Oxidase Homolog Research Articles

Calcium Chloride Enhances Defense Response Against Diaporthe nobilis by Eliciting Reactive Oxygen Species in Kiwifruit

Kiwifruit (Actinidia spp.) rot, caused by pathogenic fungi, represents the most severe disease during postharvest storage. The utilization of induced resistance against fungal pathogens in harvested fruit has been demonstrated as a promising strategy for controlling postharvest losses. In this study, the effect of calcium chloride (CaCl2) on induction of resistance to Diaporthe nobilis, one of the most epidemic pathogens of soft rot in kiwifruit, and the underlying mechanisms were investigated. Our data showed that the application of 3% CaCl2 resulted in smaller rotten lesion diameters after inoculation with the pathogen, induced expression of the respiratory burst oxidase homolog (RBOH) gene family, and increased accumulation of reactive oxygen species (ROS). Moreover, we found that the activity of three pathogenesis‐related antioxidant enzymes—peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD)—as well as the expression of important defense‐related genes, was enhanced by CaCl2 treatment. The results of our study suggest that the application of CaCl2 treatment holds promise as a green and viable method for effectively preventing and safeguarding kiwifruit against postharvest decay.

Inhibition of the invasive plant Ambrosia trifida by Sigesbeckia glabrescens extracts.

Ambrosia trifida is an invasive weed that destroys the local ecological environment, and causes a reduction in population diversity and grassland decline. The evolution of herbicide resistance has also increased the difficulty of managing A. trifida, so interspecific plant competition based on allelopathy has been used as an effective and sustainable ecological alternative. However, how to control A. trifida through interspecific competition and the underlying mechanisms are unclear. Here, we found that extracts from both the roots and leaves of the medicinal plant Sigesbeckia glabrescens suppressed the growth of A. trifida by reducing the plant height and biomass. The decrease in biomass may be explained by disruption of carbon and nitrogen metabolism. These disruptions are due to a significant decrease in the expression of genes related to nitrate absorption and transport in roots and a significant decrease in the expression of key genes related to photosynthesis and carbon fixation. Consequently, genes involved in sucrose synthesis are downregulated. In addition, increases in H2O2 content and respiratory burst oxidase homologue (RbohD) gene expression suggested that A. trifida underwent oxidative stress caused by reactive oxygen species (ROS) bursts, resulting in apoptosis due to the significant upregulation of key genes associated with apoptotic pathways. Furthermore, we identified three main allelochemicals, coumarin, ferulic acid, and 5-aminolevulinic acid (5-ALA), in S. glabrescens extracts and revealed that the combination of these three compounds could suppress the growth of A. trifida seedlings. The phenotypes and transcriptome profiles of the seedlings treated with these chemicals were the same as those of the seedlings treated with the S. glabrescens extracts. Taken together, the results of this study revealed the mechanism underlying the toxic effects of S. glabrescens on A. trifida, providing a theoretical basis for the use of interspecific plant competition for invasive weed control and further application of S. glabrescens allelochemicals in weed management.

Root apoplastic barrier mechanism: an adaptive strategy to protect against salt stress.

From soil to plant, the water and ions, enter the root system through the symplast and apoplast pathways. The latter gains significance under salt stress and becomes a major port of entry of the dissolved salts particularly the sodium ions into the root vasculature. The casparian strip (CS), a lignified barrier circumambulating the root endodermal cells' radial and transverse walls regulates the movement of water and solutes in and out of the stele. The development of CS begins with the synthesis of a protein scaffold made of CASPARIAN STRIP MEMBRANE DOMAIN PROTEINs (CASPs), followed by lignin deposition involving the enzymatic machinery viz., ENHANCED SUBERIN 1 (ESB1), RESPIRATORY BURST OXIDASE HOMOLOG F (RBOHF), and PEROXIDASE 64 (PER64), etc. Towards maintaining the integrity of the CS, the CASPARIAN STRIP INTEGRITY FACTOR 1/2-SCHENGEN 3-SCHENGEN 1 (CIF1/2-SGN3-SGN1) signaling pathway has been found to play a significant role as a barrier surveillance system, the resultant is compensatory lignification of the radial and stele-facing transversal walls of endodermis. This leads to the formation of 'U' shaped lignified structures that enable an effective apoplastic barrier mechanism to prevent the influx of sodium ions into the stele. Rice, the major staple crop is generally classified as salt-susceptible, however, root cross-sectional anatomy of selected salt-tolerant genotypes exhibits an early and enhanced lignification of the endodermis. For instance, in the salt-tolerant landrace Mundan, the development of CS is accompanied by the formation of continuous 'U' shaped lignified structures along the endodermal walls under salt stress.

SnRK1α1-mediated RBOH1 phosphorylation regulates reactive oxygen species to enhance tolerance to low nitrogen in tomato.

Nitrogen is essential for plant growth and development. SNF1-related protein kinase 1 (SnRK1) is an evolutionarily conserved protein kinase pivotal for regulating plant responses to nutrient deficiency. Here, we discovered that the expression and activity of the SnRK1 α-catalytic subunit (SnRK1α1) increased in response to low-nitrogen stress. SnRK1α1 overexpression enhanced seedling tolerance, nitrate uptake capacity, apoplastic reactive oxygen species (ROS) accumulation, and NADPH oxidase activity in tomato (Solanum lycopersicum L.) under low-nitrogen stress compared to wild type plants, while snrk1α1 mutants exhibited the opposite phenotypes. Mutation of the NADPH oxidase gene Respiratory burst oxidase homolog 1 (RBOH1) suppressed numerous nitrate uptake and metabolism genes during low-nitrogen stress. rboh1 mutants displayed lower NADPH oxidase activity, apoplastic ROS production, and seedling tolerance to low nitrogen. Silencing RBOH1 expression also compromised SnRK1α1-mediated seedling tolerance to low-nitrogen stress. SnRK1α1 interacts with and activates RBOH1 through phosphorylation of three N-terminal serine residues, leading to increased apoplastic ROS production and enhanced tolerance to low nitrogen conditions. Furthermore, RBOH1-dependent ROS oxidatively modified the transcription factor TGA4 at residue Cys-334, which increased NRT1.1 and NRT2.1 expression under low-nitrogen stress. These findings reveal a SnRK1α1-mediated signaling pathway and highlight the essential role of RBOH1-dependent ROS production in enhancing plant tolerance to low nitrogen.

Genome-Wide Identification, Classification, Expression Analysis, and Screening of Drought and Heat Resistance-Related Candidates of the Rboh Gene Family in Wheat.

Plant respiratory burst oxidase homologs (Rbohs) are key enzymes that produce reactive oxygen species (ROS), which serve as signaling molecules regulating plant growth and stress responses. In this study, 39 TaRboh genes (TaRboh01-TaRboh39) were identified. These genes were distributed unevenly among the wheat genome's fourteen chromosomes, with the exception of homoeologous group 2 and 7 and chromosomes 4A, as well as one unidentified linkage group (Un). TaRbohs were classified into ten distinct clades, each sharing similar motif compositions and gene structures. The promoter regions of TaRbohs contained cis-elements related to hormones, growth and development, and stresses. Furthermore, five TaRboh genes (TaRboh26, TaRboh27, TaRboh31, TaRboh32, and TaRboh34) exhibited strong evolutionary conservation. Additionally, a Ka/Ks analysis confirmed that purifying selection was the predominant force driving the evolution of these genes. Expression profiling and qPCR results further indicated differential expression patterns of TaRboh genes between heat and drought stresses. TaRboh11, TaRboh20, TaRboh22, TaRboh24, TaRboh29, and TaRboh34 were significantly upregulated under multiple stress conditions, whereas TaRboh30 was only elevated in response to drought stress. Collectively, our findings provide a systematic analysis of the wheat Rboh gene family and establish a theoretical framework for our future research on the role of Rboh genes in response to heat and drought stress.

Nitric oxide-mediated PpRAP2.2 regulates the transcription of PpRbohA/D to enhance resistance to Monilinia fructicola in peach fruit

Nitric oxide (NO), a critical signaling molecule, plays key roles in plant cell physiological processes, particularly in redox system. To investigate the resistance of exogenous NO to brown rot (Monilinia fructicola) in peach fruit during storage, postharvest peaches were treated with NO and c-PTIO (an NO scavenger), exploring the mechanism of NO enhancing disease resistance in peach fruit through reactive oxygen species (ROS) metabolism. The results indicated that NO significantly inhibited brown rot in postharvest peach fruit, evidenced by reduced disease incidence and smaller lesion diameters. Additionally, exogenous NO treatment decreased the expression of respiratory burst oxidase homologs (PpRbohs), which is a crucial factor in ROS metabolism. The transcription factor PpRAP2.2 was identified through yeast one-hybrid (Y1H) screening with the promoter of PpRbohD. The expression pattern of PpRAP2.2 was closely associated with superoxide anion (O2-) content and peach fruit disease resistance. PpRAP2.2, a nuclear-localized transcription factor, activated the expression of PpRbohA and PpRbohD by directly binding to their promoter. The virus-induced gene silencing (VIGS) assay showed that silencing of PpRAP2.2 facilitated peach disease resistance by reducing the transcription of PpRbohA and PpRbohD. Furthermore, the overexpression of PpRAP2.2 resulted in a significant reduction in disease incidence and conidiophores in transgenic tomatoes relative to control group under M. fructicola, suggesting that PpRAP2.2 effectively enhanced the resistance of tomatoes to M. fructicola. These results indicate that PpRAP2.2 enhances NO-mediated disease resistance in peach by regulating the expression of PpRbohA and PpRbohD. These finding provide a basis for further studies on function and regulatory mechanisms of peach fruit disease resistance.

The OXIDATIVE SIGNAL-INDUCIBLE1 kinase regulates plant immunity by linking microbial pattern-induced reactive oxygen species burst to MAP kinase activation.

Plant cell surface-localized pattern recognition receptors (PRRs) recognize microbial patterns and activate pattern-triggered immunity (PTI). Typical PTI responses include reactive oxygen species (ROS) burst controlled by the NADPH oxidase RESPIRATORY BURST OXIDASE HOMOLOG D (RbohD) and activation of the MAP kinase (MAPK) cascade composed of MAPKKK3/5-MKK4/5-MPK3/6. However, the mechanisms through which PRRs regulate and coordinate these immune responses are not fully understood. Here, we showed that Arabidopsis thaliana OXIDATIVE SIGNAL-INDUCIBLE1 (OXI1), a kinase known to be activated by ROS, is involved in the LYK5-CERK1 receptor complex, which recognizes fungal cell wall-derived chitin. The oxi1 mutant exhibits enhanced susceptibility to various pathogens and reduced chitin-induced MAPK activation and ROS burst. We showed that chitin induces the phosphorylation of OXI1 in an RbohD-dependent manner. H2O2 and chitin treatment causes the oxidation of OXI1 at Cys104 and Cys205, which is essential for the kinase activity of OXI1. These oxidation sites are required for chitin-induced MAPK activation and disease resistance. Activated OXI1 directly phosphorylates MAPKKK5 to regulate MAPK activation. Additionally, OXI1 phosphorylates RbohD, suggesting that it may activate RbohD to promote ROS burst to further enhance the long-term MAPK activation. Together, our findings reveal a pathway linking PRR-mediated ROS production to MAPK activation through OXI1.

Effect of elevated ammonium on biotic and abiotic stress defense responses and expression of related genes in cucumber (Cucumis sativus L.) plants

Ammonium (NH4+) enhances plant defense mechanisms but can be phytotoxic as the sole nitrogen source. To investigate the impact of a balanced NH4+ and NO3− ratio on plant defense parameters without adverse effects, cucumber plants (Cucumis sativus L.) were grown under control (14 mM NO3− + 2 mM NH4+) and elevated level of NH4+ (eNH4+, 8 mM NO3−+ 8 mM NH4+). Plants subjected to eNH4+ showed significantly increased shoot and root biomass by about 41% and 47%, respectively. Among the antioxidant enzymes studied, ascorbate peroxidase (EC 1.11.1.11) activity was increased up to 3.3 fold in eNH4+ compared with control plants, which was associated with enhanced resistance to paraquat. Upregulation of PATHOGENESIS RELATED PROTEIN 4 (PR4) and LIPOXYGENASE 1 (LOX1), accompanied by increased concentrations of salicylic acid and nitric oxide, conferred more excellent resistance of eNH4+ plants to powdery mildew infection. However, the expression levels of ACC OXIDASE 1 (ACO1) and RESPIRATORY BURST OXIDASE HOMOLOGS B (RBOHB) were lower in eNH4+ plants, which was consistent with decreased NADPH oxidase activity and lower leaf H2O2 levels. The biosynthesis of phenolics was enhanced, whereas the activities of polymerizing enzymes and lignin deposition were reduced by half in eNH4+ plants. Besides, a significant effect on plant biomass under salt or drought stress has not been observed between control and eNH4+ plants. These results showed that different defense pathways are distinctively affected by eNH4+ treatment, and the NH4+ to NO3− ratio may play a role in fine-tuning the plant defense response.

The histone deacetylase RhHDA15 represses petal senescence by epigenetically regulating reactive oxygen species homeostasis in rose.

Epigenetic modifications play vital roles in many biological processes. Flower senescence involves epigenetic factors that influence the chromatin state and gene expression. However, the molecular mechanism underlying the role of histone deacetylation in regulating flower senescence has not been elucidated. Here, we demonstrate that histone deacetylation is involved in flower senescence by fine-tuning reactive oxygen species (ROS) homeostasis in rose (Rosa hybrida). Our data reveal that the histone lysine deacetyltransferase RhHDA15 inhibits ROS accumulation and petal senescence by downregulating the expression of NADPH OXIDASE/RESPIRATORY BURST OXIDASE HOMOLOG (RhRboh) genes. Furthermore, the transcription factor RELATED TO ABI3/VP1 2 (RhRAV2) recruits RhHDA15 and the co-repressor TOPLESS (RhTPL) to suppress flower senescence by reducing H3 lysine 9 acetylation (H3K9ac) at the RhRbohA1/2 promoter and thus directly inhibiting precocious RhRbohA1/2 expression. Our work sheds light on an epigenetic mechanism in which histone deacetylation plays a crucial role in controlling petal senescence by precisely fine-tuning ROS homeostasis, providing insights into the regulatory network of organ senescence.

Transcription factor PagWRKY33 regulates gibberellin signaling and immune receptor pathways in Populus.

Enhanced autoimmunity often leads to impaired plant growth and development, and the coordination of immunity and growth in Populus remains elusive. In this study, we have identified the transcription factors PagWRKY33a/b as key regulators of immune response and growth maintenance in Populus. The disruption of PagWRKY33a/b causes growth issues and autoimmunity while conferring resistance to anthracnose caused by Colletotrichum gloeosporioides. PagWRKY33a/b binds to the promoters of N requirement gene 1.1 (NRG1.1) and Gibberellic Acid-Stimulated in Arabidopsis (GASA14) during infection, activating their transcription. This process maintains disease resistance and engages in GA signaling to reduce growth costs from immune activation. The oxPagWRKY33a/nrg1.1 mutant results in reduced resistance to C. gloeosporioides. Further, PagWRKY33a/b is phosphorylated and activated by mitogen-activated protein kinase kinase 1, which inhibits respiratory burst oxidase homolog D (RBOHD) and respiratory burst oxidase homolog I (RBOHI) transcription, causing reactive oxygen species bursts in wrky33a/b double mutants. This leads to an upregulation of PagNRG1.1 in the absence of pathogens. However, the wrky33a/b/nrg1.1 and wrky33a/b/rbohd triple mutants show compromised defense responses, underscoring the complexity of WRKY33 regulation. Additionally, the stability of PagWRKY33 is modulated by Ring Finger Protein 5 (PagRNF5)-mediated ubiquitination, balancing plant immunity and growth. Together, our results provide key insights into the complex function of WRKY33 in Populus autoimmunity and its impact on growth and development.

Comparative transcriptome analysis of tobacco plants reveals differences in reactive oxygen species activated defenses following the infestations of non-viruliferous and TYLCCNV-viruliferous whitefly Mediterranean

ABSTRACT Tomato yellow leaf curl China virus (TYLCCNV) viruliferous Mediterranean (MED) whitefly infestation in tobacco plants elevated hydrogen peroxide (H2O2) levels more than non-viruliferous ones. To unravel the underlying mechanisms behind the phenomenon, comparative analyses of plant transcriptome and enzyme activities of reactive oxygen species (ROS) scavengers were conducted in this study. Compared to the control, a total of 4032 and 6853 differentially expressed genes (DEGs) were identified in the plants following the infestations of non-viruliferous and TYLCCNV-viruliferous MED, respectively. Gene ontology analysis of DEGs suggested substantial transcriptional reprogramming of diverse cellular processes, particularly in the biotic stress response pathways of tobacco plants. Respiratory burst oxidase homologs related genes and important kinases putatively involved in plant defense were differentially modulated in tobacco plants in response to the non-viruliferous and TYLCCNV-viruliferous whitefly MED infestation. The analysis reported here lays out a strong foundation for future investigations into the mechanisms of ROS-associated plant defense mediating the interactions between B. tabaci and begomoviruses.

A novel root hair mutant, srh1, affects root hair elongation and reactive oxygen species levels in wheat.

Root hairs are single-celled projections on root surfaces, critical for water and nutrient uptake. Here, we describe the first short root hair mutant in wheat (Triticum aestivum L.), identified in a mutagenized population and termed here short root hair 1 (srh1). While the srh1 mutant can initiate root hair bulges, lack of subsequent extension results in very short root hairs. Due to its semi-dominant nature, heterozygous lines displayed intermediate root hair lengths compared to wild-type. Bulked segregant analysis in a BC1F3 segregating population genotyped via exome capture sequencing localized the genetic control of this mutant to a region on the long arm of chromosome 3A. Via RNA sequencing and bioinformatic analysis, we identified two promising candidate genes. The first was a respiratory burst oxidase homolog (RBOH) encoding gene TaNOX3-A, orthologous to RBOH genes controlling root hair elongation in rice (OsNOX3) and maize (ZmRTH5), that carries a missense mutation in a conserved region of the predicted protein. RBOHs are membrane bound proteins that produce reactive oxygen species (ROS) which trigger cell wall extensibility, allowing subsequent root hair elongation. Notably, reduced ROS levels were observed in srh1 root hair bulges compared to wild-type. The second candidate was the calreticulin-3 encoding gene TaCRT3-A, located within the wider srh1 interval and whose expression was significantly downregulated in srh1 root tissues. The identification of a major effect gene controlling wheat root hair morphology provides an entry point for future optimization of root hair architecture best suited to future agricultural environments.

Orchestrating ROS regulation: coordinated post-translational modification switches in NADPH oxidases.

Reactive oxygen species (ROS) are among the most important signaling molecules, playing a significant role in plant growth, development, and responses to various environmental stresses. Respiratory burst oxidase homologs (RBOHs) are key enzymes in ROS production. Plants tightly regulate the activation and deactivation of RBOHs through various post-translational modifications (PTMs), including phosphorylation, ubiquitination, S-nitrosylation, and persulfidation. These PTMs fine-tune ROS production, ensuring normal plant growth and development while facilitating rapid responses to abiotic and biotic stresses. This review discusses the effects of different PTMs on RBOH function and their biological relevance. Additionally, we examine the evolutionary conservation of PTM sites and emphasize the complex interplay between multiple PTMs regulating RBOHs.

Functions and mechanisms of CDPKs in plant responses to abiotic stress

Calcium-dependent protein kinases (CDPKs/CPKs) are members of the Ca2+-sensitive Ser/Thr protein kinase family and play a crucial role in plant growth and development and responses to abiotic stress. CDPKs are capable of rapidly sensing changes in intracellular Ca2+ signals and recognizing and phosphorylating specific substrates, thereby transmitting and amplifying Ca2+ signal cascades downstream. They are involved in plant responses to stress conditions such as drought, saline-alkali stress, and injuries and regulate plant growth and development, gene expression, ion channel activity, and stomatal movement. The autophosphorylation of CDPKs can affect their activities and substrate specificity. CDPKs have the ability to bind to and phosphorylate multiple substrates. In addition to participating in respiratory burst oxidase homolog (RBOH), mitogen-activated protein kinase (MAPK), and plant hormone signaling pathways, CDPKs can also bind to 14-3-3 proteins, which enables the regulation of plant responses to stress and promotes plant growth and development. This paper summarized the research findings on the discovery, structure, classification, and roles of CDPKs in plant responses to stress and proposed the future research directions, aiming to provide the genetic resources and a theoretical basis for improving the stress tolerance of crops.

The leucine-rich repeat receptor kinase QSK1 regulates PRR-RBOHD complexes targeted by the bacterial effector HopF2Pto.

Plants detect pathogens using cell-surface pattern recognition receptors (PRRs) such as ELONGATION Factor-TU (EF-TU) RECEPTOR (EFR) and FLAGELLIN SENSING 2 (FLS2), which recognize bacterial EF-Tu and flagellin, respectively. These PRRs belong to the leucine-rich repeat receptor kinase (LRR-RK) family and activate the production of reactive oxygen species via the NADPH oxidase RESPIRATORY BURST OXIDASE HOMOLOG D (RBOHD). The PRR-RBOHD complex is tightly regulated to prevent unwarranted or exaggerated immune responses. However, certain pathogen effectors can subvert these regulatory mechanisms, thereby suppressing plant immunity. To elucidate the intricate dynamics of the PRR-RBOHD complex, we conducted a comparative coimmunoprecipitation analysis using EFR, FLS2, and RBOHD in Arabidopsis thaliana. We identified QIAN SHOU KINASE 1 (QSK1), an LRR-RK, as a PRR-RBOHD complex-associated protein. QSK1 downregulated FLS2 and EFR abundance, functioning as a negative regulator of PRR-triggered immunity (PTI). QSK1 was targeted by the bacterial effector HopF2Pto, a mono-ADP ribosyltransferase, reducing FLS2 and EFR levels through both transcriptional and transcription-independent pathways, thereby inhibiting PTI. Furthermore, HopF2Pto transcriptionally downregulated PROSCOOP genes encoding important stress-regulated phytocytokines and their receptor MALE DISCOVERER 1-INTERACTING RECEPTOR-LIKE KINASE 2. Importantly, HopF2Pto requires QSK1 for its accumulation and virulence functions within plants. In summary, our results provide insights into the mechanism by which HopF2Pto employs QSK1 to desensitize plants to pathogen attack.

3'UTR of tobacco vein mottling virus regulates downstream GFP expression and changes in host gene expression.

Tobacco vein mottling virus (TVMV) is a member of the family Potyviridae. The 3' untranslated region (3'UTR) of viral genomic RNA has been reported to significantly impact viral infection. Nevertheless, the role of the TVMV 3'UTR during viral infection remains unknown. Here, a 3'UTR-GFP expression vector was transiently expressed in Nicotiana benthamiana, in which the 3'UTR of TVMV was introduced upstream of the green fluorescent protein (GFP) gene. Transcriptome sequencing was performed to analyze the genes associated with plant resistance. The effect of the TVMV 3'UTR on GFP expression was studied using an Agrobacterium-mediated transient expression assay, revealing that the TVMV 3'UTR significantly inhibited GFP expression. Transcriptome analysis of differentially expressed genes in 3'UTR-GFP in N. benthamiana was performed to elucidate the why the TVMV 3'UTR inhibited GFP expression. Eighty genes related to plant disease resistance were differentially expressed, including 29 upregulated and 51 downregulated genes. Significantly upregulated genes included those encoding the calcium-binding protein CML24, leucine-rich repeat receptor-like tyrosine-protein kinase, and respiratory burst oxidase homolog protein E. The significantly downregulated genes included calcium-binding protein 7, ethylene-responsive transcription factor 10, endoglucanase 5, and receptor-like protein kinase. These findings indicate that the 3'UTR of TVMV may inhibit the expression of GFP gene by inducing the expression of plant resistance genes. This study provides a theoretical basis for further research on the function and mechanism of the TVMV 3'UTR.

Root-sourced H2O2 is essential for maintaining jasmonic acid and Na+/K+ homeostasis to delay leaf senescence during salt stress in Paspalum vaginatum

Improving salt tolerance and mitigating senescence in the presence of high salinity are crucial for sustaining agricultural productivity. Previous research has demonstrated that hydrogen peroxide (H2O2), specifically H2O2 derived from roots and mediated by the respiratory burst oxidase homolog (NADPH), plays a significant role in regulating ion and plant hormone homeostasis in glycophytic plants, such as Arabidopsis. However, the extent to which root-derived H2O2 fulfils similar functions in halophytic plants remains uncertain. Therefore, our study aimed to explore the potential contribution of root-sourced H2O2 in delaying leaf senescence induced by high salinity, utilizing seashore paspalum as a model halophytic plant. The application of the NADPH-oxidase inhibitor DPI, coupled with a series of leaf senescence analyses, we revealed that root-derived H2O2 significantly retards salt-induced leaf senescence. Furthermore, through the application of hormone analysis, lipidomics, ionomics, Non-invasive Micro-test Technology (NMT), and transcriptomics, we established that NADPH-dependent H2O2 induced by salt stress in the roots was indispensable for maintaining the balance of the aging hormone, jasmonic acid (JA), and sodium ion homeostasis within this halophytic plant. Finally, by utilizing AtrbohD Arabidopsis mutants and virus-induced gene silencing (VIGs) in Paspalum vaginatum, we demonstrated the pivotal role played by root-sourced H2O2 in upholding JA homeostasis and regulating JA-triggered leaf senescence in P. vaginatum. This study offers novel insights into the mechanisms that govern plant leaf senescence and its response to salinity-induced stress.

FmRbohH Mediates ROS Generation and Enhances Pollen Tube Growth in Fraxinus mandshurica

Flowering plants require normal pollen germination and growth to be fertilized, but studies on the mechanism regulating pollen tube growth in Fraxinus mandshurica are limited. Here, we used transcriptomic data to study the oxidative phosphorylation pathway during pollen tube growth in Fraxinus mandshurica. Our study identified 8,734 differentially expressed genes during the stages S1 to S3 of pollen tube growth. Significant enrichment of the oxidative phosphorylation pathway, amino acid synthesis, protein processing in the ER, carbon metabolism, pyruvate metabolism, citrate cycle (TCA cycle), and glycolysis/gluconeogenesis were examined using the Kyoto Encyclopedia of Genes and Genomes, and 58 genes linked to ROS synthesis and scavenging during the S1–S3 stages were identified. Also, H2DCFDA staining confirmed ROS formation in the pollen and the pollen tubes, and treatment with copper (II) chloride (CuCl2) and diphenyleneiodonium (DPI) was shown to reduce ROS in the pollen tube. Reduction in ROS content caused decreased pollen germination and pollen tube length. Furthermore, FmRbohH (respiratory burst oxidase homolog H) expression was detected in the pollen and pollen tube, and an antisense oligodeoxynucleotide assay demonstrated reduced ROS and pollen tube growth in Fraxinus mandshurica. This study shed more light on the RbohH gene functions during pollen tube growth.

Actin cytoskeleton and plasma membrane aquaporins are involved in different drought response of Arabidopsis rhd2 and der1 root hair mutants

Actin cytoskeleton and reactive oxygen species are principal determinants of root hair polarity and tip growth. Loss of function in RESPIRATORY BURST OXIDASE HOMOLOG C/ROOT HAIR DEFECTIVE 2 (AtRBOHC/RHD2), an NADPH oxidase emitting superoxide to the apoplast, and in ACTIN 2, a vegetative actin isovariant, in rhd2-1 and der1-3 mutants, respectively, lead to similar defects in root hair formation and elongation Since early endosome-mediated polar localization of AtRBOHC/RHD2 depends on actin cytoskeleton, comparing the proteome-wide consequences of both mutations might be of eminent interest. Therefore, we employed a differential proteomic analysis of Arabidopsis rhd2-1 and der1-3 mutants. Both mutants exhibited substantial alterations in abundances of stress-related proteins. Notably, plasma membrane (PM)-localized PIP aquaporins showed contrasting abundance patterns in the mutants compared to wild-types. Drought-responsive proteins were mostly downregulated in rhd2-1 but upregulated in der1-3. Proteomic data suggest that opposite to der1-3, altered vesicular transport in rhd2-1 mutant likely contributes to the deregulation of PM-localized proteins, including PIPs. Moreover, lattice light sheet microscopy revealed reduced actin dynamics in rhd2-1 roots, a finding contrasting with previous reports on der1-3 mutant. Phenotypic experiments demonstrated a drought stress susceptibility in rhd2-1 and resistance in der1-3. Thus, mutations in AtRBOHC/RHD2 and ACTIN2 cause similar root hair defects, but they differently affect the actin cytoskeleton and vesicular transport. Reduced actin dynamics in rhd2-1 mutant is accompanied by alteration of vesicular transport proteins abundance, likely leading to altered protein delivery to PM, including aquaporins, thereby significantly affecting drought stress responses.

Overexpression of OsRbohH Enhances Heat and Drought Tolerance through ROS Homeostasis and ABA Mediated Pathways in Rice (Oryza sativa L.).

Respiratory burst oxidase homologs (Rbohs) are the primary producers of reactive oxygen species (ROS), which have been demonstrated to play critical roles in plant responses to abiotic stress. Here, we explored the function of OsRbohH in heat and drought stress tolerance by generating overexpression lines (OsRbohH-OE). OsRbohH was highly induced by various abiotic stress and hormone treatments. Compared to wild-type (WT) controls, OsRbohH-OE plants exhibited enhanced tolerance to heat and drought, as determined by survival rate analyses and total chlorophyll content. Histochemical staining revealed that OsRbohH-OE accumulated less ROS. This is consistent with the observed increase in catalase (CAT) and peroxidase (POD) activities, as well as a reduced electrolyte leakage rate and malondialdehyde (MDA) content. Moreover, OsRbohH-OE exhibited enhanced sensitivity to exogenous abscisic acid (ABA), accompanied by altered expression levels of ABA synthesis and catabolic genes. Further analysis indicated that transgenic lines had lower transcripts of ABA signaling-related genes (OsDREB2A, OsLEA3, OsbZIP66, and OsbZIP72) under heat but higher levels under drought than WT. In conclusion, these results suggest that OsRbohH is a positive regulator of heat and drought tolerance in rice, which is probably performed through OsRbohH-mediated ROS homeostasis and ABA signaling.