Abstract Trichomes are multicellular, non-glandular structures that play important roles in plant development, and provide protection against harmful ultraviolet radiation, insect damage and excessive water loss through transpiration. However, the regulation of trichome formation in cucumber remains incompletely understood. Although the phenotype of gl2 have been identified, no fine mapping and candidate gene for it has been determined so far. In this study, CsGL2 was mapped to a 72.5-kb region on chromosome 2, and via fine-mapping, an ethylene-responsive transcription factor ESR1, was identified as the candidate gene. We generated a CRISPR/Cas9 knockout mutant CsGL2 CRwhich was phenotypically resembled the natural gl2 mutant, demonstrating that CsESR1 is CsGL2. We further investigated whether CsGL2 interacts with CsGL1 and CsGL3, two known regulators of trichome development. Our results showed that CsGL2 directly binds to the promoter of CsGL1 and regulates its expression. Moreover, CsGL1 directly interacts with the CsGL3 protein. Genetic analysis demonstrated that CsGL3 is epistatic to CsGL1 and CsGL2, respectively. In conclusion, this study identified CsGL2 as a novel regulator of trichome development in cucumber, which is the first report of an ethylene-responsive transcription factor involved in cucumber trichome development. Furthermore, we elucidate the functional relationships among CsGL2, CsGL1 and CsGL3, providing new insights into the molecular network governing trichome development in cucumber.
Abstract Sweet cherry (Prunus avium) fruit is highly susceptible to postharvest softening, resulting in significant economic losses. This process is driven by the degradation of cell wall components, yet its transcriptional regulation remains unclear. Here, we explored the differential degradation of fruit cell walls in two sweet cherry cultivars with varying levels of fruit firmness. We identified the APETALA2/Ethylene Response Factor (AP2/ERF) family gene PavDREB2A, which is uniquely expressed during the ripening of the low-firmness cultivar. Phenotypic and transcriptomic analyses confirm its role in reducing cell wall integrity and fruit firmness. Through integrated transcriptomics, DAP-seq, and AlphaFold3 screening, we identified four cell wall-modifying genes (PG2, MAN7, EXP1, EXP4) as potential target genes of PavDREB2A. PavDREB2A directly activates these genes by binding to the DRE cis-element in their promoter regions, as shown by dual-luciferase, yeast one-hybrid, and EMSA assays. AlphaFold3 structural predictions indicate that six arginines in the AP2/ERF domain of PavDREB2A are essential for its binding and transcriptional activity. Our findings establish a PavDREB2A-mediated transcriptional pathway that promotes cell wall disassembly during ripening. This study provides both mechanistic insight into fruit texture control and potential molecular targets for breeding sweet cherry cultivars with enhanced firmness and shelf-life.
Abstract The tea plant (Camellia sinensis) possesses an array of specialized metabolites that contribute to its distinctive flavor and health-promoting properties. How they are regulated by long non-coding RNAs (lncRNAs) is not well elucidated. Here we systematically identify 65 675 high-confidence lncRNAs using the RNA sequencing dataset from the second-leaf samples of 133 accessions. Integrative co-expression analysis revealed 55 lncRNA-centered regulatory modules significantly associated with the accumulation of 57 specialized metabolites. We further show that a Gypsy-retrotransposon-derived lncRNA, CsLNC703, promotes CsMYB111-mediated regulation of the F3′H-associated flavan-3-ol biosynthetic branch, resulting in preferential accumulation of catechin (C), epicatechin (EC), and epicatechin gallate (ECG). These findings demonstrate that lncRNAs constitute an important regulatory layer controlling specialized metabolism in tea plants and reveal a transposon-derived lncRNA that modulates catechin biosynthesis. Our study provides new insights into the regulatory mechanisms underlying metabolic diversity in tea and highlights lncRNAs as potential targets for metabolic engineering and breeding of tea cultivars with improved quality traits.
Abstract Bacterial stem and root rot (BSRR), triggered by Dickeya dadantii, severely reduces sweetpotato productivity, yet the cellular mechanisms underlying root responses remain poorly understood. Here, we generated a single-cell transcriptomic landscape of 40 345 cells from infected and control sweetpotato root tips to uncover cell-type-specific defense patterns. Using canonical marker genes, we delineated seven major cell types and 16 transcriptionally distinct clusters. Cortex cells were identified as the principal responders to infection, characterized by extensive transcriptional reprogramming and enriched defense-related trajectories. Among the differentially expressed genes, IbWRKY40 emerged as a core regulator within the co-expression network of root immune responses. IbWRKY40 predominantly localized to the nucleus and exhibited strong induction approximately 9 h after pathogen exposure. Functional validation in sweet potato further suggested that IbWRKY40 negatively modulates resistance to D. dadantii. To our knowledge, this is the first report elucidating the cellular-level molecular mechanisms of sweet potato root tips in response to BSRR infection, providing valuable insights into plant defense strategies under pathogen stress.
Abstract Structural variations (SVs) represent an important source of genomic diversity and can contribute substantially to phenotypic variation in crops. However, the population-scale distribution and phenotypic effects of SVs in Theobroma cacao L. (cocoa) remain poorly understood. Here, we constructed a population-scale cocoa SV atlas using whole-genome resequencing data from 165 cocoa accessions representing ten previously defined genetic groups. Using a unified short-read-based SV detection strategy, we identified 11 271 high-confidence SVs, including deletions, duplications, and inversions. Using a framework based on term frequency-inverse document frequency (TF-IDF) algorithm, we identified 1078 fingerprint SVs for ten cocoa genetic groups of which 145 showed significant SV–trait associations. To further investigate integrated phenotypic divergence associated with functional SVs, we developed an analysis framework based on latent Dirichlet allocation (LDA). This analysis identified four latent phenotypic features showing significant divergence among cocoa genetic groups. Geographic populations from South America displayed extensive admixture of genetic groups, whereas populations outside South America showed reduced genetic diversity consistent with historical dispersal bottlenecks. Our study provides a population-scale SV resource for cocoa and demonstrates that SVs contribute to genetic differentiation, phenotypic divergence, and geographic adaptation in cocoa populations.
Abstract Among the traits that define the appeal of rose flowers, fragrance, which results from complex blends of volatile organic compounds (VOCs) holds a central cultural, sensory and economic role. These VOCs mainly include terpenoids, phenylpropanoid/benzenoid derivatives, phenolic methyl ethers and norisoprenoids, with strong variation among species and cultivars. In this review, we provide a broad, near-comprehensive and chronological synthesis of research on biosynthesis, regulation and emission of rose VOCs, with particular attention to the experimental approaches that have shaped the field, from classical biochemical and molecular studies to recent advances enabled by metabolic analyses, genomic resources and functional genomics. These developments have recently enabled to decipher major pathways leading to rose VOCs and highlighted the contribution of gene duplication, enzyme diversification and natural variation to scent diversity. Recent findings have been especially important for terpenoids. These includes the discovery of original cytosolic terpene biosynthetic pathways involving both terpene synthase-dependent and terpene synthase-independent routes, the cytosolic origin of the geraniol precursor GPP, the enzymatic conversion of geraniol to β-citronellol and the characterization of numerous terpene synthases involved in the production of major terpenes produced by rose flowers. We also discuss advances in the regulation of VOC biosynthesis, together with emerging questions regarding tissue specialization, intracellular trafficking, transport and emission of scent compounds. By integrating historical discoveries with recent findings, this review outlines how rose scent biology has been reshaped and discuss remaining challenges for understanding, breeding and engineering desirable floral fragrances.
Abstract Drought stress severely restricts the growth and yield of alfalfa (Medicago sativa L.), and ABA plays a vital role in plant drought tolerance. Here, we report that a transcription factor MsSPL12, induced by both drought and ABA treatment, mediates drought tolerance in alfalfa. Overexpression of MsSPL12 enhanced drought resistance and ABA sensitivity in alfalfa plants, while suppression of its expression led to the opposite phenotypes. Population genetic analysis across Medicago species revealed a SNP in the 5’UTR of MsSPL12 that converts the strong repressive Type I upstream open reading frame (uORF) into a weaker Type II. Disruption of the Type I uORF12 by using the CRISPR/Cas9 system markedly elevated the translation efficiency of MsSPL12, leading to enhanced ABA sensitivity and improved drought resistance in two alfalfa cultivars. Transcriptome analysis showed MsSPL12 modulates numerous drought-responsive genes, with ABA biosynthesis genes closely associated with altered ABA levels in transgenic plants. Biochemical assays verified that MsSPL12 directly activates the expression of MsBCH2 and MsCCD4, key genes in ABA biosynthesis, thereby promoting ABA accumulation. Overexpression of MsBCH2 and MsCCD4 significantly increased ABA content and drought tolerance in alfalfa plants and hairy roots, respectively. This study reveals the regulatory module of MsSPL12-mediated ABA biosynthesis in alfalfa drought tolerance, providing valuable molecular targets and uORF-based strategies for drought-tolerant alfalfa breeding.
Abstract Abscission is a fundamental developmental process in plants that can also be integrated into a defense strategy to restrict pathogen spread by shedding colonized organs. Members of Botryosphaeriaceae represent a major threat to global agriculture. These hemibiotrophic fungal pathogens colonize the vascular tissues of woody plants as endophytes, eliciting no visual symptoms. Under stress conditions, they switch to a necrotrophic lifestyle, causing destructive diseases such as stem dieback and fruit stem-end rot. Despite their economic significance, the role of the earliest, endophytic chronic colonization in triggering premature fruit drop, or abscission, remains largely unexplored. In this study, we uncover a previously overlooked phenomenon where Lasiodiplodia theobromae endophytically colonizes mango (Mangifera indica) flowers and fruitlets at the abscission zone (AZ), triggering premature abscission through activation of host ethylene-dependent defense responses. This causes the colonized fruitlets to drop prematurely, reducing the overall fruit yield. The presence of fungi initiates a cascade of events at the AZ, reprogramming the AZ cells through oxidative bursts and ethylene biosynthesis. Thereafter, the fungal secretion of ammonia leads to local cytoplasmic alkalization, ultimately accelerating callose deposition, accompanied by a depletion of auxin and cytokinin fluxes, and drives premature fruit detachment. The results elucidate two previously uncharacterized facets of the Botryosphaeriaceae endophytic/chronic colonization stage. We demonstrate that host damage and yield loss initiate before the necrotrophic transition and characterize a novel systemic defense response wherein the tree prematurely drops colonized fruits to limit pathogenic fungi colonization.
Abstract The basic leucine zipper (bZIP) transcription factor family plays crucial roles in plant development and stress responses, yet its evolutionary dynamics and functional diversification across green plants remain poorly understood. Here, we conducted a comprehensive analysis of the bZIP family across 114 green plant taxa, from algae to angiosperms, with emphasis on Rosaceae species and Malus accessions. Phylogenetic analysis showed that most bZIP subfamilies formed stable monophyletic clades across green plants, except for two atypical evolutionary patterns. An intertwined E-M-E-I complex suggested that the core domain of subfamily M originated from the group E lineage, whereas the highly divergent subclades, S1 and I1, highlighted the uneven evolutionary rates within the family. In Rosaceae, lineage-specific whole genome duplication (WGD) events, especially in Malinae subtribe and Potentilla, markedly expanded bZIP repertoires with asymmetric retention among subfamilies. Furthermore, lineage-specific pangenome in Malus identified 1251 core and 18 unique bZIP members across diverse accessions. Pan-transcriptome identified tissue-specific expression and stress-responsive co-expression modules. And functional characterization of MP_bZIP77, a gene derived from wild apple, provides a concrete example of this regulatory mechanism by demonstrating that its overexpression enhances salt tolerance through the activation of antioxidant enzyme activities. This study provides novel insights into the evolutionary origin, expansion pattern, and functional divergence of the bZIP family in green plants and Rosaceae, while laying a theoretical and genetic foundation for future molecular breeding aimed at improving stress resistance in fruit trees.
Abstract Grafting cucumber onto salt-tolerant pumpkin rootstocks is widely used to alleviate salt stress, yet the contribution of long-distance mRNA signaling to rootstock-conferred salt tolerance remains unclear. In this study, using reciprocal cucumber/pumpkin heterografts with SNP-resolved tracking, we demonstrate that salt stress rewires long-distance mRNA trafficking. Salinity imposes a pronounced directional bias, favouring mRNA export from the tolerant pumpkin to the sensitive cucumber while limiting reverse flux, irrespective of graft configuration. This identifies directional RNA mobility as an emergent property of salt stress adaptation. Among pumpkin-derived mobile transcripts, CmoHSP83 and CmoCAT2 were strongly enriched in cucumber scions and selected for functional validation. Their long-distance mobility was confirmed using transgenic hairy-root systems, species-specific RT–PCR, SNP-informed Hi-TOM sequencing, and GUS assays. Rootstock overexpression of either gene enhanced salt tolerance in grafted cucumber, accompanied by reduced ROS accumulation and improved Na +/K+ homeostasis. Collectively, these findings establish mobile mRNAs as functional long-distance signals in rootstock–scion communication under salinity and provide mechanistic insights into pumpkin rootstock-conferred systemic salt adaptation in grafted cucumber.
Abstract Ethylene is the key hormone controlling climacteric fruit ripening, a process that plays a crucial role in determining both fruit quality and shelf-life. Melon serves as an excellent model for studying ripening due to the presence of both climacteric and non-climacteric varieties. In this study, we fine-tuned ethylene production by combining the alleles of three non-climacteric QTLs, ETHQB3.5, ETHQV6.3, and ETHQV8.1, into a climacteric Cantaloupe background. This pyramiding strategy resulted in lines with reduced and delayed ethylene production, which in turn affected fruit flesh firmness and harvest timing. Regarding aroma, although overall volatile organic compounds (VOCs) production was reduced, the fruits retained the synthesis of specific ester-based compounds, preserving some of the characteristic sweet aroma of Cantaloupe. QTL pyramiding also revealed the impact of QTL-QTL interactions on ripening-related traits: ETHQV6.3 and ETHQV8.1 acted synergistically, while ETHQB3.5 showed an antagonistic effect on ETHQV6.3. This approach generated a range of climacteric behaviours, offering valuable genetic resources for breeding programs aimed at improving long-shelf life.
Abstract The adzuki bean (Vigna angularis) is an economically and nutritionally significant legume species. Using a multi-strategy sequencing approach, a gapless, telomere-to-telomere (T2T) genome assembly of an elite cultivar, Jihong 16 (JH16), was generated. The genome size was 514.85 Mb, with a Contig N50 of 48.72 Mb, 3x longer than previous published genome. The centromeres of adzuki bean were revealed for the first time and found to be composed of ~ 9-bp minisatellites — a distinct departure from the much longer satellites typically found in other plants. Low sequence conservation between the centromeres of adzuki bean and mung bean further highlights the highly active evolutionary dynamics of centromeric sequences in Vigna species. Whole-genome resequencing of 706 adzuki bean accessions was performed and three major phylogenetic groups were identified. Among these, the Southern China group exhibited the highest genetic diversity and the most rapid linkage disequilibrium (LD) decay. A set of 112 core accessions was selected to represent the global germplasm, and 12 barcode SNPs were identified for DNA fingerprinting. Leveraging high-quality phenotypic data for 39 agronomic traits collected across seven locations over two years, a genome-wide association study (GWAS) identified 9613 significant marker-trait associations (MTAs) and thousands of candidate genes, including 40 MTAs located in the centromere regions. Several genes were experimentally validated, including the flowering-time gene Vigan07G000670, overexpression of which in Arabidopsis led to early flowering phenotype and altered FT/FLC expression. Finally, genomic selection (GS) evaluations demonstrated that even a small set (500 ~ 1000) of trait-associated markers achieved high prediction accuracy (>0.8) for key agronomy traits such as seed weight, flowering date and plant height. The genomic resources and insights generated in this study provide a robust foundation for genetic improvement of adzuki bean and related legume crops.
Abstract Gray mold caused by Botrytis cinerea poses severe threats to global horticultural production. Conventional chemical fungicides face mounting restrictions due to rising pathogen resistance and stringent food safety regulations, creating an urgent need for sustainable alternatives. RNA pesticides offer an eco-friendly alternative to chemical fungicides. However, the application of RNA pesticides faces challenges including double-stranded RNA (dsRNA) instability, delivery inefficiency, and high production costs. Here, we utilized an endophyte to produce dsRNA, enabling effective control of gray mold. The Beauveria bassiana strain QSE-F1 was proved as tomato endophyte and engineered to express dsRNA targeting BcMucin, an essential gene in B. cinerea and was validated as an effective RNA interference (RNAi) target. The engineered B. bassiana (Bb-dsMucin) produced and delivered dsRNA to achieve significant silencing of BcMucin at transcriptional and translational levels. The Bb-dsMucin strain showed superior antifungal activity and gray mold control efficacy compared with control strains. The combined use of conidia and fermentative broth of Bb-dsMucin attained optimal and lasting control efficacy. Our research establishes phyllosphere-derived Microbe-induced gene silencing (MIGS) as a promising biocontrol strategy for managing gray mold, and promotes the advancement of RNA pesticides technologies.
Abstract Anthocyanins are important plant pigments that contribute to leaf coloration and stress adaptation, but how their biosynthesis is regulated across different leaf cell types remains unclear, especially in woody plants. Here, we combined single-nucleus RNA sequencing (snRNA-seq) and bulk RNA-seq to investigate anthocyanin-associated regulatory programs in Pistacia chinensis leaves. We found that epidermal cells followed a largely unidirectional transition toward high-anthocyanin states, whereas mesophyll cells differentiated into multiple functional subtypes, revealing substantial cellular heterogeneity. Although epidermal and mesophyll cells shared the core anthocyanin biosynthetic pathway, their upstream regulatory programs differed. Integrative analysis identified multiple transcription factors associated with anthocyanin accumulation, several of which showed distinct expression patterns between high-anthocyanin epidermal cells and anthocyanin-enriched mesophyll cells. Functional assays showed that PcMYB113a and PcMYB113b promote anthocyanin accumulation and interact with conserved MBW-complex components. Overall, this study provides cell-resolved insight into anthocyanin-associated regulatory programs in woody plant leaves.
Abstract High temperature is a major environmental constraint limiting the growth, development, ornamental quality, and reproductive performance of chrysanthemum (Chrysanthemum morifolium). In production systems, thermal stress often coincides with drought-like hydraulic stress, including increased vapor pressure deficit and soil water deficit, and may promote salinity or ionic stress under specific substrate, fertigation, or protected-cultivation conditions. These associated constraints may intensify high-temperature-induced developmental disorders, but their relative importance depends on cultivar, developmental stage, and cultivation context. In this review, we synthesize current knowledge of high-temperature responses in chrysanthemum, with an emphasis on developmental vulnerability and species-specific ornamental traits. We first summarize the effects of elevated temperature on vegetative growth, adventitious rooting, photosynthesis, plant architecture, flowering time, capitulum morphogenesis, floral pigmentation, postharvest performance, pollen fertility, and seed set. We then discuss how heat-responsive pathways intersect with water-status regulation, ion homeostasis, hormone signaling, reactive oxygen species metabolism, heat shock responses, transcriptional regulation, and epigenetic modulation. Particular attention is given to distinguishing direct evidence from chrysanthemum from mechanistic insights inferred from model plants or other crops. Finally, we outline adaptive strategies, including environmental control, chemical priming, grafting, germplasm evaluation, molecular breeding, and AI-assisted cultivation. This review provides a high-temperature-centered framework for understanding stress-associated developmental disorders in chrysanthemum and for guiding the future improvement of climate-resilient ornamental cultivars.
Abstract Pollen tubes are among the fastest growing polarized plant cells and must traverse the stigma, style and ovary to deliver male gametes. The style constitutes the longest and most complex phase of this journey yet the molecular and biomechanical logic of pollen tube behavior within this tissue remains poorly integrated. Here we synthesize recent genetic, live-imaging and biochemical studies to assemble a coherent sequence from ionic gradients, actin dynamics and vesicle trafficking to cell wall remodeling, and propose an integrative signal-mechanics feedback framework for long-distance guidance, arrest and reactivation. Within this framework, arrest at the stylar base is interpreted as an actively regulated checkpoint rather than passive waiting. We further place this view in a broader evolutionary context by comparing prolonged progamic phase in gymnosperms with delayed fertilization in Fagaceae, where ovule immaturity is coupled to intermittent pollen tube arrest and reactivation within the pistil. From this synthesis we derive three testable predictions: localized remodeling of the stylar base extracellular matrix may modulate pollen tube reactivation by altering mechanical resistance and engaging FER/LRX-related mechanosensory Ca2+ signaling; AGPs may support the ionic environment required for exocytosis through local Ca2+ supply; and female tissues actively maintain or restart tube quiescence in delayed-fertilization systems. Finally, we highlight several unresolved questions and methodological bottlenecks that point toward future directions. This conceptual synthesis bridges insights from model and non-model species, provides a predictive perspective on long-distance tip growth and its evolutionary diversification, and outlines a forward-looking agenda for plant reproductive biology.
Abstract Bananas (Musa spp.) are a globally important fruit and cash crop. Their postharvest ripening process directly determines fruit eating quality, commercial value, and shelf life. As a typical climacteric fruit, banana ripening is governed by the ethylene signalling pathway and involves multi-layer regulation including transcriptional reprogramming, post-translational modifications, and epigenetic regulation. In recent years, with the completion of multiple banana genome maps and the integration of multi-omics technologies, the molecular mechanisms underlying fruit quality formation have been systematically uncovered. This review summarises research progress in the past decade, focusing on the genetic contributions of different subgenomes (A, B, S, T), the regulatory networks controlling key ripening processes, and multi-layer regulatory mechanisms. In addition, we discuss the molecular basis of banana fruit responses to high- and low-temperature stresses, as well as the effects of postharvest technologies on ripening progression and quality. This review deliberately focuses on multi-layer regulatory network architecture, aiming to elucidate how these regulatory layers are hierarchically organised and functionally integrated to coordinately govern banana fruit ripening and quality formation, providing a theoretical basis for postharvest biology research, quality improvement, and preservation technology development, and offering potential targets for molecular breeding and precision postharvest management.
Abstract Grapevine (Vitis vinifera), an industry valued at approximately 108.61 billion US dollars globally, faces escalating threats from abiotic stresses that intensify under climate change and increasingly compromise berry quality, phenology, and yield. Despite decades of molecular characterization, translating stress biology knowledge into climate-resilient cultivars remains limited. We argue this gap reflects not a lack of knowledge within individual biological layers, but a fundamental failure to integrate across them. Grapevine stress tolerance operates through three interconnected regulatory layers. Transcription factor networks, WRKY, NAC, MYB, DREB, and bZIP families, constitute the most rapid layer, converging on shared ABA-mediated signaling hubs despite apparent stress-type specificity. Quantitative trait loci and genome-wide association studies capture the genomic architecture underlying these responses, yet remain critically under characterized for drought, salinity, and heavy metal tolerance. Epigenetic regulation through DNA methylation, histone modifications, and stress memory mechanisms constitutes a temporally durable third layer, uniquely important for perennial crops where adaptive chromatin states persist across growing seasons. Critically, these layers are not independent: transcription factor activity shapes the chromatin landscape, epigenetic marks modulate QTL expression, and genomic loci encode the regulatory machinery executing stress responses. Current breeding tools, marker-assisted selection, CRISPR/Cas9, and epigenomic selection, map onto these three layers but are overwhelmingly applied in isolation, limiting their collective impact. This review synthesizes knowledge gaps across all three layers within a unified hierarchical framework, arguing that deliberate cross-layer integration through multi-omics and precision breeding could enable cultivars capable of sustaining productivity under a rapidly changing climate.
Abstract Leaf color is an important trait affecting vegetable quality, yield, and market value. However, traditional methods for leaf color assessment are often subjective or destructive, which limits accurate and high-throughput phenotyping. In this study, an unmanned aerial vehicle (UAV)–based multispectral imaging platform was used to collect phenotypic data from 214 Chinese cabbage inbred lines at the rosette stage. A multispectral UNet model was applied to segment individual plants, and a membership function was used to quantify leaf color on a continuous scale. Based on these high-throughput phenotypic data, a genome-wide association study was used to identify two candidate genes, BrEMB976 and BrGSH2, on chromosome A06. Subsequent virus-induced gene silencing analysis showed that silencing these genes altered leaf color. In addition, a deep learning-based genomic selection model, BrDeepGS, was developed for leaf color prediction, which achieved a Pearson correlation coefficient of 0.853. These results demonstrate the potential of integrating UAV-based high-throughput phenotyping, candidate gene analysis, and genomic prediction for leaf color evaluation and selection in Chinese cabbage breeding.
