Earthworms and plants interact synergistically in soil ecosystems and influence nitrogen cycling processes. However, the effects of earthworms on microbial communities and nitrogen metabolism in the plant rhizosphere remain unclear. In this study, high-throughput sequencing was used to characterize the microbial communities in the drilosphere and elucidate how functional nitrogen metabolism genes respond to earthworm activity and plant growth. The results showed increased relative abundances of Proteobacteria and Bacteroidetes and decreased abundances of Patescibacteria and Chloroflexi in soil microdomains affected by earthworms. Earthworm-responsive microorganisms were clustered into specific effector modules within the bacterial co-occurrence network. In the drilosphere, these modules were positively correlated with the abundances of amoA, nrfA, and nifH. In the rhizosphere, they were positively correlated with nifH but negatively correlated with nrfA and nirK gene abundances. The observed enrichment of nitrogen-cycling functional groups was associated with elevated soil pH and nutrient levels, particularly available phosphorus. Overall, earthworm activity modulated the abundance and composition of nitrogen-metabolizing microbial communities, thereby contributing to enhanced nitrogen cycling. This study provides a foundation for understanding the beneficial effects of earthworms on soil nitrogen pools and nitrogen-functional microbiota, offering insights into sustainable agriculture in nutrient-deficient soils.

Abstract Chemically inducible expression systems enable transgene expression regulation in response to external small molecules. Tetracycline repressor (TetR)-based gene switches work in plants, but antibiotics are neither approved nor advisable for crop use. Here we report engineering of TetR mutants that respond to approved sulfonylurea (SU) herbicides instead of antibiotics. Designed variants show low-nanomolar EC 50 values for ethametsulfuron-methyl (Es) or chlorsulfuron and tightly bind the Tet operator sequence, but only in the absence of corresponding SUs. Crystal structures of two repressors in complex with their respective SU ligands reveal extensive interactions explaining their strong binding. The Es repressor-based gene switch is introduced into tobacco, soybean, maize, rice, and Arabidopsis , and robust reporter gene activation is observed upon herbicide application. Addition of a repressor-regulated siRNA targeting the repressor transcript increases the magnitude and spatial distribution of the response following herbicide treatment and results in a partially bistable gene switch. The SU repressors also function well in mammalian cell culture and may enable regulation of additional genes in conjunction with TetR.

Rice sheath blight is a major fungal disease threatening global rice production, causing substantial yield losses and lacking effective resistant cultivars. In this study, we systematically evaluated the biocontrol efficacy of Bacillus velezensis BER1 against rice sheath blight and elucidated its underlying mechanisms from three complementary perspectives: strain genomics, plant immune responses, and the phyllosphere microbiome. Genome analysis revealed that BER1 harbors 22 secondary metabolite biosynthetic gene clusters, including those encoding natural products such as fengycin and bacillaene with broad-spectrum antifungal and antibacterial activities. Application of BER1 reduced disease severity to 35.4%, outperforming conventional chemical treatment, and significantly induced the expression of rice immune-related genes (e.g., NH1 and PR1a), accompanied by increased activities of defense enzymes such as POD and PAL. 16S rRNA amplicon sequencing further showed that BER1 markedly increased the relative abundance of Bacillus in the phyllosphere while suppressing potential pathogenic taxa, and enriched functional pathways associated with secondary metabolite biosynthesis, particularly polyketide-related pathways. Compared with chemical intervention, BER1 maintained microbial community stability through ecological niche competition and sustained metabolic activity, thereby reducing pathogen risk. Collectively, these findings provide quantitative evidence supporting BER1 as a promising green and sustainable biocontrol agent against rice sheath blight and highlight the potential of microecology-based strategies for plant disease management.

In modern agriculture, Bacillus-based inoculants are widely used due to their environmental adaptability, spore formation, and functional versatility; however, systematic comparisons among different Bacillus strains remain limited. Using rice as a model crop, we compared Bacillus velezensis BER1 with a commercial Bacillus subtilis formulation in a simplified sterile-substrate pot system, focusing on rice growth, rhizosphere-associated bacterial communities, and predicted nutrient-cycling potential. The results showed that BER1 increased root and shoot dry biomass to 0.52 g and 0.77 g, respectively, while leaf total nitrogen, phosphorus, and potassium contents reached 2.9%, 0.32%, and 2.87%. Rhizosphere microbiome analysis revealed that BER1 was associated with changes in bacterial community composition and β-diversity, the enrichment of several taxa potentially related to nutrient cycling and plant growth promotion, and a more complex microbial co-occurrence network dominated by positive correlations under sterile-substrate conditions. Comparative genomic analysis demonstrated that BER1 harbors a richer repertoire of plant growth-promoting genes and secondary metabolite biosynthetic gene clusters, including NRPS/PKS modules and a specific glpQ4 gene potentially involved in organic phosphorus mobilization. Structural equation modeling suggested a potential association pathway linking bacterial inoculation, predicted microbial functional profiles, soil properties, plant nutritional status, and biomass formation. Overall, BER1 promoted rice growth in this simplified sterile-substrate pot system, likely through its genomic potential and associated rhizosphere bacterial responses.

In grape, EXO70 subunits are essential components of the exocyst tethering complex, which is involved in complex assembly, plant growth and stress-related processes. However, the exact function it performs in facilitating tolerance to drought and cold conditions has yet to be clarified. In this study, we functionally characterized a grape EXO70 isoform, VvEXO70A1, which is predominantly localized at the plasma membrane. Evolutionary analysis of the EXO70 gene family across 11 species indicated that grape EXO70 genes share a conserved phylogenetic history with their orthologs from dicotyledons. The expression of VvEXO70A1 was elevated in overexpression (OE) lines Arabidopsis and grape calli exposed to water deficit and low-temperature treatments. Compared with wild type (WT), VvEXO70A1 overexpressing Arabidopsis and grape calli exhibited enhanced tolerance under these abiotic stresses, which was characterized by both lower levels of oxidative stress markers and a more active antioxidant machinery with substantially higher activities of major ROS-scavenging enzymes, including CAT, SOD, and POD. Notably, the transgenic plants were more sensitive to abscisic acid (ABA), displaying stronger ABA-induced stomatal closure. This phenomenon was further enhanced under drought stress, with the expression of the ABA signaling gene ABF4 being significantly upregulated. Cold stress-related genes (CBF1/2/3, ICE1/2, and ICE1a/b/c) were also strongly induced when the OE lines underwent low-temperature treatment. Overall, overexpression of VvEXO70A1 functions as a positive regulator, boosting tolerance to drought and cold conditions which was accompanied by reduced ROS damage and elevated expression levels of key stress-responsive genes associated with drought and cold in both transgenic Arabidopsis and grape calli.

Global increases in the intensity and frequency of elevated temperatures is threatening ecosystem stability and crop yield. Understanding plant thermomorphogenesis is critical for developing climate-resilient crops, yet the underlying mechanisms remain to be clarified. Here, we identify the BEL1-LIKE HOMEODOMAIN transcription factor BLH1 as a critical negative regulator of thermomorphogenesis that modulates the key BRASSINAZOLE-RESISTANT 1 (BZR1)-PHYTOCHROME INTERACTING FACTOR 4 (PIF4) thermomorphogenic regulatory module. Overexpression of BLH1 or its homologs confers high-temperature (HT) insensitivity, whereas blh higher-order mutants exhibit HT hypersensitivity. BLH1 expression is directly repressed by BZR1 and is down-regulated by HT. We further demonstrate that BLH1 directly binds to the PIF4 promoter to repress its transcription and concurrently interacts with the PIF4 protein to inhibit its activity. Overexpression of BLH1 rescues the elongated hypocotyl phenotype in bzr1-1D or PIF4 overexpression plants. Our findings define a BZR1-BLH1-PIF4 regulatory axis that modulates the BZR1-PIF4-auxin-BR-BZR1 positive feedback loop, ensuring a balanced thermomorphogenic response to HT.

Soil salinity severely reduces agricultural productivity by impairing seed germination and plant growth, thereby threatening the sustainable development of eco-friendly farming worldwide. The utilization of seed nanopriming, specifically using carbon dots (CDs), alleviates the side effects of salt stress during seed germination in cotton (Gossypium hirsutum. L), but the underlying mechanisms remain poorly characterized. In this study, we investigated the impact of CDs priming on cotton seeds by assessing their physiological, biochemical, and transcriptomic responses under salinity stress. Nanopriming with CDs significantly increased seedling root length (205.2 %), germination rate (32 %) and seed vigor index (378.4 %) under salt stress compared with controls (water priming). CDs primed seeds showed significantly lower content of superoxide anion radicals (O2 center dot-) and malondialdehyde (MDA) while hydrogen peroxide (H2O2) levels increased by 42.3 %-48.5 %. Under salt stress conditions, CDs priming also resulted in a significant reduction in Na+ accumulation (15.3 % lower than the control) without affecting K+ content. Further research found that the enhanced Na+ efflux and seed germination induced by CDs priming were substantially suppressed by an NADPH oxidase inhibitor, diphenyleneiodonium chloride (DPI). Thus, we confirmed that GhRboh-mediated H2O2regulates Na+ homeostasis to promote seed germination under salinity. Transcriptome sequencing (RNA-Seq) results suggested that CDs priming-induced salt stress resistance is likely linked to oxidative stress response, MAPK signaling pathway, cellular ion homeostasis and Ca2+-binding proteins. Moreover, CDs priming treatment significantly upregulated the relative expression levels of GhRboh genes, Ca2+ influx genes and SOS1/NHX7. These results indicate that GhRboh-mediated H2O2 accumulation may modulate Na+ homeostasis via the Ca2+-dependent Na+/H+ antiporter system to increase salt tolerance in cotton seed. This study provides novel mechanistic insights into nanomaterial-based seed priming strategies for improving crop resilience in saline soils.

Lycoris are well-known bulbous ornamental plants with both ornamental and economic value, displaying diverse floral traits, particularly an extensive array of petal colors. The red pigmentation of Lycoris petals is predominantly attributed to anthocyanin accumulation. However, the underlying regulatory mechanism of anthocyanin biosynthesis in Lycoris remains elusive. In this study, we conducted a metabolomic analysis that revealed both the total anthocyanin content and the relative abundance of individual anthocyanin species are critical contributors of the color variation observed between white- and red-petaled cultivars. Integrating transcriptomic data with metabolic profiles, we identified a gene module central to anthocyanin biosynthesis, with LrMYB114 emerging as a key transcriptional activator. Functional validation through overexpression and silencing of LrMYB114 in Lycoris petal confirmed its essential role in regulating total anthocyanin, cyanidin-3-O-sambubioside (Cy3Sa), and pelargonidin-3-O-sambubioside (Pg3Sam) levels. Furthermore, it was found that LrMYB114 could interact with the bHLH transcription factor LrbHLH94 to form an LrMYB114–LrbHLH94 complex that activated anthocyanin biosynthetic genes, essential for pigment accumulation in Lycoris petals. Futhermore, LrMYB114–LrbHLH94 complex upregulates transcription of the anthocyanin biosynthetic gene LrDFR1, LrDFR3, and LrPAL, leading to enhanced Cy3Sa and Pg3Sam accumulation. These findings highlight the LrMYB114–LrbHLH94 regulatory module as a critical determinant of petal coloration, offering potential avenues for improving petal quality through genetic manipulation.

The combined application of nitrogen (N) fertilizer and straw return is a promising strategy for improving agricultural sustainability. However, how these practices interact with soil fertility to influence arbuscular mycorrhizal fungi (AMF)-mediated plant performance and ecosystem multifunctionality in paddy soils remains unclear. In this study, we collected two paddy soils with contrasting fertility levels and amended them with three straw types—wheat straw (WS), rape straw (RS), and Astragalus sinicus L. (AS)—under varying N fertilization rates for rice cultivation. We quantified AMF abundance, diversity, and community composition, together with multiple ecosystem function variables in hyphosphere soil, and examined their relationships with plant performance. The effects of N fertilization and straw incorporation on rice productivity depended strongly on initial soil fertility. In low-fertility soil, high N input (150 mg N kg−1) combined with straw return, especially AS straw, significantly increased rice N uptake and biomass. In contrast, in high-fertility soil, low N input (75 mg N kg−1) with straw return was optimal for biomass production. Partial least squares path modeling (PLS-PM) showed that N input level and straw type affected plant N uptake and growth by modifying hyphosphere AMF attributes and soil ecosystem functions. Regarding AMF attributes, greater plant biomass in both soils was associated with a higher relative abundance of Glomus sp. clB, as identified by random forest analysis. By integrating 16 ecological functions, we found that N fertilization under straw-return conditions significantly enhanced most soil functions and the ecosystem multifunctionality index, both of which were closely associated with increased rice N uptake and biomass. Overall, our findings demonstrate that optimizing N and straw inputs can improve crop productivity by regulating AMF communities and soil functions, although the optimal strategy is fertility dependent. We therefore advocate site-specific, fertility-based nutrient management that leverages beneficial AMF taxa to maximize both yield and ecosystem multifunctionality in sustainable rice production.

Methane (CH4) emissions from paddies, driven by methanogens and methanotrophs, are major agricultural CH4 sources. While fertilization-induced soil nutrient dynamics and rice growth regulate CH4 fluxes, mechanistic responses to phosphorus (P) availability remain understudied relative to carbon and nitrogen. Based on a long-term paddy experiment without P input in Tai Lake region since 1980, we investigated P addition effects on CH4 emissions, microbial functional genes and communities across four treatments: chemical nitrogen and potassium fertilizer (CNK) vs. CNK+P, chemical NK combined with organic fertilizer (MNK) vs. MNK+P. Relative to CNK and MNK, P addition reduced CH4 emissions by 19.9 % and 35.4 %, respectively, with flux variation positively correlated with mcrA gene at tillering. Unlike genes, alpha diversity and community composition of methanogens/methanotrophs were unaffected by P addition, and treatment differences were mainly driven by historical N fertilizer types. Notably, P addition suppressed microbial P-mining capacity, evidenced by decreased phosphatase activities and phoC and phoD gene abundances, thereby reducing methanogenic substrate levels, reflected by lower dissolved organic carbon and root-secreted organic acids. Furthermore, P input did not disrupt the dominance of type I methanotrophs shaped by historical fertilization, which showed superior CH4 oxidation efficiency. The reduction of CH4 emissions under P addition was driven by the substrate limitation of methanogens, without altering the dominant position of type I methanotrophs. This study highlighted the critical role of P availability in regulating paddy CH4 emissions and advocated optimized P management to balance rice productivity and CH4 mitigation.

Maize lethal necrosis (MLN) is a severe disease caused by the combined infection of maize chlorotic mottle virus (MCMV) and a potyvirus, most often sugarcane mosaic virus (SCMV). This disease seriously threatens food security across sub-Saharan Africa (SSA). We investigated a major-effect quantitative trait locus for resistance on chromosome 6, named the maize lethal necrosis susceptibility locus 1 (qMLNS1), derived from the Thai line KS23-6. Fine mapping and CRISPR-Cas9 editing of the candidate genes within the narrowed 105 kb interval revealed a peroxisomal peptidase as the underlying cause of susceptibility. Confocal microscopy confirmed the localization of the MLNS1 protein within peroxisomes. Targeted knockout of the Mlns1 gene in the susceptible elite line CML536 from SSA conferred resistance comparable to KS23-6 in field trials conducted in Naivasha, Kenya. This knockout specifically blocked MCMV accumulation without affecting SCMV. The edited lines showed no yield penalty or agronomic defects under disease-free conditions. Our findings uncover a mechanistic link between a peroxisomal enzyme and viral susceptibility. They also establish a rapid, scalable gene editing strategy for incorporating MLN resistance into elite germplasm, offering a model for combating similar viral diseases in staple crops globally.

Splicing dysregulation frequently occurs in cancers, yet the functional implications of most mis-spliced genes remain elusive. Through a comprehensive pan-cancer analysis of splicing dysregulation, we identified universally mis-spliced genes in cancers and were surprised to find their significant functional enrichment in cell migration. Interestingly, the STE20-like protein kinase (SLK) gene encoding a scaffold protein showed a significant splicing shift from SLK-S to SLK-L isoforms across various cancer types. Detailed studies demonstrated this splicing shift promoted cancer metastasis in cellular and animal models. Mechanistically, RNA-binding Fox-1 Homolog 2 (RBFOX2) protein was identified as a splicing factor that regulates SLK splicing. The two isoforms interact with occludin with different affinities, contributing to their unique activities in cancer metastasis. Notably, the antisense oligonucleotides designed to suppress SLK-L splicing effectively inhibited cancer migration and invasion. Collectively, this study shows a new splicing switch with a key role in controlling cancer metastasis, shedding light on new cancer therapy via splicing manipulation.

Selenium (Se) deficiency is a widespread health issue. Investigating the differences in Se bioavailability in agricultural products can provide valuable insights into prioritizing food choices for Se biofortification strategies. This study investigated the effects of food matrix (rice and vegetables) and Se dose on Se retention in tissue by administering SeMet, SeCys2, MeSeCys, and Se-rich rice and vegetables to mice at varying doses. Results showed that following ingestion of vegetables and rice, Se accumulation in mouse liver and kidneys exhibited non-linear increases with Se doses. Additionally, by giving mice celery, rice, SeMet, SeCys2, and MeSeCys with different Se doses, Se concentration in liver and kidneys of both male and female mice increased non-linearly with dose, plateauing at higher dose levels, suggesting relative lower retention when doses were higher. This was not due to lower Se absorption in the intestine at higher Se doses, as Se fecal elimination factors (FEFs) in both female and male mice were stable across SeMet doses. However, higher Se doses caused increased urinary excretion, as evidenced by urinary Se excretion factors (UEFs) rising with escalating SeMet doses, leading to lower increases in Se tissue retention under higher Se doses. Further comparing Se tissue retention from rice and celery with same concentrations and species, celery consumption resulted in significantly lower Se accumulation and higher FEFs, suggesting lower Se bioavailability for celery than rice. Our findings demonstrate that Se dose and food matrix collectively shape Se bioavailability, offering a framework to address global deficiencies while balancing efficacy and safety.

IntroductionThe Malvaceae family shows rich species diversity and has substantial economic and medicinal value. However, the frequent interspecific hybridization among members of this family has resulted in confused phylogenetic relationships among the groups, limiting the usefulness of traditional classification methods.MethodsThis study aimed to investigate the phylogenetic relationships among selected taxa of Malvaceae by evaluating 23 chloroplast (CP) genomes, including three newly assembled CP genomes. Among these three genomes, the CP genome of Hibiscus schizopetalus L. was reported for the first time, while the CP genomes of Alcea rosea L. and Hibiscus grewiifolius L., which have been deposited in NCBI, were re-analyzed here alongside newly generated data for comparative purposes. In addition, 20 downloaded CP genomes encompassing 13 genera were analyzed using SNPs in whole CP genomes data.ResultsThe results showed that the genomes ranged from 160,403 to 161,978 base pairs in length and consisted of small single copies (SSCs) and large single copies (LSCs) separated by two inverted repeat sequences (IRs), forming a typical quadripartite circular structure. The entire genome sequence showed relative conservation across species in terms of structure, GC content, codon usage, and gene composition. The mutation sites were mainly located in the LSC and SSC regions, and the variability in the non-coding regions was higher than that in the coding regions. The nucleotide polymorphism (Pi) analysis identified the non-coding regions such as ndhF-rpl32 and psbZ-trnG as high variable hotspots. A maximum likelihood phylogenetic tree was constructed based on SNPs in whole CP genomes data. The phylogenetic analysis divided these 23 species into five highly supported clades. It also revealed a close sister-group relationship between Abelmoschus and Hibiscus species, suggesting that Hibiscus may have a separate lineage from okra species. DiscussionIn conclusion, the increasing availability of CP genome resources will enhance our understanding of the classification and evolutionary patterns of the Malvaceae family. The development of molecular markers will provide important molecular evidence for precise identification and classification revision of plants in this family.

Soil microorganisms play a key role in plant adaptation to salt stress. However, how wood vinegar-modified biochars from different feedstocks regulate microbial communities and crop growth in saline soils remains unclear. In this study, biochars and wood vinegar-modified biochars prepared from maize straw and waste wood were applied at a rate of 1% (w/w) in a pot experiment. Wood vinegar modification enhanced the soil amelioration potential of biochar, but its effects were strongly feedstock-dependent. Wood vinegar-modified maize straw biochar (VMB) showed the most consistent growth-promoting effect, significantly increasing soybean biomass and nutrient uptake. Compared with maize straw biochar (MB), VMB improved soybean growth by reducing the plant Na+/K+ ratio, promoting root development, enhancing microbial P turnover, increasing labile P pools, reshaping the rhizosphere microbial community, and improving microbial network stability. In contrast, wood vinegar-modified wood biochar (VWB) showed weaker and less consistent effects than wood biochar (WB). Compared with pristine biochars, wood vinegar-modified biochars had lower pH, higher dissolved organic carbon (DOC) content and cation exchange capacity, and greater abundances of phenolic hydroxyl and carboxyl groups, thereby creating a more favorable soil environment for microorganisms and plants. Maize straw-derived biochar had lower pH and EC and higher specific surface area, DOC, total N, and available P than wood-derived biochar, which may explain its stronger response to modification. Overall, VMB promoted soybean growth by improving soil P availability and microbial network stability, providing a theoretical basis for feedstock selection and sustainable utilization of saline soils.

Fatty acyl-CoA reductase (FAR) proteins play a crucial role in plant growth and development. Previous studies on FAR proteins primarily focused on their involvement in the synthesis of suberin and waxes, while limited attention has been given to their roles in biotic and abiotic stress responses. In this study, 31 FAR genes were identified across Gossypium arboreum, G. raimondii, G. hirsutum, and G. barbadense. Bioinformatics analyses revealed that cotton FAR genes cluster into three distinct groups, with conserved motifs, exon-coding sequences, and domains among genes within the same group. Promoter analysis identified numerous cis-acting elements associated with plant hormone regulation in GhFAR genes, highlighting their regulatory potential. Gene family expansion was predominantly driven by whole-genome duplication events. Expression profiling demonstrated that FAR genes are expressed in various cotton tissues, with most showing reduced expression under abiotic stresses but significantly increased expression following Verticillium dahliae infection. Functional analysis using virus-induced gene silencing (VIGS) of GhFAR3 revealed that its silencing compromises disease resistance by reducing suberin accumulation in roots. This study provides a comprehensive genome-wide identification and characterization of FAR genes in cotton, emphasizing their potential biological functions. Furthermore, it establishes GhFAR3 as a key player in enhancing cotton resistance to V. dahliae, mediated through suberin biosynthesis in roots.

Cysteine metabolism is essential for plants to alleviate cadmium (Cd2+) stress. Investigating the function of serine acetyltransferase (SAT), the pivotal enzyme in cysteine synthesis, in combating Cd2+ stress is highly significant. This study conducted a bioinformatics analysis of the SAT gene family and identified key candidate genes, GhSERAT1;1 and GhSERAT1;2, that respond to Cd2+ stress. Plants subjected to gene silencing of GhSERAT1;1 and GhSERAT1;2 through virus-induced gene silencing exhibited a notable reduction in cysteine and glutathione levels, an increase in intracellular malondialdehyde content, and heightened sensitivity to Cd2+ stress. Compared with non-silenced plants, those with silenced genes displayed poorer growth conditions, decreased biomass, and more pronounced damage to chloroplast and leaf structures when exposed to Cd2+ stress. This study integrated the primary enzyme involved in cysteine synthesis with Cd2+ stress, elucidating the relationship between Cd2+ and cysteine. These findings significantly enhance our understanding of cysteine synthesis genes and contribute to developing Cd2+-resistant plant breeding strategies.