Heavy metal contamination of the environment is becoming alarmingly serious with the rapid pace of urbanization and unrestricted industrialization. Cobalt (Co) is one of the toxic and non-essential metals for plants; however, it is widely recognized as a beneficial component for numerous enzymes and coenzymes within biological systems. Nevertheless, if its level increases, it leads to severe toxicity and plants are highly susceptible to the toxic effects of elevated cobalt concentration. Herein, we investigated the effect of different levels of Co stress on maize seedlings using physiological, biochemical, ultrastructural, and molecular markers in order to depict a panoramic view of the maize response mechanism. The findings revealed that Co stress inhibited plant growth and development by causing oxidative stress, reduced nutrient uptake, and photosynthetic efficiency. The increasing concentration of Co led to the overproduction of reactive oxygen species (ROS) in root and shoot that principally caused oxidative damage as monitored by high MDA contents and ultrastructural damages. The in vivo detection of ROS stained with ROS fluorescent (H2DCFDA and DHE) further confirmed the overproduction of ROS under stress conditions. In response to stress, a substantial upward trend was noticed in plant antioxidant defense activities with the increasing Co concentration. However, a sharp decline was observed in antioxidant enzymes under the highest used concentration, while the level of ROS continued to rise, indicating a disruption in cellular redox homeostasis. Furthermore, the qPCR results showed that Co stress differentially expressed antioxidant-related genes. Collectively, we provide the first report tracking down the underlying mechanism of Co stress-induced oxidative damage in maize.

Global crises of biodiversity loss and climate change highlighted the urgent need to understand the trade-off and synergy between carbon sequestration and biodiversity and the driving mechanisms behind them. However, existing research on driving mechanisms primarily focused on the influence of independent variables on dependent variables, neglecting the interaction between independent variables. The InVEST and PLUS models were utilized in this study to evaluate the dynamics of carbon sequestration and biodiversity in the Chishui River Basin, Southwest of China. By integrating the factor detection of the Geographic Detector and the Multi-scale Geographically Weighted Regression (MGWR) model, this study identified the key drivers affecting trade-off and synergy and revealed their spatial heterogeneity. Furthermore, utilizing the interaction detection of the Geographic Detector and the Geographic Cross-Convergence Mapping (GCCM) model, this study analyzed the interaction direction and interaction intensity among key drivers. The results showed that: (1) From 2012-2035, the total values of carbon sequestration and biodiversity exhibited an overall increasing trend, with 58.49 % of the area showing trade-off effects, 23.86 % showing positive synergy, and 17.65 % showing negative synergy. (2) Elevation, slope, topographic relief, temperature, and land use intensity were identified as key factors influencing trade-off and synergy, and these factors displayed significant spatial heterogeneity. (3) When multiple drivers interacted, their explanatory power for trade-off and synergy increased significantly. The interaction patterns among drivers presented a "topography-*climate <-> human activity <-> distance-*soil" structure, where human activity interacted bidirectionally with other factors, with the strongest interaction occurring between climate factors and human activity. This study advanced the understanding of the driving mechanisms behind trade-off and synergy through MWGR and GCCM model. Based on the supplydemand theory, a synergistic management framework was proposed to emphasize the "patternprocess-service-feedback" nexus. The hotspot areas identified through scenario simulation could be used for synergistic enhancement of carbon sequestration and biodiversity.

The mechanism underlying the ability of rice to germinate underwater is a largely enigmatic but key research question highly relevant to rice cultivation. Moreover, although rice is known to accumulate salicylic acid (SA), SA biosynthesis is poorly defined, and its role in underwater germination is unknown. It is also unclear whether peroxisomes, organelles essential to oilseed germination and rice SA accumulation, play a role in rice germination. Here, we show that submerged imbibition of rice seeds induces SA accumulation to promote germination in submergence. Two submergence-induced peroxisomal Oryza sativa cinnamate:CoA ligases (OsCNLs) are required for this SA accumulation. SA exerts this germination-promoting function by inducing indole-acetic acid (IAA) catabolism through the IAA-amino acid conjugating enzyme GH3. The metabolic cascade we identified may potentially be adopted in agriculture to improve the underwater germination of submergence-intolerant rice varieties. SA pretreatment is also a promising strategy to improve submerged rice germination in the field.

Background The growth and development of rice ( Oryza sativa L.) are affected by multiple factors, such as ROS homeostasis and utilization of iron. Here, we demonstrate that OsUGE2 , a gene encoding a UDP-glucose 4-epimerase, controls growth and development by regulating reactive oxygen species (ROS) and iron (Fe) level in rice. Knockout of this gene resulted in impaired growth, such as dwarf phenotype, weakened root growth and pale yellow leaves. Biochemical analysis showed that loss of function of OsUGE2 significantly altered the proportion and content of UDP-Glucose (UDP-Glc) and UDP-Galactose (UDP-Gal). Cellular observation indicates that the impaired growth may result from decreased cell length. More importantly, RNA-sequencing analysis showed that knockout of OsUGE2 significantly influenced the expression of genes related to oxidoreductase process and iron ion homeostasis. Consistently, the content of ROS and Fe are significantly decreased in OsUGE2 knockout mutant. Furthermore, knockout mutants of OsUGE2 are insensitive to both Fe deficiency and hydrogen peroxide (H 2 O 2 ) treatment, which further confirmed that OsUGE2 control rice growth possibly through Fe and H 2 O 2 signal. Collectively, these results reveal a new pathway that OsUGE2 could affect growth and development via influencing ROS homeostasis and Fe level in rice.

Rural tourism plays a crucial role in promoting the process of urban-rural integration and regionally co-ordinated development.Drawing on theories of tourism system drive and system science,this study constructed a rural tourism system dynamics model with four subsystems:demand,supply,media,and support.Subsequently,we predicted the optimal mechanism for rural tourism development using scenario simulation methods.The find-ings were four-fold.(1)The rural tourism development dynamics system constructed in this study overcame the limitations of previous subsystem divisions.(2)Under the development scenarios of Natural development mecha-nism(NDM),Demand driven mechanism(DDM),Supply driven mechanism(SDMe),Media driven mechanism(MDM),Support driven mechanism(SDMu),and Synergistic driven mechanism(SDMy),the rural tourism devel-opment index values in 2035 were 0.678,0.702,0.755,0.715,0.776,and 0.836,respectively.Among these sce-narios,SDMy emerged as the ideal mechanism for rural tourism development in Yangshuo County.(3)Based on the characteristics of the rural tourism development index,rural tourism in Yangshuo County has undergone three stages:fluctuating growth,rapid development,and recession.(4)Yangshuo County's rural tourism supply index grows slowly and is always below 0.8,which is a key link for future structural optimization.This study proposes a direction for rural tourism development in Yangshuo and a later impetus,which can accelerate the process of ur-ban-rural integration in Yangshuo and similar areas.

The neurotoxin β-methylamino-L-alanine (BMAA) produced by cyanobacteria is widely present in foods and dietary supplements, posing a significant threat to human health. Ganglioside GM1 (GM1) has demonstrated potential for treating neurodegenerative diseases; however, its ability to prevent BMAA-induced neurotoxicity remains uncertain. In this study, zebrafish embryos were treated with Ganglioside GM1 to investigate its neuroprotective effects against BMAA exposure and the underlying mechanisms. Exposure to BMAA (400 μM) resulted in increased malformation rate and heart rates in zebrafish embryos at 72 h post-fertilization, along with the decreased survival rates. Conversely, GM1 intervention rescued BMAA-induced movement disorders and brain cell apoptosis, and oxidative stress was alleviated. In addition, GM1 inhibited the neurotoxic effects of BMAA in zebrafish embryos, as indicated by the up-regulation of genes related to neuron development (gpx1a, bdnf, ngfb, and islet-1) and the down-regulations of neurodegeneration-related genes (cdk5, gfap, and nptxr). GM1 treatment restored 261 differentially expressed genes (DEGs) identified through RNA sequencing, with the most enriched DEGs related to the mitogen-activated protein kinase (MAPK) signaling pathway (P < 0.05, 47 genes). GM1 modulated MAPK-targeted gene expression at the mRNA level. These findings suggest that GM1 alleviates BMAA-induced neurotoxicity in the early-life stage of zebrafish embryos. The neuroprotective mechanism may involve the MAPK pathway, offering new insights into lipid signaling for the prevention of neurotoxic hazards to biological health.

Alternative splicing serves as a pivotal source of complexity in the transcriptome and proteome, selectively connecting various coding elements to generate a diverse array of mRNAs. This process encodes multiple proteins with either similar or distinct functions, contributing significantly to the intricacies of cellular processes. The role of alternative splicing in mammalian immunity has been well studied. Remarkably, the immune system of fish shares substantial similarities with that of humans, and alternative splicing also emerges as a key player in the immune processes of fish. In this review, we offer an overview of alternative splicing and its associated functions in the immune processes of fish, and summarize the research progress on alternative splicing in the fish immunity. Furthermore, we review the impact of alternative splicing on the fish immune system's response to external stimuli. Finally, we present our perspectives on future directions in this field. Our aim is to provide valuable insights for the future investigations into the role of alternative splicing in immunity.

The Roquin family is a recognized RNA-binding protein family that plays vital roles in regulating the expression of pro-inflammatory target gene mRNA during the immune process in mammals. However, the evolutionary status of the Roquin family across metazoans remains elusive, and limited studies are found in fish species. In this study, we discovered that the RC3H genes underwent a single round of gene duplication from a primitive ancestor during evolution from invertebrates to vertebrates. Furthermore, there were instances of species-specific gene loss events or teleost lineage-specific gene duplications throughout evolution. Domain/motif organization and selective pressure analysis revealed that Roquins exhibit high homology both within members of the family within the same species and across species. The three rc3h genes in zebrafish displayed similar expression patterns in early embryos and adult tissues, with rc3h1b showing the most prominent expression among them. Additionally, the promoter regions of the zebrafish rc3h genes contained numerous transcription factor binding sites similar to those of mammalian homologs. Moreover, the interaction protein network of Roquin and the potential binding motif in the 3’-UTR of putative target genes analysis both indicated that Roquins have the potential to degrade target mRNA through mechanisms similar to those of mammalian homologs. These findings shed light on the evolutionary history of Roquin among metazoans and hypothesized their role in the immune systems of zebrafish.

Root hairs are crucial in the uptake of essential nutrients and water in plants. This study showed that a zinc finger protein, GIS3 is involved in root hair growth in Arabidopsis. The loss-of-function gis3 and GIS3 RNAi transgenic line exhibited a significant reduction in root hairs compared to the wild type. The application of 1-aminocyclopropane-1-carboxylic acid (ACC), an exogenous ethylene precursor, and 6-benzyl amino purine (BA), a synthetic cytokinin, significantly restored the percentage of hair cells in the epidermis in gis3 and induced GIS3 expression in the wild type. More importantly, molecular and genetic studies revealed that GIS3 acts upstream of ROOT HAIR DEFECTIVE 2 (RHD2) and RHD4 by binding to their promoters. Furthermore, exogenous ACC and BA application significantly induced the expression of RHD2 and RHD4, while root hair phenotype of rhd2-1, rhd4-1, and rhd4-3 was insensitive to ACC and BA treatment. We can therefore conclude that GIS3 modulates root hair development by directly regulating RHD2 and RHD4 expression through ethylene and cytokinin signals in Arabidopsis.

It is widely known that heavy metals (HM) severely impair plant growth and development, posing a serious threat to food security and human health. Anthropogenic activities and an increasing trend toward industrialization combined with a rising population are further exacerbating environmental conditions, posing a massive global challenge to sustainable agriculture. Cobalt (Co) is one of the toxic and nonessential HM and has become a major pollutant affecting global food production and quality. High Co concentrations negatively affect plant growth and development by causing oxidative damage, nutrient deficiencies, and impairment of the photosynthetic apparatus. In response, plants show a range of changes in metabolism and growth that help them avoid stress-induced changes. Recent advances in “omics” technology (transcriptomics, metabolomics, proteomics, etc.) have helped track down transcription factors, metabolites, and stress-triggered proteins that play a role in stress tolerance and that can be used for metal tolerance or the development of resilient plant systems. Advances in omics have generated extensive resources for genomics, proteomics, transcriptomics, and metabolomics of plants that respond flexibly to provocations triggered by heavy metals. This chapter will brief Co induce changes and insights into omics approaches to understand Co-stress tolerance.

Nitrate is an important nitrogen source and signaling molecule that regulates plant growth and development. Although several components of the nitrate signaling pathway have been identified, the detailed mechanisms are still unclear. Our previous results showed that OsMADS25 can regulate root development in response to nitrate signals, but the mechanism is still unknown. Here, we try to answer two key questions: how does OsMADS25 move from the cytoplasm to the nucleus, and what are the direct target genes activated by OsMADS25 to regulate root growth after it moves to the nucleus in response to nitrate? Our results demonstrated that OsMADS25 moves from the cytoplasm to the nucleus in the presence of nitrate in an OsNAR2.1-dependent manner. Chromatin immunoprecipitation sequencing, chromatin immunoprecipitation qPCR, yeast one-hybrid, and luciferase experiments showed that OsMADS25 directly activates the expression of OsMADS27 and OsARF7, which are reported to be associated with root growth. Finally, OsMADS25-RNAi lines, the Osnar2.1 mutant, and OsMADS25-RNAi Osnar2.1 lines exhibited significantly reduced root growth compared with the wild type in response to nitrate supply, and expression of OsMADS27 and OsARF7 was significantly suppressed in these lines. Collectively, these results reveal a new mechanism by which OsMADS25 interacts with OsNAR2.1. This interaction is required for nuclear accumulation of OsMADS25, which promotes OsMADS27 and OsARF7 expression and root growth in a nitrate dependent manner.

Trichomes are common appendages originating and projecting from the epidermal cell layer of most terrestrial plants. They act as a first line of defense and protect plants against different types of adverse environmental factors. GL3/EGL3-GL1-TTG1 transcriptional activator complex and GIS family genes regulate trichome initiation through gibberellin (GA) signaling in Arabidopsis. Here, our novel findings show that TOE1/TOE2, which are involved in developmental timing, control the initiation of the main-stem inflorescence trichome in Arabidopsis. Phenotype analysis showed that the 35S:TOE1 transgenic line increases trichome density of the main-stem inflorescence in Arabidopsis, while 35S:miR172b, toe1, toe2 and toe1toe2 have the opposite phenotypes. Quantitative RT-PCR results showed that TOE1/TOE2 positively regulate the expression of GL3 and GL1. In addition, protein-protein interaction analysis experiments further demonstrated that TOE1/TOE2 interacting with GIS/GIS2/ZFP8 regulate trichome initiation in Arabidopsis. Furthermore, phenotype and expression analysis also demonstrated that TOE1 is involved in GA signaling to control trichome initiation in Arabidopsis. Taken together, our results suggest that TOE1/TOE2 interact with GIS to control trichome development in Arabidopsis. This report could provide valuable information for further study of the interaction of TOE1/TOE2 with GIS in controlling trichome development in plants.

Heavy metal (HM) contamination and high environmental temperature (HT) are caused by anthropogenic ac-tivities that negatively impact soil microbial communities and agricultural productivity. Although HM con-taminations have deleterious effects on microbes and plants; there are hardly any reports on the combined effects of HM and HT. Here, we reported that HT coupled with cadmium (Cd) accumulation in soil and irrigated water could seriously affect crop growth and productivity, alternatively influencing the microbial community and nutrient cycles of paddy soils in rice fields. We analyzed different mechanisms of plants and microflora in the rhizospheric region, such as plant rhizospheric nitrification, endophytes colonization, nutrient uptake, and physiology of temperature-sensitive (IR64) and temperature-resistant Huanghuazhan (HZ) rice cultivars against different Cd levels (2, 5 and 10 mg kg-1) with rice plants grown under 25 degrees C and 40 degrees C temperatures. Conse-quently, an increment in Cd accumulation was observed with rising temperature leading to enhanced expression of OsNTRs. In contrast, a greater decline in the microbial community was detected in IR64 cultivar than HZ. Similarly, ammonium oxidation, root-IAA, shoot-ABA production, and 16S rRNA gene abundance in the rhizo-sphere and endosphere were significantly influenced by HT and Cd levels, resulting in a significant decrease in the colonization of endophytes and the surface area of roots, leading to a decreased N uptake from the soil. Overall, the outcomes of this study unveiled the novel effects of Cd, temperature, and their combined effect on rice growth and functions of the microbial community. These results provide effective strategies to overcome Cd-phytotoxicity on the health of endophytes and rhizospheric bacteria in Cd-contaminated soil by using temperature-tolerant rice cultivars.

Zc3h12 family is an important RNA-binding protein family regulating mRNA of inflammatory cytokines in mammals. However, there are few studies on their post-transcriptional level regulation of inflammatory cytokines in fish. Here, we investigated the evolution of zebrafish Zc3h12 family and explored their immunomodulatory role. Phylogenetic and syntenic analysis indicated the number of zc3h12 family members had increased ranging from a single member in invertebrates to a single copy of four members in mammals. As the most evolutionarily diverse group of vertebrates, the number of zc3h12 family members was more complex and diverse in the teleost, each member experienced different fates and followed different rules in multiple rounds of whole-genome duplication events. Thereinto, zebrafish contained three zc3h12 genes, among which zc3h12aa and zc3h12ab were duplicated from the same gene. Zebrafish Zc3h12 family could recognize the 3'-UTR regions of inflammatory cytokines through binding to the specific RNA secondary structure and negatively regulate their expression. Deletion of either Zc3h12 domains or mutation of the key amino acid in RNAase domain attenuated their modulatory effect, suggesting both domain and RNAase activity are important to the immunomodulatory role. These results elucidated the evolution of Zc3h12 family and uncovered Zc3h12-mediated post-transcriptional regulation of cytokines in zebrafish.

Reactive oxygen species (ROS) are highly reactive molecules, generated by nicotinamide adenine dinucleotide phosphate oxidases encoded by respiratory burst oxidase homologs. The functions of the OsRbohs gene family in rice are diverse and poorly understood. OsRbohI was recently identified as a newly evolved gene in the rice OsRbohs gene family. However, the function of OsRbohI in regulating rice growth is not yet reported. In this study, our results indicate that knockout (KO) OsRbohI mutants showed significantly shorter shoot and primary roots, along with lower ROS content than the control lines, whereas the overexpression (OE) lines displayed contrasting results. Further experiments showed that the abnormal length of the shoot and root is mainly caused by altered cell size. These results indicate that OsRbohI regulates rice shoot and root growth through the ROS signal. More importantly, RNA-seq analysis and jasmonic acid (JA) treatment demonstrated that OsRbohI regulates rice growth via the JA synthesis and signaling pathways. Compared with the control, the results showed that the KO mutants were more sensitive to JA, whereas the OE lines were less sensitive to JA. Collectively, our results reveal a novel pathway in which OsRbohI regulates rice growth and development by affecting their ROS homeostasis through JA synthesis and signaling pathway.

KEY MESSAGE:Root hairs are required for water and nutrient acquisition in plants. Here, we report a novel mechanism that OsUGE1 is negatively controlled by OsGRF6 to regulate root hair elongation in rice. Root hairs are tubular outgrowths generated by the root epidermal cells. They effectively enlarge the soil-root contact area and play essential roles for nutrient and water absorption. Here, in this study, we demonstrated that the Oryza sativa UDP-glucose 4-epimerase 1-like (OsUGE1) negatively regulated root hair elongation and was directly targeted by Oryza sativa growth regulating factor 6 (OsGRF6). Knockout mutants of OsUGE1 using CRISPR-Cas9 technology showed longer root hairs than those of wild type. In contrast, overexpression lines of OsUGE1 displayed shorter root hair compared with those of wild type. GUS staining showed that it could specifically express in root hair. Subcellular localization analysis indicates that OsUGE1 is located in endoplasmic reticulum, nucleus and plasma membrane. More importantly, ChIP-qPCR, Yeast-one-hybrid and BiFC experiments revealed that OsGRF6 could bind to the promoter of OsUGE1. Furthermore, knockout mutants of OsGRF6 showed shorter root hair than those of wild type, and OsGRF6 dominantly expressed in root. In addition, the expression level of OsUGE1 is significantly downregulated in Osgrf6 mutant. Taken together, our study reveals a novel pathway that OsUGE1 is negatively controlled by OsGRF6 to regulate root hair elongation in rice.

CD248, also known as endosialin or tumor endothelial marker 1, is a type I single transmembrane glycoprotein. CD248 has been demonstrated to be upregulated in cancers, tumors and many fibrotic diseases in human and mice, such as liver damage, pulmonary fibrosis, renal fibrosis, arthritis and tumor neovascularization. However, no definite CD248 orthologs in fish have been documented so far. In this study, we report the identification of cd248a and cd248b in the zebrafish. Both the phylogenetic analysis and the conserved synteny strongly suggested that zebrafish cd248a and cd248b are orthologs of the human CD248. Both cd248a and cd248b exhibited similar and dynamic expression pattern in early development, both genes had weak maternal expression, the zygotic transcripts were first seen in anterior somites and head mesenchyme, then shifted to eyes and head mesenchyme, later expanded to branchial arches, and gradually declined with development. The expression profiles of cd248a and cd248b were upregulated upon LPS (Lipopolysaccharide) challenge. Both Cd248a protein and Cd248b protein were localized on the cell membrane and cytoplasm, and overexpression of cd248a and cd248b induced the expression of pro-inflammatory cytokines, in vitro and in vivo. Moreover, deficiency of cd248a or cd248b both downregulated the expression of pro-inflammatory cytokines and upregulated anti-inflammatory cytokine. Additionally, loss of cd248a or cd248b both downregulated the expression of pro-inflammatory cytokines after LPS treatment. Taken together, these results indicated that cd248a and cd248b in zebrafish were involved in immune response and would provide further information to understand functions of Cd248 protein in innate immunity of fish.

Many studies have demonstrated that receptor interacting protein kinase-1 acts as a crucial mediator in the regulation of immune response, but evidence remains lacking for its direct interaction with bacteria. In this study, we found that challenge with lipopolysaccharide and lipoteichoic acid resulted in a significantly increased transcriptional expression of receptor interacting protein kinase-1 in zebrafish, suggesting the receptor interacting protein kinase-1 is implicated in anti-infectious responses. In accordance, we found that recombinant receptor interacting protein kinase-1 was not only able to bind to Gram-negative and -positive bacteria via interaction with lipopolysaccharide and lipoteichoic acid, but also agglutinate both Gram-negative and -positive bacteria in a Ca2+-dependent manner.

The pollution of nanoparticles (NPs) has linked with severe negative effects on crop productivity. Thus, effective strategies are needed to mitigate the phytotoxicity of NPs. The aim of present study was to evaluate the efficacy of exogenously applied melatonin (MT) in mitigating the toxic effects of copper oxide nanoparticles (CuO NPs) from maize seedlings. Therefore, we comprehensively investigated the inhibitory effects of MT against CuO NPs-induced toxicity on morpho-physiological, biochemical and ultrastructural levels in maize. Our results show that CuO NPs (300 mg L-1) exposure displayed significantly reduction in all plant growth traits and induced toxicity in maize. Furthermore, 50 μM MT provided maximum plant tolerance against CuO NPs-induced phytotoxicity. It was noticed that MT improved plant growth, biomass, photochemical efficiency (Fv/Fm), chlorophyll contents (Chl a and Chl b), SPAD values and gas exchange attributes (stomatal conductance, net photosynthetic rate, intercellular CO2 concentration and transpiration rate) under CuO NPs stress. In addition, MT enhanced the antioxidant defense system and conferred protection to ultrastructural (mainly chloroplast, thylakoids membrane and plastoglobuli) damages and stomatal closure in maize plants subjected to CuO NPs stress. Together, it can be stated that the exogenous supply of MT improves the resilience of maize plants against the CuO NPs-induced phytotoxicity. Our current findings can be useful for the enhancement of plant growth and yield attributes in CuO NPs-contaminated soils. The reported information can provide insight into the MT pathways that can be used to improve crop stress tolerance in a challenging environment.

The enormous use of metal-based nanoparticles (NPs) in different sectors may result in enhanced accumulation in agricultural soil, which could impose negative effects on crop productivity. Hence, strategies are needed to explore the mechanisms of copper oxide nanoparticle (CuO NP)-induced toxicity in crops. The present study aimed to investigate the involvement of ethylene in CuO NP-induced toxicity in rice seedlings. Here, our results indicate that 450 mg L −1 of CuO NPs induced toxic effects in rice seedlings. Thus, it was evidenced by the reduced plant biomass accumulation, enhanced oxidative stress indicators, and cellular ultrastructural damages. More importantly, the exogenous supply of ethylene biosynthesis and signaling antagonists cobalt (Co) and silver (Ag) respectively provided tolerance and improved the defense system of rice seedlings against CuO NP toxicity. The ethylene antagonists could significantly reduce the extent of ultrastructural and stomatal damage by controlling the ROS accumulation in rice seedlings under CuO NP stress. Furthermore, Co and Ag augmented the antioxidant defense system against CuO NP-induced toxicity. Contrary to that, all oxidative damage attributes were further enhanced exogenous application of ethylene biosynthesis precursor [1-aminocyclopropane-1-carboxylic acid (ACC)] in the presence of CuO NPs. In addition, ACC could increase the CuO NP-induced stomatal and ultrastructural damages by reducing the ROS-scavenging ability in rice seedlings. Taken together, these results indicate the involvement of ethylene in CuO NP-induced toxicity in rice seedlings.