Flavonoids produced by legume roots act as signaling molecules that induce the expression of nod genes in symbiotic rhizobia. However, the role of flavonoids in root exudates under intercropping systems in promoting soybean nodulation remains unclear. Two consecutive years of field experiments were conducted using maize–soybean strip intercropping with interspecific row spacings of 30cm (MS30), 45cm (MS45), and 60cm (MS60), along with sole cropping of soybean(SS) and maize (MM). Root interactions were manipulated using either no root barrier (NB) or a polyethylene plastic barrier (PB) to assess the relationship between flavonoids in root exudates and soybean nodulation. We found that root–root interaction between soybean and maize increased nodule number and fresh weight in intercropped soybean, with enhancement gradually increasing as interspecific distancewidened. The proportion of nodules with diameters exceeding 0.4cm was higher in intercropped soybean under NB compared to PB. Additionally, the expression of nodule-related genes-GmENOD40, GmNIN2b, and GmEXPB2-was up-regulated. Furthermore, compared to monocropping, isoflavone secretion by soybean roots decreased, whereas flavonoid and flavonol secretion by both maize and soybean roots increased under intercropping. The abundance of differentially secreted flavonoid metabolites in the rhizosphere of both species declined when rootcontact was prevented by the barrier. In soybean roots, the expression of GmCHS8and GmIFS1was up-regulated, while GmICHGwas down-regulated under root interaction. Most flavonoid and flavonol compounds showed positive correlations with nodule diameter. Nodule number, fresh weight, and the proportion of nodules larger than 0.2cm increased in diverse soybean genotypes treated with maize root exudates, which contributed to enhanced nitrogen fixation capacity. Therefore, maize–soybean strip intercropping, combined with optimal row spacing, enhances the positive effects of underground root interactions and improves nodulation and nitrogen fixation in intercropped soybean.

Hydraulic theory predicts a positive coupling between leaf hydraulic conductance (K_leaf) and stomatal conductance (g_s); however, this theory has not been fully supported by observations, and underlying mechanisms are poorly understood. Partitioning K_leafinto inside-xylem (K_x) and outside-xylem (K_ox) components offers a refined framework for elucidating the regulation of g_sby leaf hydraulics. While optimal planting density may enhance water use efficiency (WUE) through modulation of g_s, corresponding changes in leaf hydraulic properties and their influence on gas exchange remain unclear. We examined relationships among K_x, K_ox, g_s, leaf photosynthetic rate (A_N), and WUE, and analyzed the structural determinants of K_ox in cotton grown under eight planting densities: 12, 18, 24, 36, 48, 60, 72, and 84 plants m^–2. Results showed that as planting density increased, K_leafand A_Nremained stable, whereas K_oxand g_sdeclined significantly. Leaf thickness and the volume fraction of inter-cellular air space were key structural factors influencing K_ox. Neither K_leafnor K_xcorrelated with A_Nor g_s; however, K_oxexhibited a significant positive correlation with g_s. Furthermore, K_oxwas negatively correlated with WUE. These findings indicate that K_oxmodulates g_sto minimize water loss without compromising A_N, thereby enhancing WUE in cotton across varying planting densities.

Softening of fleshy fruits during ripening and postharvest is a programmed physiological process that substantially impacts fruit quality and shelf life. However, the molecular mechanism underlying peach softening remains largely unknown.Lateral organ boundary (LOB) domain (LBD) proteins are pivotal regulators of plant growth and development. To date, certain LOB/LBD transcription factors are seemingly implicated in fruit softening. In this study, we identified 42 LOB/LBDgenes in the peach genome. Expression analysis showed a significant upregulation of PpLOB1transcripts toward peach fruit ripening. PpLOB1 was classified into Class II subgroup, and showed high sequence similarity to several softening-related LOB/LBD transcription factors. Transient transformation assays showed that PpLOB1positively modulates peach softening. Further experiments demonstrated that PpLOB1 directly targets and activates the promoters of pectatelyase1(PpPL1) and PpPL15, thereby contributing to the regulation of fruit softening. Additionally, PpNAP4 up-regulates PpLOB1expression by binding to its promoter. Meanwhile, our findings revealed that PpNAP4 and PpNAP6 cooperatively modulate the expression of PpLOB1. Taken together, our findings revealed a novel regulatory module involving PpNAP4 and PpLOB1 that modulates peach fruit softening.

Cold stress represents a critical constraint on crop productivity, particularly in temperate climates. Despite the established role of abscisic acid (ABA) in cold stress responses, the precise mechanisms through which transcription factors mediate ABA-dependent cold tolerance remain elusive. Here, we identify VaMYB4a, a MYBtranscription factor from VitisamurensisRupr. (Amur grape), as a key regulator of cold tolerance. It integrates ABA signaling with the CBF(C-repeat binding factors)-COR(cold-regulated) pathway to orchestrate cold stress adaptation. Through a combination of overexpression and CRISPR/Cas9-mediated knockout lines in Arabidopsisthaliana, grape callus, and Vitisvinifera L. seedlings, we demonstrate that VaMYB4aenhances freezing tolerance by promoting osmotic regulation, reactive oxygen species (ROS) scavenging, and stomatal closure. VaMYB4a functions as a homo-dimer, with its C-terminal domain being essential for transcriptional activation. Mechanistically, VaMYB4a directly upregulates CBFand CORgenes while fine-tuning ABA signaling components such as ABI1and ABF4. Notably, ABA exhibits a dual role: enhancing VaMYB4a-mediated freezing tolerance under short-term stress but attenuating its effects during prolonged cold exposure, revealing an intricate regulatory crosstalk between cold and hormonal pathways. Our work not only advances the molecular understanding of cold adaptation but also provides a promising genetic target for developing stress-resilient grape varieties to mitigate the impacts of climate change.

Heat stress reduces theanine content in tea plants, but the underlying molecular mechanism remains unclear. In this study, a temperature gradient treatment (20°C, 25°C, 30°C, and 35°C) was performed to unveil the effect of heat stress on biosynthesis and accumulation of theanine. We found that heat stress induced metabolic changes, characterized by decreased theanine content and increased catechin levels. In addition, heat stress up-regulated the expression of the class B heat shock transcription factor gene CsHSFB2c, while significantly suppressing the transcription of key theanine biosynthetic genes CsTS1and CsGS1. Functional studies showed that silencing CsHSFB2cincreased theanine content, while its overexpression significantly decreased theanine levels. Consistent with these changes, silencing CsHSFB2cupregulated the expression of CsTS1and CsGS1, while overexpression of CsHSFB2cdownregulated their expression. Yeast one-hybrid (Y1H) and dual-luciferase reporter gene (Dual-LUC) assays showed that CsHSFB2c directly binds to the promoters of CsTS1and CsGS1and inhibits their expression. These results demonstrate that CsHSFB2c mediates heat-induced suppression of theanine biosynthesis by directly inhibiting the expression of CsTS1and CsGS1. This study provides a theoretical basis for improving the heat resistance and quality of tea plants viamolecular breeding.

Caffeic acid-O-methyltransferase (COMT) is a crucial enzyme in the phenylpropanoid metabolic pathway, with significant roles in both the lignin and coumarin pathways. The function of COMTin plant disease resistance has been demonstrated in several species. Our research identified the potato COMTgene family on a genome-wide scale and StCOMT1as a candidate gene for enhancing potato disease resistance under DON induction through phylogenetic analyses combined with previously identified metabolic differences and weighted gene co-expression network analysis (WGCNA) results. In order to better understand the function of StCOMT1, heterologous expression and overexpression assays were conducted. StCOMT1 is localized in chloroplasts and was found to catalyze the methylation of substrates to produce ferulic acid and melatonin in vitro. Physiological parameters showed that, compared with wild-type potato plants, StCOMT1-overexpressing plants infected with Fusariumsporotrichioidesexhibited smaller lesion areas and lower reactive oxygen species (ROS) levels. High-performance liquid chromatography (HPLC) and RT-qPCR analyses revealed organ-specific accumulation of coumarin-related compounds and organ-specific expression of their corresponding genes in StCOMT1-overexpressing plants post-inoculation. The results indicate that StCOMT1overexpression in potatoes enhanced resistance to F. sporotrichioidesby enhancing reactive oxygen species clearance and promoting organ-specific accumulation of coumarin-related compounds.

Flavonols possess significant medical value and are essential for plant stress resistance. These compounds constitute primary components of the nutritional value in onions, particularly in edible portions. While the flavonol biosynthetic pathway has been extensively studied, its regulatory mechanisms in onions remain incompletely understood. This investigation identified flavonol biosynthesis and regulatory genes through analysis of transcriptome and metabolomics data from different developmental stages of ‘SA1’. Two R2R3-MYB transcription factors, AcMYB12 and AcMYB29, were identified as positive regulators of onion flavonol biosynthesis. Transcriptional activation assays demonstrated that both could activate AcCHS, AcF3´H, and AcFLS.Yeast one-hybrid assays confirmed their direct binding to these gene promoters. The expression levels of flavonol pathway genes and flavonol contents in AcMYB12/AcMYB29-overexpressing onion calli and Arabidopsis plants were significantly higher than those in the control group. Transient silencing assays revealed partial functional redundancy between these two transcription factors. Notably, their regulatory capabilities exhibited significant differences. AcMYB12 predominantly regulates flavonol accumulation, while AcMYB29 specifically influences quercetin. Further investigation of the molecular mechanisms underlying differential regulation indicated variations in cis-elements within flavonol pathway gene promoters and differences in binding activity between transcription factors and cis-elements.

Fusarium head blight (FHB), mainly caused by fungus Fusariumgraminearum, is a devastating wheat disease worldwide, leading to reduced yield production and compromised grain quality due to contamination by mycotoxins, such as deoxynivalenol (DON). Manipulating the specific gene expression in microorganisms through RNA interference (RNAi) presents an opportunity for new-generation double-stranded RNA (dsRNA)-based formulations to combat a large number of plant diseases. Here, we applied both spray-induced gene silencing (SIGS) and host-induced gene silencing (HIGS) to target five virulence-related and DON-synthesized genes in F.graminearum, including protein kinase gene Gpmk1, zinc finger protein gene FgChy1, transcription factor FgSR, DON synthesis gene TRI5and the cell-end marker protein gene FgTeaA, aiming to effectively control FHB in wheat. Direct spraying of individual or combined small interfering RNA (siRNAs) from the fungus showed reduced expression of target genes and suppressed pathogenic symptoms during F.graminearuminfection in wheat leaves, with the combination of all five siRNAs demonstrating superior resistance. Furthermore, we generated transgenic wheat lines expressing chimeric RNAi cassettes targeting these five genes, and two independent lines exhibited strong resistance to FHB and Fusarium crown rot, and the reduced DON accumulation. Notably, the HIGS transgenic lines did not adversely impact plant growth and yield traits. Collectively, our findings support that SIGS and HIGS represent effective strategies targeting key pathogenic genes for bolstering disease resistance in crops.

Nicotinamide mononucleotide (NMN), a precursor in nicotinamide adenine dinucleotide (NAD) biosynthesis, has long been recognized for its pivotal role in medicine. Recent investigations have suggested its potential as a plant immunity inducer for controlling fungal diseases. However, whether NMN confers plant broad-spectrum resistance against diverse phytopathogens, and its underlying mechanisms remain ambiguous. In this study, we investigate the effect of NMN against multiple phytopathogens in tobacco. Our results demonstrate that tobacco pretreated with NMN exhibits enhanced resistance against RalstoniasolanacearumCQPS-1, PseudomonassyringaeDC3000 ΔhopQ1-1, Phytophthoraparasitica, and tobacco mosaic virus (TMV). NMN displays effectiveness within the concentration range of 50–600μmol L^–1, with 75μmol L^–1NMN exhibiting the most pronounced effect. The impact of NMN pretreatmentcould persist for up to 10 days. Beyond tobacco, NMN pretreatment enhances disease resistance in tomato and pepper plants against diverse pathogens, underscoring NMN’s capacity to confer broad-spectrum disease resistance in crops. Moreover, RT-qPCR analysis reveals that NMN significantly upregulates the expression of the pattern-triggered immunity (PTI) marker gene NbCYP71D20and salicylic acid (SA) marker gene NbPR1a. This suggests that NMN enhances plant resistance by inducing both PTI and SA-mediated immunity. Interestingly, the positive impactof NMN on plant disease resistance is not significantly compromised in both NMNadenylyltransferase(NMNAT)-silenced plants and NAD receptor mutant lecrk-I.8, suggesting the existence of NAD-independent signaling pathways for NMN-induced plant immunity. In conclusion, our study establishes that the bioactive molecule NMN imparts broad-spectrum disease resistance in plants, offering a simple, environmental-friendly, and promising strategy for safeguarding crops against diverse phytopathogens. These findings also provide valuable insights for future in-depth studies into the functional mechanisms of NMN.

The Loxostegesticticalis(Lepidoptera: Pyralidae) is a major migratory pest of agriculture and animal husbandry in Asia and Europe. Utilizing plant volatile organic compounds (pVOCs) as attractants for monitoring and controlling pests is considered an environmentally friendly and effective method. However, limited knowledge exists regarding applying pVOCsto manage L.sticticalis. Here, volatile compounds released byChenopodiumalbum, Setariaviridis, and Medicagosativa,the three preferred oviposition plantsforL.sticticalisfemales,were collectedusing dynamic headspace sampling techniques. A total of 55distinctcompounds were identified through gas chromatography-mass spectrometry (GC-MS), and16 compounds in the concentration range from 0.001 to 100 µg µL^–1elicited consistently enhanced electrophysiological responses in both male and female L.sticticalis. Subsequently, the attraction potential of four bioactive compounds-linalool, cis-anethole, trans-2-hexenal, and 1-octen-3-ol-were further confirmed by indoor behavioral bioassays. The blends of linalool, cis-anethole, trans-2-hexenal, and 1-octen-3-ol mixed at ratios of 5:1:5:10 (formulation No. 25) and 5:1:1:10 (formulation No. 21) were highly attractive to L.sticticalisadults. Field-trapping assays indicated that lure No. 2 baited with formulation 21 demonstrated superior efficacy in field trapping. These findings suggest that pVOC-based attractants can be effectively employed for monitoring and mass trapping L.sticticalisadults, providing insights into the development of botanical attractants.