Apples are one of the most valued tree fruit crops around the world. Currently, a few highly popular and economically successful apple cultivars dominate the commercial production and serve as main genetic contributors to the development of new apple cultivars. This limited level of genetic diversity grown as a clonally propagated monoculture renders the apple industry vulnerable to the wide range of weather events, pests, and pathogens. Wild apple species are an excellent source of beneficial alleles for the wide range of biotic and abiotic stressors challenging apple production. However, the biological barriers of breeding with small-fruited wild apples greatly limit their use. Using a closely related wild species of apple such as Malus sieversii can improve the efficiency of breeding efforts and broaden the base of available genetics. M. sieversii is the main progenitor of the domesticated apple, native to Central Asia. The similarity of fruit morphology to domesticated apples and resistances to abiotic and biotic stresses makes it appealing for apple breeding programs. However, this important species is under threat of extinction in its native range. Preserving the wild apple forests in Central Asia is vital for ensuring the sustainable protection of this important genetic resource. The insufficient awareness about the complete range of challenges and opportunities associated with M. sieversii hinders the maximization of its potential benefits. This review aims to provide comprehensive information on the cultural and historical context of M. sieversii, current genetic knowledge for breeding, and the conservation challenges of wild apple forests.

Triterpene (C30 isoprene compounds) represents the most structurally diverse class of natural products and has been extensively exploited in the food, medicine, and industrial sectors. Decades of research on medicinal triterpene biosynthetic pathways have revealed their roles in stress tolerance and shaping microbiota. However, the biological function and mechanism of triterpenes are not fully identified. Even this scientific window narrows down for horticultural trees. The lack of knowledge and a scalable production system limits the discovery of triterpene pathways. Recent synthetic biology research revealed several important biosynthetic pathways that define their roles and address many societal sustainability challenges. Here, I review the chemical diversity and biosynthetic enzymes involved in triterpene biosynthesis of horticultural trees. This review also outlines the integrated Design-Build-Test-Learn (DBTL) pipelines for the discovery, characterization, and optimization of triterpene biosynthetic pathways. Further, these DBTL components share many fundamental and technical difficulties, highlighting opportunities for interdisciplinary collaboration between researchers worldwide. This advancement opens up unprecedented opportunities for the bioengineering of triterpene compounds toward development and scaleup processes.

As one of the grave environmental hazards, soil salinization seriously limits crop productivity, growth, and development. When plants are exposed to salt stress, they suffer a sequence of damage mainly caused by osmotic stress, ion toxicity, and subsequently oxidative stress. As sessile organisms, plants have developed many physiological and biochemical strategies to mitigate the impact of salt stress. These strategies include altering root development direction, shortening the life cycle, accelerating dormancy, closing stomata to reduce transpiration, and decreasing biomass. Apart from being a prime energy source, light is an environmental signal that profoundly influences plant growth and development and also participates in plants' response to salt stress. This review summarizes the regulatory network of salt tolerance by light signals in plants, which is vital to further understanding plants' adaptation to high salinity. In addition, the review highlights potential future uses of genetic engineering and light supplement technology by light-emitting diode (LED) to improve crop growth in saline-alkali environments in order to make full use of the vast saline land.

Although plant secretory tissues play important roles in host defense against herbivores and pathogens and the attraction of insect pollinators, their genetic control remains elusive. Here, it is focused that current progress has been made in the genetic regulatory mechanisms underpinning secretory tissue development in land plants. C1HDZ transcription factors (TFs) are found to play crucial roles in the regulation of internal secretory tissues in liverworts and Citrus as well as external secretory tissues in peach. C1HDZ TFs regulate secretory tissue development via synergistic interaction with AP2/ERF and MYC TFs. Thus, a set of genes are speculated to be recruited convergently for the formation of secretory tissues in land plants.

Gene transcription is governed by a complex regulatory system involving changes in chromatin structure, the action of transcription factors, and the activation of cis-regulatory elements. Postharvest fruits are threatened by Penicillium expansum, a leading causal agent of blue mold disease and one of the most economically significant postharvest pathogens worldwide. However, information on its transcription regulatory mechanism is lagging. Here, we conducted an assay for transposase accessible chromatin sequencing (ATAC-seq) for P. expansum during vegetative growth and infection phase and then studied the function of a basic leucine zipper (bZIP) transcription factor PeAtf1. Results highlighted the role of promoter regions in gene transcription and the significant difference in P. expansum between these two phases. Six footprint-supported cis-regulatory elements of active transcription factors were obtained and analyzed. We then identified a homolog of the bZIP regulator Atf1, PeAtf1, and found it positively regulated vegetative growth, reproduction, and osmotic stress response in P. expansum. Furthermore, PeAtf1 deletion enhanced the fungus's tolerance to oxidative, cell wall, and membrane stresses, which might contribute to the virulence of deletion mutants in apple fruits, leading to similar pathogenicity between mutants and the wild type. Overall, this study provides new insights into the transcription regulatory profile of P. expansum, aiding in the future development of strategies to control P. expansum.

Flowering date in perennial fruit trees is an important trait for fruit production. Depending on the winter and spring temperatures, flowering of olive may be advanced, delayed, or even suppressed. Deciphering the genetic control of flowering date is thus key to help selecting cultivars better adapted to the current climate context. Here, we investigated the genetic determinism of full flowering date stage in cultivated olive based on capture sequencing data of 318 genotypes from the worldwide olive germplasm bank of Marrakech, Morocco. The genetic structure of this collection was organized in three clusters that were broadly attributed to eastern, central, and western Mediterranean regions, based on the presumed origin of genotypes. Flowering dates, collected over 7 years, were used to estimate the genotypic best linear unbiased predictors, which were then analyzed in a genome-wide association study. Loci with small effects were significantly associated with the studied trait, by either a single- or a multi-locus approach. The three most robust loci were located on chromosomes 01 and 04, and on a scaffold, and explained 7.1%, 6.2%, and 6.5% of the trait variance, respectively. A significantly higher accuracy in the best linear unbiased predictors of flowering date prediction was reported with Ridge- compared to LASSO-based genomic prediction model. Along with genomic association results, this suggests a complex polygenic determinism of flowering date, as seen in many other fruit perennials. These results and the screening of associated regions for candidate genes open perspectives for further studies and breeding programs targeting flowering date.

Alfalfa is one of the most economically valuable forage crops in the world. However, molecular cytogenetic studies in alfalfa lag far behind other cash crops and have reached a bottleneck. Here, we developed a novel chromosome identification system by designing 21 oligo probes in specific regions of each chromosome, which can be used as a barcode to simultaneously distinguish all chromosomes in a cell. Using this system, we revealed the chromosome karyotype features and evolutionary differences among 10 cultivated alfalfa varieties. Interestingly, we also found two chromosomal variation types, i.e. aneuploidy and large chromosomal segment deletions in the seeds of three alfalfa varieties. Variation frequency analysis showed that only 7/173 seeds in those three alfalfa varieties had chromosome aberrations, which indicated that the inheritance and meiosis of alfalfa had evolved to a relatively stable state. Remarkably, 4/7 variation seeds were chromosome 2 aberrations, suggesting that chromosome 2 appears to be more susceptible to natural chromosomal aberrations than other chromosomes during inheritance. DNA sequence variation analysis showed that the difference of presence and absence variations (PAVs) among homologous copies of chromosome 2 was larger than that of the other seven chromosomes. We suggest that such large PAV divergence among homologous copies may provide the physical basis for natural chromosome 2 aberrations propensity. Our study provides a valuable and efficient tool for alfalfa’s molecular cytogenetics and sheds new insights into the propensity for natural chromosome aberrations during autopolyploid inheritance.

The circadian clock mediates metabolic functions of plants and rhythmically shapes structure and function of microbial communities in the rhizosphere. However, it is unclear how the circadian rhythm of plant hosts regulates changes in rhizosphere bacterial and fungal communities and nutrient cycles. In the present study, we measured diel changes in the rhizosphere of bacterial and fungal communities, and in nitrogen (N) and phosphorus (P) cycling in 20-year-old tea plantations. The fungal communities were more stable in their responses to circadian changes than bacterial communities in the rhizosphere of the cultivars LJ43 and ZC108. Nevertheless, fungal genera with circadian rhythms were more numerous and had a higher abundance at midnight. Organic P and N mineralization in the rhizosphere was more intensive in LJ43 under day-night alterations, while inorganic N and P cycling was more easily affected by circadian rhythms in ZC108. The rhizosphere denitrification encoded by the genes AOA and AOB was intensive in the morning, irrespective of tea cultivar. Genes related to rhizosphere N fixation (nifH) and denitrification (nosZ and nirK) expressed at greater levels in ZC108, and they reached a peak at midnight. Moreover, the diel rhythm of rhizosphere microbial communities in ZC108 largely regulated dial changes in N and P cycling. These results suggested that the bacterial and fungal communities in the rhizosphere respond differently to circadian rhythms, and they vary between tea cultivars. The timing of bacterial and fungal cycling largely regulates rhizosphere N and P cycling and their ecological functions.

Resveratrol is an important phytoalexin that adapts to and responds to stressful conditions and plays various roles in health and medical therapies. However, it is only found in a limited number of plant species in low concentrations, which hinders its development and utilization. Chalcone synthase (CHS) and stilbene synthase (STS) catalyze the same substrates to produce flavonoids and resveratrol, respectively. However, it remains unclear how CHS and STS compete in metabolite synthesis. In this study, two CHS2 mutant cell lines (MT1 and MT2) were generated using CRISPR/Cas9 genome editing. These CHS2 mutant cell lines exhibited abundant mutations in CHS2, leading to the premature termination of protein translation and subsequent CHS2 knockout. Amplicon sequencing confirmed comprehensive CHS2 knockout in MT1, whereas the wild-type sequence remained predominant in the MT2 cell line. Transcriptome and RT-qPCR results showed a significant downregulation of genes involved in flavonoid biosynthesis, including CHS2, CHS3, F3H, F3’H, DFR, FLS, LDOX, among others, resulting in decreased flavonoid accumulation, such as anthocyanins, proanthocyanidins, quercetin, and kaempferol. Conversely, STS genes involved in stilbenoid biosynthesis were upregulated competing with the flavonoid pathway. Consequently, there was a marked increase in stilbenoids, including resveratrol, piceatannol, piceid, and pterostilbene, with a 4.1-fold increase in resveratrol and a 5.3-fold increase in piceid (a derivative of resveratrol) observed in CHS2 mutant cell lines. This research demonstrates that CHS2 mutation induces a shift from flavonoid biosynthesis towards stilbenoid biosynthesis, offering new insights into metabolite biosynthesis and regulation, as well as an alternative solution for natural resveratrol production, and a novel breeding approach for eliminating non-target agronomic traits using CRISPR-Cas9.

Cherries are one of the economically important fruit crops in the Rosaceae family, Prunus genus. As the first fruits of the spring season in the northern hemisphere, their attractive appearance, intensely desirable tastes, high nutrients content, and consumer-friendly size captivate consumers worldwide. In the past 30 years, although cherry geneticists and breeders have greatly progressed in understanding the genetic and molecular basis underlying fruit quality, adaptation to climate change, and biotic and abiotic stress resistance, the utilization of cherry genomic data in genetics and molecular breeding has remained limited to date. Here, we thoroughly investigated recent discoveries in constructing genetic linkage maps, identifying quantitative trait loci (QTLs), genome-wide association studies (GWAS), and validating functional genes of edible cherries based on available de novo genomes and genome resequencing data of edible cherries. We further comprehensively demonstrated the genetic architecture of the main agronomic traits of edible cherries by methodically integrating QTLs, GWAS loci, and functional genes into the identical reference genome with improved annotations. These collective endeavors will offer new perspectives on the availability of sequence data and the construction of an interspecific pangenome of edible cherries, ultimately guiding cherry breeding strategies and genetic improvement programs, and facilitating the exploration of similar traits and breeding innovations across Prunus species.

The recessive genic male sterility (RGMS) method has several benefits in hybrid seed production; however, it is seldom employed in industrial hybrid seed production owing to the difficulty of producing an ample number of pure male-sterile seeds. In this study, we present an efficient methodology for developing a two-line strategy to produce hybrid seed through targeted mutation of BnaMS1 and BnaMS2 in conjunction with the RUBY reporter in Brassica napus. In this method, male-sterile lines were successfully created directly from different elite rapeseed breeding lines through CRISPR/Cas9-mediated mutagenesis and enhanced Agrobacterium-mediated transformation. To establish an efficient transgenic maintainer, three seed production technology (SPT) cassettes carrying a functional BnaMS1 gene linked to different reporters (DsRed, BnaA07.PAP2, and RUBY) were tested and compared in rapeseed. The results indicated that the PMR-based reporter possesses advantages such as phenotypic stability and ease of identification at early stages, making it an ideal tool for rapid and efficient screening. Subsequently, ideal transgenic maintainer lines with a single hemizygous copy of the SPT cassette were successfully developed in the context of Bnams1Bnams2 double mutants. The progeny from crossing the maintainer line with its male-sterile counterpart exhibited a 1:1 segregation pattern of nontransgenic male-sterile and male-fertile maintainer plants, distinguishable by seedling color. This biotechnological approach to male sterility offers promising prospects for improving the propagation of recessive genic male-sterile plants and the development of hybrid seeds in rapeseed. Furthermore, it is simple to adapt this technique to more Brassica crops.

Appropriate root system architecture (RSA) can improve alfalfa yield, yet its genetic basis remains largely unexplored. This study evaluated six RSA traits in 171 alfalfa genotypes grown under controlled greenhouse conditions. We also analyzed five yield-related traits in normal and drought stress environments and found a significant correlation (0.50) between root dry weight (RDW) and alfalfa dry weight under normal conditions (N_DW). A genome-wide association study (GWAS) was performed using 1 303 374 single-nucleotide polymorphisms (SNPs) to explore the relationships between RSA traits. Sixty significant SNPs (−log₁₀(P) ≥ 5) were identified, with genes within the 50 kb upstream and downstream ranges primarily enriched in GO terms related to root development, hormone synthesis, and signaling, as well as morphological development. Further analysis identified 19 high-confidence candidate genes, including AUXIN RESPONSE FACTORs (ARFs), LATERAL ORGAN BOUNDARIES-DOMAIN (LBD), and WUSCHEL-RELATED HOMEOBOX (WOX). We verified that the forage dry weight under both normal and drought conditions exhibited significant differences among materials with different numbers of favorable haplotypes. Alfalfa containing more favorable haplotypes exhibited higher forage yields, whereas favorable haplotypes were not subjected to human selection during alfalfa breeding. Genomic prediction (GP) utilized SNPs from GWAS and machine learning for each RSA trait, achieving prediction accuracies ranging from 0.70 for secondary root position (SRP) to 0.80 for root length (RL), indicating robust predictive capability across the assessed traits. These findings provide new insights into the genetic underpinnings of root development in alfalfa, potentially informing future breeding strategies aimed at improving yield.

Angiosperms are prolific producers of structurally diverse terpenes, which are essential for plant defense responses, as well as the formation of floral scents, fruit flavors, and medicinal constituents. Terpene synthase genes (TPSs) play crucial roles in the biosynthesis of terpenes. This study specifically focuses on the catalytic products of 222 functionally characterized TPSs in 24 angiosperms, which mainly comprise monoterpenes, sesquiterpenes, diterpenes, and sesterterpene. Our systematic analysis of these TPSs uncovered a significant expansion of the angiosperm-specific TPS-a, b, and g subfamilies in comparison to the TPS-e/f and c subfamilies. The expanded subfamilies can be further partitioned into distinct branches, within which considerable functional innovation and diversification have been observed. Numerous TPSs exhibit bifunctional or even trifunctional activities in vitro, yet they exhibit only a single activity in vivo, which may be largely determined by their inherent properties, subcellular localization, and the availabilities of endogenous substrates. Additionally, we explored the biological functions of terpenes in various organs and tissues of angiosperms. We propose that the expansion and functional divergence of TPSs contribute to the adaptability and diversity of angiosperms, facilitating the production of a broad spectrum of terpenes that enable diverse interactions with the environment and other organisms. Our findings provide a foundation for comprehending the correlation between the evolutionary features of TPSs and the diversity of terpenes in angiosperms, which is significant for terpene biosynthesis research.

Ambient temperature is a pivotal factor in the regulation of the flowering process in plants. In this study, we found that high ambient temperature exerts an inhibitory effect on the flowering of Osmanthus fragrans “Sjigui”. However, the underlying molecular mechanisms remain not fully understood. Through transcriptome analysis, a differently expressed C3H gene OfC3H49 was identified, which is induced by high ambient temperature. OfC3H49 was demonstrated to delay the flowering process of Arabidopsis and downregulate the expression of flowering-related genes in O. fragrans calli. Further investigation indicates that OfC3H49 as a transcriptional repressor directly suppresses the expression of the OfSOC1B thereby causing a delay in flowering time. Furthermore, a WRKY transcription factor, OfWRKY17, was identified to be responsive to high ambient temperature, directly binding to the OfC3H49 promoter and enhance OfC3H49 expression. Overexpression of OfWRKY17 in Arabidopsis resulted in a significant delay in flowering and induced the expression of OfC3H49 in O. fragrans calli. Collectively, our findings delineate a regulatory module, OfWRKY17-OfC3H49, which is activated by high ambient temperature and functions as a negative regulator of flowering by suppressing the expression of OfSOC1B in O. fragrans. This study provides novel insights into the molecular mechanisms involved in ambient temperature-mediated flowering control and contributes to the development of molecular breeding strategies for O. fragrans.

In the context of organic farming, the introduction of a local product to wider markets and an evaluation of storage effects, metabolic and transcriptomic variations in two broccoli rabe genotypes from production cycles of two different years were studied by comparing florets of stored fresh (SF) and packaged (P) for 4 days with those harvested fresh from the field (H). Twenty-five hydrosoluble compounds, including amino acids, carbohydrates, and organic acids, were quantified by untargeted nuclear magnetic resonance (NMR). Principal component analysis produced a neat separation among the three commodity statuses with P being the most divergent and SF closer to H. In the packaged florets, carbohydrate levels dropped significantly (over −52%), while the levels of amino acids and organic acids varied. There was an increase in stress-responsive phenylalanine and valine (over 30%) and succinic and α-ketoglutaric acids (over 75%). Compound correlation analyses indicated a carbohydrate sink towards γ-aminobutyric acid (GABA) and lactic acid (LA) metabolism under hypoxic conditions in packaged florets. RNA-seq analysis revealed that over 4000 genes were differentially expressed in SF vs H and 8000 in P vs H. Several CAR and AA pathways were significantly enriched in S and even more significantly in P, when compared to H. A map of gene expression (175 genes) and metabolite contents (14 compounds) was constructed to elucidate the gene routes that lead to accumulation of GABA and LA, known for healthy properties, in P. WGCNA and promoter binding site analyses enabled the identification of transcription factors (bZIP, WRKY, ERF types), interactions, and targeted genes encoding key enzymes in GABA and LA accumulation.

Fresh-cut fruit browning severely affects the appearance of fruit. Light treatment can effectively inhibit fresh-cut apple fruit browning, but the regulatory mechanism remains unknown. Here, we discovered that violet LED (Light-Emitting-Diode) light treatment significantly reduced fresh-cut apple fruit browning. Metabolomic analysis revealed that violet LED light treatment enhanced the phenolic accumulation of fresh-cut apple fruit. Transcriptomic analysis showed that the expression of phenolic degradation genes POLYPHENOL OXIDASE (MdPPO) and PEROXIDASE (MdPOD) was reduced, and the expression of phenolic synthesis gene PHENYLALANINE AMMONIA LYASE (MdPAL) was activated by violet LED light treatment. Moreover, two ELONGATED HYPOCOTYL 5 (MdHY5 and MdHYH) transcription factors involved in light signaling were identified. The expression of MdHY5 and MdHYH was activated by violet LED light treatment. Violet LED light treatment no longer inhibited fresh-cut apple fruit browning in MdHY5- or MdHYH- silenced fruit. Further experiments revealed that MdHY5 and MdHYH suppressed MdPPO and MdPOD expression and promoted MdPAL expression by binding to their promoters. In addition, MdHY5 and MdHYH bound to each other’s promoters and enhanced their expression. Overall, our findings revealed that violet LED light-activated MdHY5 and MdHYH formed a positive transcriptional loop to regulate the transcription of MdPPO, MdPOD, and MdPAL, which in turn inhibited the degradation of phenolics and promoted the synthesis of phenolics, thus inhibiting fresh-cut apple fruit browning. These results provide a theoretical basis for improving the appearance and quality of fresh-cut apple fruit.

Sugar limitation has dramatic consequences on plant cells, which include cell metabolism and transcriptional reprogramming, and the recycling of cellular components to maintain fundamental cell functions. There is however no description of the contribution of epigenetic regulations to the adaptation of plant cells to limited carbon availability. We investigated this question using nonphotosynthetic grapevine cells (Vitis vinifera, cv Cabernet Sauvignon) cultured in vitro with contrasted glucose concentrations. Sugar depletion in the culture medium led to a rapid cell growth arrest and a major metabolic shift that include the depletion in soluble sugar and total amino acids and modulation of the cell redox status. Consistently, flux modeling showed a dramatic slowdown of many pathways required for biomass accumulation such as cell wall and protein synthesis. Sugar depletion also resulted in a major transcriptional reprogramming, characterized by the induction of genes involved in photosynthesis, and the repression of those related to sucrose mobilization or cell cycle control. Similarly, the epigenetic landscape was deeply modified. Glucose-depleted cells showed a higher global DNA methylation level than those grown with glucose. Changes in DNA methylation mainly occurred at transposable elements, and at genes including some of those differentially expressed, consistent with an important role for methylation to the adaptation of cells to limited sugar availability. In addition, genes encoding histone modifiers were differentially expressed suggesting that additional epigenetic mechanisms may be at work in plant cells under carbon shortage.

Oligonucleotide (Oligo)-based fluorescence in situ hybridization (FISH) represents a highly effective methodology for identifying plant chromosomes. Longan is a commercially significant fruit species, yet lacking basic chromosomal markers has hindered its cytogenetic research. In this study, we developed a cost-effective oligo-based system for distinguishing chromosomes of longan (Dimocarpus longan Lour., 2n = 2x = 30). For this system, each synthesized oligo contained two chromosome-specific sequences that spanned a distance of over 200 kb, and a PCR-based flexible amplification method coupled with nested primers was used for probe labeling. The use of these oligo-based barcodes enabled the marking of 36 chromosomal regions, which allowed for the unambiguous distinction of all 15 chromosomes in both longan and lychee (Litchi chinensis Sonn., 2n = 2x = 30) species. Based on the identification of individual chromosomes, we constructed karyotypes and detected genome assembly errors involving the 35S ribosomal RNA gene (35S rDNA) in longan and lychee. Developing oligo-based barcodes offers considerable promise for advancing cytogenetic research in longan, lychee, and their related species. Furthermore, this cost-effective synthesis system can be referred to the development of new oligo libraries among other species.

Drought stress and lateral branches are both important factors affecting crop yield. Cucumber is a widely planted vegetable crop that requires a large amount of water during its production and varieties with few lateral branches are preferred. However, the mechanisms regulating cucumber drought tolerance and lateral branch development remain largely unclear. The MADS-box transcription factor AGAMOUS-LIKE 16 (CsAGL16) was recently found to be a key positive regulator in cucumber shoot branching acting by stimulating abscisic acid (ABA) catabolism. In this study, we demonstrated that cucumber TCP interactor containing EAR motif protein 1 (CsTIE1) directly interacts with CsAGL16 at protein level and promotes lateral branch outgrowth through the CsAGL16-CsCYP707A4 mediated ABA pathway in cucumber. Additionally, mutation of CsAGL16 resulted in decreased drought tolerance, while overexpression of CsAGL16 significantly enhanced drought tolerance in cucumber. Similarly, the drought resistance of Cstie1 mutants was significantly reduced. However, overexpression of CsAGL16 can enhance the drought tolerance of Cstie1 mutants and promote their lateral branch outgrowth. These results indicated that the CsTIE1-CsAGL16 module was crucial for both lateral branch development and drought response, providing a strategy for cultivating drought-tolerant cucumber varieties with appropriate branch outgrowth.

Root development is a complex process involving phytohormones and transcription factors. Our previous research has demonstrated that BcWRKY33A is significantly expressed in Bok choy roots under salt stress, and heterologous expression of BcWRKY33A increases salt tolerance and promotes root development in transgenic Arabidopsis. However, the precise molecular mechanisms by which BcWRKY33A governs root development remain elusive. Here, we investigated the role of BcWRKY33A in both root elongation and root hair formation in transgenic Bok choy roots. Our data indicated that overexpression of BcWRKY33A stimulated root growth and stabilized root hair morphology, while silencing BcWRKY33A prevented primary root elongation and resulted in abnormal root hairs morphology. Meanwhile, our research uncovered that BcWRKY33A directly binds to the promoters of BcLRP1 and BcCOW1, leading to an upregulation of their expression. In transgenic Bok choy roots, increased BcLRP1 and BcCOW1 transcript levels improved primary root elongation and root hair formation, respectively. Additionally, we pinpointed BcWRKY25 as a NaCl-responsive gene that directly stimulates the expression of BcWRKY33A in response to salt stress. All results shed light on the regulatory mechanisms governing root development by BcWRKY25-BcWRKY33A-BcLRP1/BcCOW1 module and propose potential strategies for improving salt tolerance in Bok choy.

Leucine-rich repeat receptor-like kinases (LRR-RLKs) have emerged as key regulators of herbivory perception and subsequent defense initiation. While their functions in grass plants have been gradually elucidated, the roles of herbivory-related LRR-RLKs in woody plants remain largely unknown. In this study, we mined the genomic and transcriptomic data of tea plants (Camellia sinensis) and identified a total of 307 CsLRR-RLK members. Phylogenetic analysis grouped these CsLRR-RLKs into 14 subgroups along with their Arabidopsis homologs. Gene structure and conserved domain analyses revealed notable similarities among subgroup members. Among the identified CsLRR-RLKs, we focused on two plasma membrane-localized LRR-RLKs, CsLRR-RLK44, and CsLRR-RLK239, which do not form homodimers or heterodimers with each other. Both respond strongly to herbivory, and their expression patterns significantly correlate with herbivore resistance phenotypes across different tea accessions. CsLRR-RLK44 and CsLRR-RLK239 act upstream of mitogen-activated protein kinase (MPK) cascades and modulate the expression of defense-related MPKs and WRKY transcription factors. Additionally, silencing CsLRR-RLK44 or CsLRR-RLK239 reduced the levels of herbivory-induced jasmonates, thereby weakening the plant resistance to tea geometrid larvae (Ectropis obliqua). Our work is the first to demonstrate that in woody plants, LRR-RLKs are essential for enhancing herbivore resistance through the activation of the canonical signaling, including MPKs, WRKYs, and jasmonates. Furthermore, our study extends mechanistic insights into how LRR-RLKs initiate plant defenses from grasses to economically important tree species.

Sugars act as signaling molecules to modulate various growth processes and enhance plant tolerance to various abiotic and biotic stresses. Moreover, sugars contribute to the postharvest flavor in fleshy fruit crops. To date, the regulation of sugar metabolism and its effect in plant growth, fruit ripening, postharvest quality, and stress resistance remains not fully understood. In this study, we investigated the role of tomato gene encoding a vacuolar invertase, hydrolyzing sucrose to glucose and fructose. SlVI is specifically expressed during the tomato fruit ripening process. We found that overexpression of SlVI resulted in increased leaf size and early flowering, while knockout of SlVI led to increased fruit sucrose content, enhanced fruit firmness, and elevated resistance of postharvest fruit to Botrytis cinerea. Moreover, the content of naringenin and total soluble solids was significantly increased in SlVI knockout fruit at postharvest stage. Transcriptome analysis showed a negative feedback regulation triggered by sucrose accumulation in SlVI knockout fruit resulting in a downregulation of BAM3 and AMY2, which are critical for starch degradation. Moreover, genes associated with cell wall, cutin, wax, and flavonoid biosynthesis and pathogen resistance were upregulated in SlVI knockout fruit. Conversely, the expression levels of genes involved in cell wall degradation were decreased in knockout fruit. These results are consistent with the enhanced postharvest quality and resistance. Our findings not only provide new insights into the relationship between tomato fruit sucrose content and postharvest fruit quality, but also suggest new strategies to enhance fruit quality and extend postharvest shelf life.

Fruit firmness is an important trait for characterizing the quality and value of apple. It also serves as an indicator of fruit maturity, as it is a complex trait regulated by multiple genes. Resequencing techniques can be employed to elucidate variations in such complex fruit traits. Here, the whole genomes of 294 F₁ hybrids of ‘Fuji’ and ‘Cripp's Pink’ were resequenced, and a high-density binmap was constructed using 5014 bin markers with a total map distance of 2213.23 cM and an average map distance of 0.44 cM. Quantitative trait loci (QTLs) of traits related to fruit were mapped, and an A-T allele variant identified in the coding region of MdNAC5 was found to potentially regulate fruit firmness and ripening. The overexpression of MdNAC5^A resulted in higher production of methionine and 1-aminocyclopropanecarboxylic acid compared to MdNAC5^T, leading to reduced fruit firmness and accelerated ripening in apples and tomatoes. Furthermore, the activities of MdNAC5^A and MdNAC5^T were enhanced through their differential binding to the promoter regions of MdACS1 and MdERF3. Spatial variations in MdNAC5^A and MdNAC5^T caused changes in MdACS1 expression following their interaction with MdERF3. Ultimately, utilizing different MdNAC5 alleles offers a strategy to manipulate fruit firmness in apple breeding.

DNA methylation is a stable epigenetic mark that plays a crucial role in plant life processes. However, the specific functions of DNA methylation in grape berry development remain largely unknown. In this study, we performed whole-genome bisulfite sequencing on ‘Kyoho’ grape and its early-ripening bud mutant ‘Fengzao’ at different developmental stages. Our results revealed that transposons (TEs) and gene flanking regions exhibited high levels of methylation, particularly in ‘Fengzao’, attributed to CHH site methylation. Interestingly, the methylation patterns in these two cultivars showed distinct dynamics during berry development. While methylation levels of genes and TEs increased gradually in ‘Kyoho’ throughout berry development, ‘Fengzao’ did not display consistent changes. Notably, ‘Fengzao’ exhibited higher methylation levels in promoters compared to ‘Kyoho’, suggesting that hypermethylation of promoters may contribute to its early ripening phenotype. Integration of methylome and transcriptome data highlighted differentially methylated genes (DMGs) and expressed genes (DEGs) associated with secondary metabolite biosynthesis, with 38 genes identified as potential candidates involved in grape berry development. Furthermore, the study identified a jasmonate-induced oxygenase gene (JOX1) as a negative regulator of ripening in Arabidopsis and grapes, indicating that hypermethylation of JOX1 may play a role in the early ripening of ‘Fengzao’. Overall, our findings provide insights into the distinct DNA methylation patterns during grape berry development, shedding light on the epigenetic regulatory mechanisms underlying the early-ripening bud mutant.

Branched-chain amino acids (BCAAs) are essential amino acids in tomato (Solanum lycopersicum) required for protein synthesis, which also modulate growth and abiotic stress responses. To date, little is known about their uptake and transport in tomato especially under abiotic stress. Here, the tomato amino acid permease 6 (SlAAP6) gene was identified as an amino acid transporter that restored mutant yeast cell growth on media with a variety of amino acids, including BCAAs. Overexpression of SlAAP6 (SlAAP6-OE) in tomato raised the BCAA content and elevated the fresh weight, while SlAAP6 knockouts (slaap6) showed reduced levels of neutral and basic amino acids in seedling tissues and lower total free amino acid distribution to shoots. In comparison to wild type and slaap6 mutants, SlAAP6-OE alleviated root limited growth by elevated BCAA transport and upregulated the expression of root-growth-related genes by increasing BCAAs in vivo. As SlAAP6 serves as a positive regulator for BCAA abundance, SlAAP6-OE lines showed greater salinity tolerance, while slaap6 mutants exhibited increased salt sensitivity. The salt tolerance of SlAAP6-OE plants was further enhanced by the application of exogenous BCAAs. In addition, BCAA supplementation reduced the accumulation of H₂O₂ in root under salt stress conditions. Based on these findings, SlAAP6-mediated uptake and transport of BCAAs facilitated growth and salt tolerance in tomato. By characterizing this key amino acid transporter, this study provides a novel approach to simultaneously enhance tomato nutritional quality, growth and development, and stress resistance through genetic improvement.

Glycerophosphodiester phosphodiesterase 1 (GDPD1) plays an important function in the abiotic stress responses and participates in the accumulation of sn-glycerol-3-phosphate (G3P) in plants, which is key to plant systemic acquired resistance (SAR). However, the role of GDPD1 in plant responses to biotic stress remains poorly understood. This study characterized the antivirus function of the GDPD1 gene (designated as ClGDPD1) from Eureka lemon. ClGDPD1 is located in the membrane and endoplasmic reticulum, where it interacts with the citrus yellow vein clearing virus (CYVCV) coat protein (CP). Compared to individually expressed ClGDPD1 or coexpressed ClGDPD1+CP_140-326, transiently coexpressed ClGDPD1+CP or ClGDPD1+CP_1-139 significantly upregulated the key substance contents and genes expression involved in glycerophospholipid metabolism. Over-expression of ClGDPD1 significantly facilitated the accumulation of G3P, upregulated the expression of SAR-related genes, and increased the resistance of transgenic Eureka lemon to CYVCV infection. Furthermore, exogenous glycerol treatment and over-expression of ClGPDH increased the G3P content and reduced CYVCV titers in plants or hairy roots. These results indicated that the enhanced resistance of ClGDPD1 transgenic Eureka lemon to CYVCV may be due to facilitating G3P accumulation through the interaction of ClGDPD1 with CP. Our findings provide novel insights into the role of ClGDPD1 as an important regulatory center in mediating the citrus defense response to viral infections.

Prefertilization hybridization barriers are the main causes of intersubgeneric hybridization challenges in water lily. However, the mechanism underlying low compatibility between pollen and stigma of water lily remains unclear. This study demonstrates that CBL-interacting protein kinase 6 (CIPK6) responded to the signaling exchange between incompatible pollen and stigma through interactions with SNF1-related kinase 1 (SnRK1) and promotes the accumulation of SnRK1 protein. Activated SnRK1 interacted with 9-cis-epoxycarotenoid dioxygenase 2 (NCED2) to promote its degradation, thereby inhibiting abscisic acid (ABA) synthesis. A decrease in ABA content in the stigma impaired the ABA-mediated removal of reactive oxygen species (ROS), ultimately resulting in the rejection of the incompatible pollen by the stigma. Our results highlight the essential role of the NpCIPK6-NpSnRK1-NpNCED2 module in conferring intersubgeneric hybridization barriers in water lily by interfering with ABA synthesis and promoting ROS accumulation. This study offers valuable mechanistic insights into cellular signaling and reproductive barriers in water lily as well as across other biological contexts.

GRAS, termed after gibberellic acid insensitive (GAI), RGA (repressor of GA1), and SCR (scarecrow), is a plant-specific transcription factor crucial for plant development and stress response. However, understanding of the functions played by the GRAS members and their target genes in citrus is limited. In this study, we identified a cold stress-responsive GRAS gene from Poncirus trifoliata, designated as PtrPAT1, by yeast one-hybrid library screening using the promoter of PtrBADH-l, a betaine aldehyde dehydrogenase (BADH)-like gene. PtrPAT1, belonging to the PAT1 subfamily, was localized in the nucleus and plasma membrane, exhibited transactivation activity and showed a remarkable upregulation under cold stress. Overexpression of PtrPAT1 elevated BADH activity, increased glycine betaine (GB) accumulation, and conferred enhanced cold tolerance in transgenic tobacco plants compared with wild type, while downregulation in trifoliate orange by virus-induced gene silencing (VIGS) resulted in opposite trends. Furthermore, the activities of two antioxidant enzymes, including peroxidase (POD) and superoxide dismutase (SOD), were significantly increased in the overexpression plants, but remarkably decreased in the VIGS line, consistent with accumulation patterns of the reactive oxygen species (ROSs). PtrPAT1 was demonstrated to interact with and activate the PtrBADH-l promoter through the putative PAT1-binding motif with the core sequence of TTTCATGT, indicating that PtrBADH-l is a target gene of PtrPAT1. Taken together, these results demonstrate that PtrPAT1 positively affects cold tolerance through the regulation of GB biosynthesis by modulating PtrBADH-l expression.