated antioxidant enzymes, resulting in efficient ROS scavenging and reduced oxidative damage. Metabolomic analysis revealed that the overexpression of VlARO profoundly reshaped the metabolic landscape following B. cinerea inoculation, with the flavonoid biosynthesis pathway most prominently enriched. Notably, the accumulation of guaijaverin, a flavonoid glycoside, showed strong positive correlation with VlARO transcript levels. Further results demonstrated that guaijaverin directly inhibits the mycelial growth of B. cinerea in vitro and that its exogenous application significantly reduced disease severity in grape calli. Our findings establish the mechanism for VlARO in grapevine immunity by promoting guaijaverin accumulation. This study provides a promising genetic target for disease-resistant grapevine breeding and lays a theoretical foundation for developing eco-friendly fungicides based on guaijaverin.
TEX1 contributes to mRNA export and SA-mediated defense while modulating ABA-driven susceptibility, linking RNA processing to immune regulation in Arabidopsis TEX1, a subunit of the multi-protein THO/TREX complex conserved in eukaryotes, facilitates the synthesis, splicing, and nuclear-cytoplasmic export of mRNAs. However, its mechanism in plants remains to be thoroughly established, particularly its contribution to plant immunity and the mRNA export mechanism in response to pathogen challenges. Here, we showed that TEX1 contributes to mRNA export and hormonal signaling during pathogen infection. The TEX1 mutant was impaired in plant resistance to the Pseudomonas syringae pv. tomato DC3000 and exhibited a compromised salicylic acid (SA) accumulation and reduced expression of SA-regulated genes. Moreover, we show that the pattern-triggered immunity (PTI) response was compromised in the tex1-4 mutant, as it contributes to a reduced defense phenotype upon flg22, leading to reduced flg22-induced defense responses. TEX1 also positively regulates effector-triggered immunity (ETI) against Pto carrying AvrRpm1 and AvrRpt2. Epistasis analysis revealed that TEX1-mediated resistance is SA-dependent. The tex1-4 mutant accumulated elevated abscisic acid (ABA) levels and increased ABA-mediated susceptibility, accompanied by enhanced ABA biosynthesis and reduced expression of catabolic genes. Analysis with sid2-2 further indicated that ABA-induced susceptibility depends on SA signaling, supporting SA-ABA antagonism. Furthermore, the tex1-4 mutation exhibited defective bulk nuclear mRNA export to the cytoplasm upon pathogen infection, as observed by in situ hybridization using 48-mer fluorescein-labeled oligo(dT) probe. Thus, our findings suggest that mRNA export is a crucial component of plant immune activation, with TEX1 serving as a contributing factor that mechanistically links mRNA export with hormone-regulated immune signaling to promote effective plant defense.
Black carbon is an underappreciated compound agricultural stressor that simultaneously disrupts plant physiology, agroecosystem functioning, andcrop productivity, highlighting the need for realistic exposure assessment and integrated strategies for climate-resilient agriculture. Black carbon (BC), a carbonaceous particulate generated through the incomplete combustion of fossil fuels, biomass, and biofuels, is recognized as a major short-lived climate pollutant with significant implications for atmospheric processes and agricultural sustainability. Unlike engineered biochar, atmospheric BC acts as an environmental stressor through deposition on plant surfaces and accumulation in agroecosystems, yet its direct impacts on crop physiology remain insufficiently understood. This review synthesizes current knowledge on the physicochemical characteristics, environmental pathways, and plant stress mechanisms associated with atmospheric BC while explicitly distinguishing it from intentionally applied biochar. Available evidence indicates that BC influences plant performance through multiple interconnected pathways, including reduced light availability, stomatal obstruction, disruption of photosynthesis, oxidative stress induced by excessive reactive oxygen species (ROS), chloroplast dysfunction, hormonal imbalance, and alterations in nutrient cycling and soil microbial communities. However, much of the mechanistic evidence is derived from studies on biochar, carbon nanomaterials, or other particulate pollutants, highlighting a critical gap in plant-specific evidence under realistic atmospheric BC exposure. Regional studies, particularly from the Indo-Gangetic Plain, demonstrate that elevated BC and associated aerosol loading contribute to reduced crop productivity and increased food security risks, although these impacts often reflect the combined influence of multiple atmospheric stressors rather than BC alone. The review further examines current limitations in BC exposure quantification, emphasizing the absence of agronomic dose–response thresholds and standardized field-based assessment methods. Emerging approaches, including leaf-based biomonitoring and isotopic analyses, are discussed as promising tools for future exposure assessment. Finally, key research priorities are identified, including the generation of plant-specific mechanistic evidence, development of realistic dose–response frameworks, integration of BC into crop simulation models, and implementation of long-term field studies to improve risk assessment and support evidence-based mitigation strategies for sustainable agriculture. Conceptual overview of black carbon as a multidimensional agricultural stressor
Integrated physiological, biochemical, histochemical, and structural analyses demonstrated that exogenous kinetin effectively alleviates arsenic-induced oxidative damage in Ocimum basilicum. Arsenic (As) contamination poses a severe threat to crop productivity by inducing oxidative stress through excessive reactive oxygen species (ROS) accumulation. This study aimed to investigate the protective role of exogenous kinetin (KN), a synthetic cytokinin, in alleviating As-induced phytotoxicity in Ocimum basilicum L. through a comprehensive morphological, physiological, biochemical, histochemical, and structural approach. Plants were exposed to 5 and 10 mg kg−1 As stress concentrations, with or without exogenous 5 µM KN supplementation. Key parameters assessed included growth and biomass, photosynthetic parameters, ROS accumulation, lipid peroxidation, antioxidant enzyme activities, non-enzymatic antioxidants, and leaf microstructure. As stress significantly inhibited plant growth, reduced soil plant analysis development (SPAD) chlorophyll values, and caused oxidative damage, as indicated by elevated levels of malondialdehyde (MDA), hydrogen peroxide (H2O2), and superoxide radicals (O2•⁻). The activities of enzymatic and non-enzymatic antioxidant enzymes were considerably enhanced under As exposure. Microscopic analysis of roots revealed enhanced ROS accumulation along with compromised leaf epidermal integrity under As stress. Exogenous KN application reversed these effects by restoring growth parameters, enhancing photosynthetic and chlorophyll fluorescence parameters, reinforcing membrane integrity, and significantly upregulating both enzymatic and non-enzymatic antioxidant defense systems. KN further promoted proline accumulation and preserved the structural integrity of root and leaf tissues disrupted by As toxicity. These findings collectively establish that exogenous KN effectively confers As stress tolerance in O. basilicum through coordinated ROS homeostasis, enhanced antioxidant defense, and structural preservation. This study highlights KN as a promising and cost-effective phyto-protectant strategy for sustaining the productivity, biochemical performance, and medicinal quality of O. basilicum cultivated in As-contaminated soils, with broader implications for safe herb production and sustainable agriculture.
We established a non-sterile root transformation system in peach seedlings. Using this system, we demonstrated that BA treatment inhibits plant growth and lateral root emergence by SA-mediated disruption of auxin distribution. Allelopathic autotoxins, particularly benzoic acid (BA), are recognized as primary contributors to peach (Prunus persica) replant disease; however, the molecular mechanisms by which BA disrupts root development remain poorly understood. BA treatment significantly reduced stem and root length and inhibited lateral root emergence without affecting lateral root initiation. To investigate the underlying mechanism at cellular resolution, we established a non-sterile Agrobacterium rhizogenes-based root transformation system achieving 27.11
When ectopically expressed in hybrid poplar, the gymnosperm protein PdeLAZY1 confers LAZY1-like activity, whereas deletion of Domain V abolishes this activity without conferring TAC1-like function. The IGT gene family regulates plant architecture by controlling lateral organ angles. TAC1 is proposed to have evolved from an ancestral LAZY-like gene through loss of the conserved C-terminal Domain V, but this has not been tested functionally. We expressed PdeLAZY1, a LAZY1 homolog from the gymnosperm Pinus densiflora, and its Domain V-deleted variant (PdeLAZY1ΔV) under the 35S promoter in wild-type hybrid poplar (clone BH) and in a CRISPR-generated tac1 mutant. In wild-type poplar, PdeLAZY1 significantly reduced petiole angle, showing that a gymnosperm LAZY1 is competent to promote upright growth in an angiosperm context, whereas PdeLAZY1ΔV significantly increased it, indicating that Domain V is required for this activity. In the tac1 background, however, neither construct significantly increased petiole angle relative to the untransformed control, so PdeLAZY1ΔV did not complement loss of TAC1. Domain V is therefore required for the LAZY1-type activity of PdeLAZY1, but its loss alone is not sufficient to confer TAC1-like function in poplar. Because these are ectopic overexpression experiments, they test competence rather than endogenous function; within this limit, the data are consistent with a stepwise model in which changes beyond Domain V loss were required for TAC1 evolution.
Triacylglycerols (TAGs), once considered passive storage reserves confined to seeds, are now recognized as dynamic components of plant lipid metabolism with pivotal roles in stress adaptation. Far from being inert carbon depots, TAGs function as metabolic buffers that integrate energy storage, membrane lipid remodeling, and cellular protection under fluctuating environmental conditions. Abiotic stresses such as temperature extremes, drought, salinity, and nutrient limitation disrupt membrane integrity and lipid homeostasis, leading to the release of free fatty acids and the accumulation of reactive oxygen species (ROS). TAG biosynthesis under these conditions acts as a protective sink, sequestering toxic lipid intermediates, preserving membrane stability, and maintaining redox balance. The dynamic turnover between membrane lipids and TAGs allows plants to rapidly reorganize their lipid composition, stabilizing membranes during cold or heat stress, buffering osmotic imbalance during drought and salinity, and mitigating photodamage under nutrient deprivation. Emerging evidence highlights that TAG accumulation is not merely a byproduct of stress but a strategically regulated process linked to hormonal and redox signaling pathways. This reprogramming of lipid metabolism supports energy redistribution, detoxification of reactive lipid species, and recovery following stress exposure. This review summarizes recent advances in understanding the biosynthetic, regulatory, and physiological roles of TAGs during abiotic stress. It discusses how TAG metabolism operates at the intersection of lipid remodeling, cellular homeostasis, and stress signaling, positioning TAGs as key determinants of plant resilience and promising targets for the biotechnological enhancement of crop stress tolerance.
The NAC transcription factor AtANAC070 functions in zinc tolerance by directly activating AtMTP1 transcription to promote vacuolar zinc sequestration and homeostasis in A. thaliana. Zinc (Zn) is an essential micronutrient for plant growth, but it becomes toxic when present in excess. An initial screen of Arabidopsis thaliana T-DNA insertion mutants suggested a positive role of AtANAC070 in tolerance to excess Zn. AtANAC070 expression was induced under excess Zn, and loss of function of AtANAC070 led to increased Zn sensitivity and higher Zn accumulation. Conversely, AtANAC070 overexpression enhanced Zn tolerance and reduced Zn accumulation. Yeast one-hybrid assays identified Metal Tolerance Protein 1 (AtMTP1), which encodes a key transporter mediating vacuolar sequestration of excess Zn, as a downstream target of AtANAC070. Dual-luciferase reporter and real-time quantitative PCR (RT-qPCR) assays confirmed that AtANAC070 directly binds to the AtMTP1 promoter to activate its expression. The atmtp1 mutant accumulated more Zn than the WT and was more sensitive to excess Zn, whereas AtMTP1-overexpressing lines showed the opposite phenotype. The atanac070 atmtp1 double mutant displayed Zn sensitivity comparable to that of atmtp1 mutant, while AtMTP1 overexpression in the atanac070 background reduced Zn accumulation and restored Zn tolerance. These results indicate that AtANAC070 contributes to Zn homeostasis under excess Zn by promoting AtMTP1 expression.
A total of 164 APETALA2/ethylene-responsive factor (AP2/ERF) genes were identified in Hydrangea macrophylla, and HmaERF87 positively contributes to leaf spot resistance. The APETALA2/ethylene-responsive factor (AP2/ERF) transcription factor family plays important roles in plant stress responses, but its contribution to disease resistance in Hydrangea macrophylla (hydrangea) remains poorly understood. In this study, 164 AP2/ERF genes were identified in the H. macrophylla genome and classified into APETALA2 (AP2), ethylene-responsive factor (ERF), dehydration-responsive element-binding (DREB), and related to ABI3/VP1 (RAV) subfamilies. Their chromosomal distribution, conserved motifs, gene structures, and duplication patterns were analyzed. A total of 46 pathogen-responsive H. macrophylla AP2/ERF (HmaERF) genes were identified from the RNA sequencing (RNA-seq) dataset of resistant and susceptible cultivar leaves collected before and after Corynespora cassiicola inoculation. Promoter analysis revealed that the HmaERF genes with upregulated expression post-C. cassiicola infection showed a higher frequency and copy number of jasmonate-responsive cis-regulatory elements, suggesting their possible involvement in hormone-mediated defense responses. Three infection-induced candidate genes, including HmaERF56, HmaERF87, and HmaERF129, were selected for functional validation using virus-induced gene silencing (VIGS) in hydrangea leaf discs. Silencing of HmaERF87 expression via VIGS significantly increased lesion development after C. cassiicola inoculation, whereas the transient overexpression of HmaERF87 reduced the area of leaf disc lesions. Subcellular localization showed that the HmaERF87 protein was localized in the nucleus, and yeast assays indicated that its transcriptional activation activity was mainly associated with the C-terminal region of the protein. These results support a role for HmaERF87 as a positive regulator of H. macrophylla resistance to leaf spot disease and provide a candidate gene for further studies of disease resistance in hydrangea.
Quantitative mass spectrometry of Arabidopsis telomerase uncovered AtLa1, a homolog of ciliate and yeast proteins that promotes telomerase maturation. AtLa1 is essential for telomerase function in vivo, and in vitro it engages the same region of AtTR bound by AtNAP57, homologous to a telomerase accessory from mammals. Striking divergence is evident in the biogenesis pathways and protein complements of telomerase from mammals and single-cell organisms. However, little is known about plant telomerase. Along with catalytic subunit TERT and templating RNA TR, we previously showed Arabidopsis thaliana telomerase is also associated with AtNAP57, a dyskerin homolog essential for mammalian telomerase biogenesis. Here we employ quantitative mass spectrometry (qMS) to uncover additional Arabidopsis telomerase constituents. We report AtLa1 as a new telomerase-associated protein. RNA-IP assays confirmed AtLa1 association with AtTR, while transient RNAi-mediated knockdown of AtLa1 strongly diminished telomerase activity, supporting a functional role for AtLa1 in telomere maintenance in vivo. In vitro binding studies revealed AtLa1 contacts AtTR via the UUU-3′OH and a plant-specific P1a-P1b-P4 three-way junction (TWJ). Notably, the TWJ is also required for AtNAP57 association with AtTR. However, this protein was not detected in our qMS experiment using overexpressed AtTERT and AtTR, perhaps because the purification scheme enriched for RNP assembly intermediates. La-related proteins serve as RNA chaperones and are associated with a wide variety of telomerase complexes. Therefore, we postulate that AtNAP57 and AtLa1 compete for AtTR or bind sequentially during telomerase biogenesis. Further exploration of Arabidopsis telomerase may offer novel insights into telomerase evolution and mechanisms of biogenesis.
The overexpression of MdDIR7 can alleviate osmotic and oxidative damage, increase the expression of MdFRO2 and MdIRT1, and improve plant performance under Fe-deficient conditions. Iron (Fe) is an essential trace element that serves as a critical component in numerous cellular processes, such as photosynthesis, respiration, and nitrogen-sulfur assimilation. Fe deficiency commonly occurs in alkaline and calcareous soils worldwide and severely affects plant growth and development. Members of the Dirigent (DIR) family have been identified and shown to play a role in abiotic stress response in several species, but have not been identified in apple and whether they also play an important role in Fe-deficiency stress has not been revealed. In this study, 41 members of the DIR family were identified from the apple genome, and they were unevenly distributed across 12 chromosomes of the apple (not distributed on chromosome 8). Real-time fluorescence quantitative PCR (RT-qPCR) indicated that the expression levels of the MdDIR7 gene in the leaves and roots of Malus hupehensis were significantly higher than those of the other members of this family under Fe-deficiency stress. Therefore, MdDIR7 was selected for further functional analysis using stable transformation, root transformation, and virus-induced gene silencing (VIGS). The results showed that root overexpression of MdDIR7 significantly increased photosynthetic pigment contents and improved chlorophyll fluorescence parameters, reduced relative electrical conductivity (REC) and malondialdehyde (MDA) content, increased antioxidant enzyme activity (SOD, CAT), reduced the accumulation of reactive oxygen species (ROS), and was associated with Fe-deficiency stress. Similar results were also observed in apple calli. On the contrary, we found that silenced MdDIR7 (TRV-MdDIR7) under the stress led to greater damage than the TRV plants. In addition, root overexpression of MdDIR7 upregulated the expression of Fe uptake-related genes MdFRO2 and MdIRT1 and significantly increased total Fe content, whereas silencing of MdDIR7 downregulated the expression of MdFRO2 and MdIRT1. In summary, the overexpression of MdDIR7 gene was associated with improved performance under Fe-deficiency conditions.
