Ethylene Signaling Pathway Research Articles

Ethylene response factor ERF.D7 activates auxin response factor 2 paralogs to regulate tomato fruit ripening.

Despite the obligatory role of ethylene in climacteric fruit ripening and the identification of 77 ethylene response factors (ERFs) in the tomato (Solanum lycopersicum) genome, the role of few ERFs has been validated in the ripening process. Here, using a comprehensive morpho-physiological, molecular, and biochemical approach, we demonstrate the regulatory role of ERF D7 (SlERF.D7) in tomato fruit ripening. SlERF.D7 expression positively responded to exogenous ethylene and auxin treatments, most likely in a ripening inhibitor-independent manner. SlERF.D7 overexpression (OE) promoted ripening, and its silencing had the opposite effect. Alterations in its expression modulated ethylene production, pigment accumulation, and fruit firmness. Consistently, genes involved in ethylene biosynthesis and signaling, lycopene biosynthesis, and cell wall loosening were upregulated in the OE lines and downregulated in RNAi lines. These transgenic lines also accumulated altered levels of indole-3-acetic acid at late-breaker stages. A positive association between auxin response factor 2 (ARF2) paralog's transcripts and SlERF.D7 mRNA levels and that SlARF2A and SlARF2B are direct targets of SlERF.D7 underpinned the perturbed auxin-ethylene crosstalk for the altered ripening program observed in the transgenic fruits. Overall, this study uncovers that SlERF.D7 positively regulates SlARF2A/B abundance to amalgamate auxin and ethylene signaling pathways for controlling tomato fruit ripening.

Elucidation of the interactome of the sucrose transporter StSUT4: sucrose transport is connected to ethylene and calcium signalling.

Sucrose transporters of the SUT4 clade show dual targeting to both the plasma membrane as well as to the vacuole. Previous investigations revealed a role for the potato sucrose transporter StSUT4 in flowering, tuberization, shade avoidance response, and ethylene production. Down-regulation of StSUT4 expression leads to early flowering, tuberization under long days, far-red light insensitivity, and reduced diurnal ethylene production. Sucrose export from leaves was increased and a phase-shift of soluble sugar accumulation in source leaves was observed, arguing for StSUT4 to be involved in the entrainment of the circadian clock. Here, we show that StSUT4, whose transcripts are highly unstable and tightly controlled at the post-transcriptional level, connects components of the ethylene and calcium signalling pathway. Elucidation of the StSUT4 interactome using the split ubiquitin system helped to prove direct physical interaction between the sucrose transporter and the ethylene receptor ETR2, as well as with the calcium binding potato calmodulin-1 (PCM1) protein, and a calcium-load activated calcium channel. The impact of calcium ions on transport activity and dual targeting of the transporter was investigated in detail. For this purpose, a reliable esculin-based transport assay was established for SUT4-like transporters. Site-directed mutagenesis helped to identify a diacidic motif within the seventh transmembrane spanning domain that is essential for sucrose transport activity and targeting, but not required for calcium-dependent inhibition. A link between sucrose, calcium and ethylene signalling has been previously postulated with respect to pollen tube growth, shade avoidance response, or entrainment of the circadian clock. Here, we provide experimental evidence for the direct interconnection of these signalling pathways at the molecular level by direct physical interaction of the main players.

Genome-wide identification of long intergenic non-coding RNAs for Ralstonia solanacearum resistance in tomato (Solanum lycopersicum).

There is growing evidences indicating that long intergenic ncRNAs (lincRNAs) play key roles in plant development and stress responses. To research tomato lincRNA functions during the interaction between tomato and Ralstonia solanacearum, RNA-seq data of tomato plants inoculated with R. solanacearum was analyzed. In this study, 315 possible lincRNAs were identified from RNA-seq data. Then 23 differentially expressed lincRNAs between tomato plants inoculated with R. solanacearum and control were identified and a total of 171 possible target genes for these differentially expressed lincRNAs were predicted. Through GO and KEGG analysis, we found that lincRNA might be involved in jasmonic acid and ethylene signaling pathways to respond to tomato bacterial wilt infection. Furthermore, lincRNA may also be involved in regulating the expression of AGO protein. Subsequently, analysis of expression patterns between differentially expressed lincRNAs and adjacent mRNAs by qRT-PCR revealed that part of lincRNAs and their possible target genes exhibited positive correlation. Taken together, these results suggest that lincRNAs play potential roles in tomato against R. solanacearum infection and will provide fundamental information about the lincRNA-based plant defense mechanisms.

Molecular dissection of rose and Botrytis cinerea pathosystems affected by ethylene

Botrytis cinerea (B. cinerea) is a necrotrophic pathogen that causes significant growth and postharvest commercial losses in cut roses. Postharvest disease severity is affected by ethylene levels during transport and storage conditions. In this study, we identified the relationship between ethylene, fungal growth, and ethylene inhibitors during B. cinerea infection in cut ‘Pink Beauty’ roses. The results suggest that cut rose susceptibility to gray mold disease is associated with the developmental stage of flowers, especially the level of senescence, which is stimulated by ethylene biosynthesis in petals. The mRNA levels of ethylene biosynthesis genes in petals were related to the severity of gray mold; however, their expression levels decreased when the cut roses were completely macerated by B. cinerea. B. cinerea infections in cut flowers activated ethylene biosynthesis and signaling pathways, leading to the activation of RhERFs in the petals. Ethylene regulated the development of B. cinerea infections in the cut flowers directly by binding to the receptors or indirectly by promoting the ethylene response in the host tissues. The suppression of ethylene responses in petals and inhibition of the fungal histidine kinase receptor of B. cinerea by 1-methylcylopropene (1-MCP) made cut rose flowers resistant to such necrotrophic pathogens. We also established a working model for ethylene binding and plant and fungus actions in the rose-B. cinerea pathosystem. Understanding the relationship between ethylene and gray mold disease in cut roses and the interaction between ethylene inhibitors and B. cinerea will help improve postharvest treatments for reducing B. cinerea damage in cut flowers.

A 1-aminocyclopropane-1-carboxylic-acid (ACC) dipeptide elicits ethylene responses through ACC-oxidase mediated substrate promiscuity.

Plants produce the volatile hormone ethylene to regulate many developmental processes and to deal with (a)biotic stressors. In seed plants, ethylene is synthesized from 1-aminocyclopropane-1-carboxylic acid (ACC) by the dedicated enzyme ACC oxidase (ACO). Ethylene biosynthesis is tightly regulated at the level of ACC through ACC synthesis, conjugation and transport. ACC is a non-proteinogenic amino acid, which also has signaling roles independent from ethylene. In this work, we investigated the biological function of an uncharacterized ACC dipeptide. The custom-synthesized di-ACC molecule can be taken up by Arabidopsis in a similar way as ACC, in part via Lysine Histidine Transporters (e.g., LHT1). Using Nano-Particle Assisted Laser Desoprtion/Ionization (Nano-PALDI) mass-spectrometry imaging, we revealed that externally fed di-ACC predominantly localizes to the vasculature tissue, despite it not being detectable in control hypocotyl segments. Once taken up, the ACC dimer can evoke a triple response phenotype in dark-grown seedlings, reminiscent of ethylene responses induced by ACC itself, albeit less efficiently compared to ACC. Di-ACC does not act via ACC-signaling, but operates via the known ethylene signaling pathway. In vitro ACO activity and molecular docking showed that di-ACC can be used as an alternative substrate by ACO to form ethylene. The promiscuous nature of ACO for the ACC dimer also explains the higher ethylene production rates observed in planta, although this reaction occurred less efficiently compared to ACC. Overall, the ACC dipeptide seems to be transported and converted into ethylene in a similar way as ACC, and is able to augment ethylene production levels and induce subsequent ethylene responses in Arabidopsis.

Genome-Wide Analysis of the ERF Family and Identification of Potential Genes Involved in Fruit Ripening in Octoploid Strawberry.

Ethylene response factors (ERFs) belonging to the APETALA2/ERF superfamily acted at the end of the ethylene signaling pathway, and they were found to play important roles in plant growth and development. However, the information of ERF genes in strawberry and their involvement in fruit ripening have been limited. Here, a total of 235 ERF members were identified from 426 AP2/ERF genes at octoploid strawberry genome level and classified into six subgroups according to their sequence characteristics and phylogenetic relationship. Conserved motif and gene structure analysis supported the evolutionary conservation of FaERFs. Syntenic analysis showed that four types of duplication events occurred during the expansion of FaERF gene family. Of these, WGD/segmental duplication played a major role. Transcriptomic data of FaERF genes during fruit ripening and in response to abscisic acid screened one activator (FaERF316) and one repressor (FaERF118) that were involved in fruit ripening. Transcriptional regulation analysis showed some transcription factors related to ripening such as ABI4, TCP15, and GLK1 could bind to FaERF316 or FaERF118 promoters, while protein–protein interaction analysis displayed some proteins associated with plant growth and development could interact with FaERF118 or FaERF316. These results suggested that FaERF118 and FaERF316 were potential genes to regulate strawberry ripening. In summary, the present study provides the comprehensive and systematic information on FaERF family evolution and gains insights into FaERF’s potential regulatory mechanism in strawberry ripening.

New Inhibitors of the Human p300/CBP Acetyltransferase Are Selectively Active against the Arabidopsis HAC Proteins

Histone acetyltransferases (HATs) are involved in the epigenetic positive control of gene expression in eukaryotes. CREB-binding proteins (CBP)/p300, a subfamily of highly conserved HATs, have been shown to function as acetylases on both histones and non-histone proteins. In the model plant Arabidopsis thaliana among the five CBP/p300 HATs, HAC1, HAC5 and HAC12 have been shown to be involved in the ethylene signaling pathway. In addition, HAC1 and HAC5 interact and cooperate with the Mediator complex, as in humans. Therefore, it is potentially difficult to discriminate the effect on plant development of the enzymatic activity with respect to their Mediator-related function. Taking advantage of the homology of the human HAC catalytic domain with that of the Arabidopsis, we set-up a phenotypic assay based on the hypocotyl length of Arabidopsis dark-grown seedlings to evaluate the effects of a compound previously described as human p300/CBP inhibitor, and to screen previously described cinnamoyl derivatives as well as newly synthesized analogues. We selected the most effective compounds, and we demonstrated their efficacy at phenotypic and molecular level. The in vitro inhibition of the enzymatic activity proved the specificity of the inhibitor on the catalytic domain of HAC1, thus substantiating this strategy as a useful tool in plant epigenetic studies.

Early infection response of fungal biotroph Ustilago maydis in maize.

Common smut, caused by Ustilago maydis (DC.) Corda, is a destructive fungal disease of maize worldwide; it forms large tumors, reducing corn yield and quality. However, the molecular defense mechanism to common smut in maize remains unclear. The present study aimed to use a leading maize inbred line Ye478 to analyze the response to U. maydis inoculation. The histological and cytological analyses demonstrated that U. maydis grew gradually to the host cells 6 h post-inoculation (hpi). The samples collected at 0, 3, 6, and 12 hpi were analyzed to assess the maize transcriptomic changes in response to U. maydis. The results revealed differences in hormone signaling, glycometabolism, and photosynthesis after U. maydis infection; specific changes were detected in jasmonic acid (JA), salicylic acid (SA), ethylene (ET), and abscisic acid (ABA) signaling pathways, glycolysis/gluconeogenesis, and photosystems I and II, probably related to defense response. MapMan analysis demonstrated that the differentially expressed genes between the treatment and control groups were clustered into light reaction and photorespiration pathways. In addition, U. maydis inoculation induced chloroplast swelling and damage, suggesting a significant effect on the chloroplast activity and subsequent metabolic process, especially hexose metabolism. A further genetic study using wild-type and galactinol-sucrose galactosyltransferase (gsg) and yellow-green leaf-1 (ygl-1) mutants identified that these two U. maydis-induced genes negatively regulated defense against common smut in maize. Our measurements showed the pathogen early-invasion process, and the key pathways of both chlorophyll biosynthesis and sugar transportation were critical modified in the infected maize line, thereby throwing a light on the molecular mechanisms in the maize-U. maydis interaction.

Perfluorobutanoic Acid (PFBA) Induces a Non-Enzymatic Oxidative Stress Response in Soybean (Glycine max L. Merr.).

Short-chain perfluoroalkyl substances (PFAS) are generally considered to be of less environmental concern than long-chain analogues due to their comparatively shorter half-lives in biological systems. Perfluorobutanoic acid (PFBA) is a short-chain PFAS with the most root–shoot transfer factor of all PFAS. We investigated the impact of extended exposure of soybean plants to irrigation water containing environmentally relevant (100 pg–100 ng/L) to high (100 µg–1 mg/L) concentrations of PFBA using phenotypical observation, biochemical characterization, and transcriptomic analysis. The results showed a non-monotonous developmental response from the plants, with maximum stimulation and inhibition at 100 ng/L and 1 mg/L, respectively. Higher reactive oxygen species and low levels of superoxide dismutase (SOD) and catalase (CAT) activity were observed in all treatment groups. However transcriptomic analysis did not demonstrate differential expression of SOD and CAT coding genes, whereas non-enzymatic response genes and pathways were enriched in both groups (100 ng/L and 1 mg/L) with glycine betaine dehydrogenase showing the highest expression. About 18% of similarly downregulated genes in both groups are involved in the ethylene signaling pathway. The circadian rhythm pathway was the only differentially regulated pathway between both groups. We conclude that, similar to long chain PFAS, PFBA induced stress in soybean plants and that the observed hormetic stimulation at 100 ng/L represents an overcompensation response, via the circadian rhythm pathway, to the induced stress.

Comparative Transcriptome Analysis Reveals the Mechanisms Underlying Differential Seed Vigor in Two Contrasting Peanut Genotypes

Seed vigor is an important agronomic trait, and wide variation exists among peanut accessions. However, the detailed regulatory mechanisms underlying differences in seed vigor between varieties are not known in peanut yet. Here, we performed a comparative transcriptome analysis of germinating seeds in two contrasting peanut accessions, namely A86 (high-vigor variety) and A279 (low-vigor variety). A total of 583 and 860 differentially expressed genes (DEGs) were identified at two imbibition stages between A86 and A279, respectively. Pathway enrichment tests highlighted the cell wall remodeling-, hormone signaling-, transcriptional regulation-, and oxidative stress-related DEGs, which may explain to a certain extent the difference in seed vigor between the two cultivars. Among them, the largest number of cell wall remodeling-related DEGs were extensions followed by cellulose synthases, fasciclin-like arabinogalactan proteins, polygalacturonases, expansins, and pectinesterases and the hormone signaling-related DEGs belonged mainly to the auxin and ethylene signaling pathway. The majority of transcriptional regulation-related DEGs were MYB, FAR1, and bHLH transcription factors, and the oxidative stress-related DEGs were mainly peroxidases. Further physiological analyses indicated that differences in seed vigor between A86 and A279 may be associated with differences in the ROS-scavenging abilities mediated by peroxidases. Moreover, we identified 16 DEGs homologous to known Arabidopsis regulators of seed dormancy and germination, suggesting that these DEGs would play similar functional roles during peanut seed germination. Our results not only provide important insights into the difference in seed vigor between varieties, but offer candidate genes that are worth investigating in future studies.

Comparative Transcriptome Analyses between Resistant and Susceptible Varieties in Response to Soybean Mosaic Virus Infection

Soybean mosaic virus (SMV) is a worldwide and hardly controlled virus disease in soybean. Kefeng-1 is an elite variety resistant to SMV in China. In order to discover resistance genes and regulation networks in Kefeng-1, we analyzed transcriptome data of resistant (Kefeng-1) and susceptible (NN1138-2) soybean varieties in response to infection of the SMV strain SC18 at 0, 6, and 48 hours post-inoculation (hpi) and 5 days post-inoculation (dpi). Many differentially expressed genes (DEGs) were identified with Kefeng-1 and NN 1138-2. Based on the enrichment analysis for gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, we found that 48 hpi was the best time point for the defense response of the two soybean varieties in response to the SMV infection. The expression of seven candidate genes was further verified by qRT-PCR and was relatively consistent with the results of RNA-Seq. The expression of genes for Glyma.11G239000 and Glyma.18G018400, members of the ethylene-insensitive 3/ethylene-insensitive3-like (EIN3/EIL) protein family involved in ETH, were downregulated in NN1138-2 but not in Kefeng-1 and the expression of Glyma.14G041500 was upregulated in Kefeng-1 at 5 dpi. The expression of jasmonic acid repressor genes (TIFY/JAZ) was downregulated in NN1138-2 but not in Kefeng-1. NPR1 involved in the salicylic acid signaling pathway was downregulated in NN1138-2 at 48 hpi but upregulated in Kefeng-1. It shows that ethylene, jasmonic acid, and salicylic acid signaling pathways may be involved in the disease resistance process to the SMV strain SC18. Our findings would help to understand the molecular mechanism of soybean resistance to SMV.

Gene Expression, Histology and Histochemistry in the Interaction between Musa sp. and Pseudocercospora fijiensis.

Bananas are the main fruits responsible for feeding more than 500 million people in tropical and subtropical countries. Black Sigatoka, caused by the fungus Pseudocercospora fijiensis, is one of the most destructive disease for the crop. This fungus is mainly controlled with the use of fungicides; however, in addition to being harmful to human health, they are associated with a high cost. The development of resistant cultivars through crosses of susceptible commercial cultivars is one of the main focuses of banana breeding programs worldwide. Thus, the objective of the present study was to investigate the interaction between Musa sp. and P. fijiensis through the relative expression of candidate genes involved in the defence response to black Sigatoka in four contrasting genotypes (resistant: Calcutta 4 and Krasan Saichon; susceptible: Grand Naine and Akondro Mainty) using quantitative real-time PCR (RT–qPCR) in addition to histological and histochemical analyses to verify the defence mechanisms activated during the interaction. Differentially expressed genes (DEGs) related to the jasmonic acid and ethylene signalling pathway, GDSL-like lipases and pathogenesis-related proteins (PR-4), were identified. The number and distance between stomata were directly related to the resistance/susceptibility of each genotype. Histochemical tests showed the production of phenolic compounds and callosis as defence mechanisms activated by the resistant genotypes during the interaction process. Scanning electron microscopy (SEM) showed pathogenic structures on the leaf surface in addition to calcium oxalate crystals. The resistant genotype Krasan Saichon stood out in the analyses and has potential for use in breeding programs for resistance to black Sigatoka in banana and plantains.

Ethylene pretreatment induces phenolic biosynthesis of fresh-cut pitaya fruit by regulating ethylene signaling pathway

Understanding the role of signals involved in wounding responses allows designing and improving preservation technology of fresh-cut products. The present work evaluated the role of ethylene as a signal molecule on phenolic biosynthesis induced by wounding in fresh-cut pitaya fruit at the physiological and gene expression levels. Ethylene pretreatment positively promoted the increases of phenolics and flavonoids of fresh-cut pitaya fruit by improving activities and relative gene expressions of the pivotal enzymes in phenylpropanoid pathway. It accelerated reactive oxygen species (ROS) production at the beginning of storage but caused lower ROS levels during the middle and later periods. Moreover, fresh-cut pitaya fruit pretreated with ethylene showed lower ethylene production rate in comparison with the control, but higher expression levels of ethylene receptor genes (HuETR1 and HuETR2), transcription factor genes (HuEIN3s and HuERF1s) and lower expression levels of the negative ethylene regulator HuCTR1s. On the contrary, the ethylene receptor inhibitor, 1-MCP pretreatment not only reduced the wound-induced phenolic accumulation, prevented the activation of ROS and phenylpropane metabolism, but also inhibited the up-regulation of HuETR1, HuETR2, HuEIN3s and HuERF1s. Taken together, our present work suggested that ethylene pretreatment induced phenolic biosynthesis in fresh-cut pitaya fruit by regulating ethylene signaling pathway.

Comparative transcriptome analysis of broccoli seedlings under different Cd exposure levels revealed possible pathways involved in hormesis

As a non-essential heavy metal, Cd has long been considered to cause plant damage and reduce crop productivity. Recently, accumulating studies have shown that low-level Cd shows beneficial effects on plants, called hormesis. However, the mechanism of low-level Cd triggered hormesis is largely unknown. In this study, we found that Cd showed a hormesis effect on broccoli seedlings. The comparative transcriptome analysis of broccoli seedlings under low-level and high-level Cd showed that the plants responded distinctively to different concentrations of Cd exposure. KEGG analysis suggested that plant hormone signaling and secondary metabolism, especially glucosinolate homeostasis, were important under both low and high-level Cd treatment. Interestingly, ethylene signaling pathway, auxin signaling pathway, glucosinolate degradation, and glutathione redox showed opposite responses to low and high-level Cd. Low-level Cd inhibited while high-level Cd promoted ethylene signaling. Low-level Cd enhanced auxin biosynthesis while high-level repressed auxin biosynthesis. Glucosinolate degradation was enhanced by low-level Cd and inhibited by high-level Cd. As an important indicator of cell redox state, the ratio of reduced glutathione to oxidized glutathione (GSH/GSSG) increased under low-level and decreased under high-level Cd. Since the above biological processes play important roles in both growth and Cd detoxification, their opposite response to different levels of Cd indicated their possible involvement in hormesis. The potential functions of these processes in hormesis are discussed. Our study shed a light on the underlying mechanisms of the Cd-mediated hormesis effect.

Identification and Characterization of Long Non-coding RNA in Tomato Roots Under Salt Stress.

As one of the most important vegetable crops in the world, the production of tomatoes was restricted by salt stress. Therefore, it is of great interest to analyze the salt stress tolerance genes. As the non-coding RNAs (ncRNAs) with a length of more than 200 nucleotides, long non-coding RNAs (lncRNAs) lack the ability of protein-coding, but they can play crucial roles in plant development and response to abiotic stresses by regulating gene expression. Nevertheless, there are few studies on the roles of salt-induced lncRNAs in tomatoes. Therefore, we selected wild tomato Solanum pennellii (S. pennellii) and cultivated tomato M82 to be materials. By high-throughput sequencing, 1,044 putative lncRNAs were identified here. Among them, 154 and 137 lncRNAs were differentially expressed in M82 and S. pennellii, respectively. Through functional analysis of target genes of differentially expressed lncRNAs (DE-lncRNAs), some genes were found to respond positively to salt stress by participating in abscisic acid (ABA) signaling pathway, brassinosteroid (BR) signaling pathway, ethylene (ETH) signaling pathway, and anti-oxidation process. We also construct a salt-induced lncRNA-mRNA co-expression network to dissect the putative mechanisms of high salt tolerance in S. pennellii. We analyze the function of salt-induced lncRNAs in tomato roots at the genome-wide levels for the first time. These results will contribute to understanding the molecular mechanisms of salt tolerance in tomatoes from the perspective of lncRNAs.

Loss of GSNOR increases abiotic stress sensitivity via regulating MAPK-ethylene cascade signaling in Solanum lycopersicum L.

Nitric oxide (NO), as a redox molecule, played important role in plant response to environmental stress. Here, we found that loss of tomato S-Nitrosoglutathione reductase ( SlGSNOR ), a critical regulator of NO balance, led to global sensitivity to heat, salt, bicarbonate, and paraquat stresses in tomato, suggesting that SlGSNOR was a positive regulator in tomato against abiotic stresses. In addition, under these stresses, loss of SlGSNOR induced excessive NO accumulation in the roots of the transgenic plant. Bicarbonate accumulation has been a limiting factor of tomato production in the north of China, and there were less reports on the regulation of bicarbonate stress compared with other abiotic stress, therefore, in the following experiment, the mechanism of SlGSNOR function focused on investigating bicarbonate tolerance. Proteomes data indicated that loss of SlGSNOR triggered the expression of proteins in MAPK and ethylene signaling pathway under bicarbonate condition. Importantly, loss of SlGSNOR increased ethylene emission under bicarbonate condition, meaning that ethylene signaling participated in SlGSNOR -mediated tomato bicarbonate stress tolerance. Interestingly, under bicarbonate stress, silencing SlMAPK3 or SlACO1 in SlGSNOR -RNAi tomato partially compromised the sensitivity and ethylene emission. Mechanistically, under bicarbonate stress, silencing SlMAPK3 in SlGSNOR -RNAi tomato plants suppressed the transcription of SlACO1 , whereas silencing SlACO1 in SlGSNOR -RNAi tomato activated the transcription of SlMAPK3 . Collectively, these data demonstrated that SlGSNOR -mediated tomato abiotic stress tolerance was depended on SlMAPK3 - SlACO1 cascade signaling, largely. In addition, the incorporation of global abiotic stress tolerance traits into crop plants may succeed by manipulating GSNOR expression. • Tomato GSNOR was a multiple stress-induced gene. • SlGSNOR was positively regulated tomato heat, salt, bicarbonate, and paraquat tolerance. • SlGSNOR -mediated bicarbonate tolerance was depended on SlMAPK3 - SlACO1 cascade signaling.

Ethylene insensitive 2 (EIN2) as a potential target gene to enhance Fusarium head blight disease resistance

Fusarium head blight (FHB) caused by Fusarium graminearum (Fg) severely affects cereal crops, especially wheat and barley. FHB results in significant yield loss, reduces grain quality and contaminates grains with mycotoxin. The development of FHB-resistant cereal cultivars can be expedited through CRISPR gene editing. The Arabidopsis ethylene insensitive 2 (AtEIN2) plays a key role in ethylene signaling pathway and is critical for monitoring plant growth and defense responses. RNAi down-regulation of the wheat homolog TaEIN2 has been shown to enhance wheat FHB resistance. Here we generated site-specific mutations in AtEIN2 by CRISPR-editing. Detached inflorescence infection assays revealed that AtEIN2 knock-out (KO) mutants displayed enhanced Fg resistance and substantially reduced Fg spore production in planta. Gene expression profiling of defense genes revealed that impairment of AtEIN2 resulted in down-regulation of the ethylene signaling pathway while the salicylic acid signaling pathway was unaffected. Complementation of AtEIN2-KO plants with a barley orthologue, HvEIN2, restored Fg susceptibility, indicating that HvEIN2 is functionally equivalent to its Arabidopsis counterpart and, hence, may have a similar role in conditioning barley Fg susceptibility. These results provide insight into the defense role of EIN2 and a molecular and functional foundation for manipulating HvEIN2 to enhance FHB resistance in barley.

Attenuation of ethylene signaling increases cotton resistance to a defoliating strain of Verticillium dahliae

The severity of Verticillium wilt on cotton caused by defoliating strains of Verticillium dahliae has gradually increased and threatens production worldwide. Identification of the molecular components of leaf defoliation may increase cotton tolerance to V. dahliae. Ethylene, a major player in plant physiological processes, is often associated with senescence and defoliation of plants. We investigated the cotton–V. dahliae interaction with a focus on the role of ethylene in defoliation and defense against V. dahliae. Cotton plants inoculated with V. dahliae isolate V991, a defoliating strain, accumulated more ethylene and showed increased disease symptoms than those inoculated with a non-defoliating strain. In cotton with a transiently silenced ethylene synthesis gene (GhACOs) and signaling gene (GhEINs) during cotton–V. dahliae interaction, ethylene produced was derived from cotton and more ethylene increased cotton susceptibility and defoliation rate. Overexpression of AtCTR1, a negative regulator in ethylene signaling, in cotton reduced sensitivity to ethylene and increased plant resistance to V. dahliae. Collectively, the results indicated precise regulation of ethylene synthesis or signaling pathways improve cotton resistant to Verticillium wilt.

Activation of plant immunity by exposure to dinitrogen pentoxide gas generated from air using plasma technology.

Reactive nitrogen species (RNS) play an important role in plant immunity as signaling factors. We previously developed a plasma technology to partially convert air molecules into dinitrogen pentoxide (N2O5), an RNS whose physiological action is poorly understood. To reveal the function of N2O5 gas in plant immunity, Arabidopsis thaliana was exposed to plasma-generated N2O5 gas once (20 s) per day for 3 days, and inoculated with Botrytis cinerea, Pseudomonas syringae pv. tomato DC3000 (Pst), or cucumber mosaic virus strain yellow (CMV(Y)) at 24 h after the final N2O5 gas exposure. Lesion size with B. cinerea infection was significantly (P < 0.05) reduced by exposure to N2O5 gas. Propagation of CMV(Y) was suppressed in plants exposed to N2O5 gas compared with plants exposed to the air control. However, proliferation of Pst in the N2O5-gas-exposed plants was almost the same as in the air control plants. These results suggested that N2O5 gas exposure could control plant disease depending on the type of pathogen. Furthermore, changes in gene expression at 24 h after the final N2O5 gas exposure were analyzed by RNA-Seq. Based on the gene ontology analysis, jasmonic acid and ethylene signaling pathways were activated by exposure of Arabidopsis plants to N2O5 gas. A time course experiment with qRT-PCR revealed that the mRNA expression of the transcription factor genes, WRKY25, WRKY26, WRKY33, and genes for tryptophan metabolic enzymes, CYP71A12, CYP71A13, PEN2, and PAD3, was transiently induced by exposure to N2O5 gas once for 20 s peaking at 1–3 h post-exposure. However, the expression of PDF1.2 was enhanced beginning from 6 h after exposure and its high expression was maintained until 24–48 h later. Thus, enhanced tryptophan metabolism leading to the synthesis of antimicrobial substances such as camalexin and antimicrobial peptides might have contributed to the N2O5-gas-induced disease resistance.

High light stress induces H2O2 production and accelerates fruit ripening in tomato

Increased synthesis of H2O2 is observed during the initiation of fruit ripening. However, its association with plant cell processes triggering the maturation of fruit has not yet been demonstrated. The aim of this work is to investigate whether H2O2 participates in the tomato ripening process and particularly through its association with the ethylene signaling pathway. The experiments were carried out with two ethyl methanesulfonate mutant lines of Micro-Tom tomato deficient in GDP-L-galactose phosphorylase activity and displaying lower ascorbic acid content than the corresponding parental genotype (i.e. wild type). Plants were subjected to a high irradiance (HI) treatment to stimulate H2O2 synthesis. HI treatment enhanced H2O2 production and reduced the timing of fruit ripening in both mutants and wild-type fruits. These results could be linked to an increase of the expression of H2O2-related genes and changes in the expression of ethylene-related genes. The fruit H2O2 production increased or decreased after applying the treatments that induced ethylene synthesis or blocked its action, respectively. The results presented in this work give an evidence of the association of redox and hormonal components during fruit ripening in which H2O2 participates downstream in the events regulated by ethylene.