Ethylene Response Factor Research Articles

Osmanthus fragrans Ethylene Response Factor OfERF1-3 Delays Petal Senescence and Is Involved in the Regulation of ABA Signaling

Osmanthus fragrans is widely used in gardening, but the short flowering period of O. fragrans affects its ornamental and economic value. ERF, as a plant ethylene response factor, is an important link in the regulation of plant senescence. In this study, we conducted a comprehensive analysis of the functional role of OfERF1-3 within the petals of O. fragrans. Specifically, the OfERF1-3 gene was cloned and subjected to rigorous sequence analysis. Subsequently, to evaluate its expression patterns and effects, gene overexpression experiments were carried out on both Nicotiana tabacum and O. fragrans. The results showed that OfERF1-3-overexpressing tobacco plants exhibited longer petal opening times compared with those of wild plants. Measurements of physiological parameters also showed that the flowers of overexpressed tobacco plants contained lower levels of malondialdehyde (MDA) and hydrogen peroxide (H2O2) than those of the wild type. There was a lower expression of senescence marker genes in overexpressed tobacco and O. fragrans. A yeast two-hybrid assay showed that OfERF1-3 interacted with OfSKIP14 in a manner related to the regulation of ABA. In summary, OfERF1-3 can play a delaying role in the petal senescence process in O. fragrans, and it interacts with OfSKIP14 to indirectly affect petal senescence by regulating the ABA pathway.

The ERF transcription factor ZbERF3 promotes ethylene-induced anthocyanin biosynthesis in Zanthoxylum bungeanum

Ethylene regulates fruit ripening, and in Zanthoxylum bungeanum, fruit color deepened with increasing of ethylene during fruit ripening. However, the molecular mechanism of this physiological process was still unclear. In this study, through the combined analysis of transcriptome and metabolome, it was found that ethylene release was consistent with anthocyanin synthesis, and ethylene response factors (ERFs) were significantly related to anthocyanin biosynthesis during the fruit ripening of Z. bungeanum. Ethylene treatment significantly induced fruit coloration and promoted anthocyanin synthesis and the expression of ZbERF3. Furthermore, Yeast one-hybrid assays and Luciferase reporter assays demonstrated that ZbERF3 directly bound to the promoter of ZbMYB17 and transcriptionally activated its expression. What's more, it was demonstrated that ZbMYB17 directly bound to the promoter of ZbANS, promoting anthocyanin biosynthesis. Overall, this study revealed the mechanism of ERF and MYB synergistically regulating the coloration of Z. bungeanum fruit.

ERFVII-controlled hypoxia responses are in part facilitated by MEDIATOR SUBUNIT 25 in Arabidopsis thaliana.

Flooding impairs plant growth through oxygen deprivation, which activates plant survival and acclimation responses. Transcriptional responses to low oxygen are generally associated with the activation of group VII ETHYLENE-RESPONSE FACTOR (ERFVII) transcription factors. However, the exact mechanisms and molecular components by which ERFVII factors initiate gene expression are not fully elucidated. Here, we show that the ERFVII factors RELATED TO APETALA 2.2 (RAP2.2) and RAP2.12 cooperate with the Mediator complex subunit AtMED25 to coordinate gene expression under hypoxia in Arabidopsis thaliana. Respective med25 knock-out mutants display reduced low-oxygen stress tolerance. AtMED25 physically associates with a distinct set of hypoxia core genes and its loss partially impairs transcription under hypoxia due to decreased RNA polymerase II recruitment. Association of AtMED25 with target genes requires the presence of ERFVII transcription factors. Next to ERFVII protein stabilisation, also the composition of the Mediator complex including AtMED25 is potentially affected by hypoxia stress as shown by protein-complex pulldown assays. The dynamic response of the Mediator complex to hypoxia is furthermore supported by the fact that two subunits, AtMED8 and AtMED16, are not involved in the establishment of hypoxia tolerance, whilst both act in coordination with AtMED25 under other environmental conditions. We furthermore show that AtMED25 function under hypoxia is independent of ethylene signalling. Finally, functional conservation at the molecular level was found for the MED25-ERFVII module between A. thaliana and the monocot species Oryza sativa, pointing to a potentially universal role of MED25 in coordinating ERFVII-dependent transcript responses to hypoxia in plants.

PfERF106, a novel key transcription factor regulating the biosynthesis of floral terpenoids in Primula forbesii Franch

BackgroundFlowers can be a source of essential oils used in the manufacture of substances with high economic value. The ethylene response factor (ERF) gene family plays a key role in regulating secondary metabolite biosynthesis in plants. However, until now, little has been known about the involvement of ERF transcription factors (TFs) in floral terpenoid biosynthesis.ResultsIn this study, an aromatic plant, Primula forbesii Franch., was used as research material to explore the key regulatory effects of PfERF106 on the biosynthesis of terpenoids. PfERF106, which encodes an IXb group ERF transcription factor, exhibited a consistent expression trend in the flowers of P. forbesii and was transcriptionally induced by exogenous ethylene. Transient silencing of PfERF106 in P. forbesii significantly decreased the relative contents of key floral terpenes, including (z)-β-ocimene, sabinene, β-pinene, γ-terpinene, linalool, eremophilene, α-ionone, and α-terpineol. In contrast, constitutive overexpression of PfERF106 in transgenic tobacco significantly increased the relative contents of key floral terpenes, including cis-3-hexen-1-ol, linalool, caryophyllene, cembrene, and sclareol. RNA sequencing of petals of PfERF106-silenced plants and empty-vector control plants revealed 52,711 expressed unigenes and 9,060 differentially expressed genes (DEGs). KEGG annotation analysis revealed that the DEGs were enriched for involvement in secondary metabolic biosynthetic pathways, including monoterpene and diterpene synthesis. Notably, 10 downregulated DEGs were determined to be the downstream target genes of PfERF106 affecting the biosynthesis of terpenoids in P. forbesii.ConclusionThis study characterized the key positive regulatory effects of PfERF106 on the biosynthesis of terpenoids, indicating high-quality genetic resources for aroma improvement in P. forbesii. Thus, this study advances the artificial and precise directional regulation of metabolic engineering of aromatic substances.

Genome-wide analysis of the ERF Family in Stephania japonica provides insights into the regulatory role in Cepharanthine biosynthesis.

Cepharanthine (CEP), a bisbenzylisoquinoline alkaloid (bisBIA) extracted from Stephania japonica, has received significant attention for its anti-coronavirus properties. While ethylene response factors (ERFs) have been reported to regulate the biosynthesis of various alkaloids, their role in regulating CEP biosynthesis remains unexplored. Genome-wide analysis of the ERF genes was performed with bioinformatics technology, and the expression patterns of different tissues, were analyzed by transcriptome sequencing analysis and real-time quantitative PCR verification. The nuclear-localized ERF gene cluster was shown to directly bind to the promoters of several CEP-associated genes, as demonstrated by yeast one-hybrid assays and subcellular localization assays. In this work, 59 SjERF genes were identified in the S. japonica genome and further categorized into ten subfamilies. Notably, a SjERF gene cluster containing three SjERF genes was found on chromosome 2. Yeast one-hybrid assays confirmed that the SjERF gene cluster can directly bind to the promoters of several CEP-associated genes, suggesting their crucial role in CEP metabolism. The SjERFs cluster-YFP fusion proteins were observed exclusively in the nuclei of Nicotiana benthamiana leaves. Tissue expression profiling revealed that 13 SjERFs exhibit high expression levels in the root, and the qRT-PCR results of six SjERFs were consistent with the RNA-Seq data. Furthermore, a co-expression network analysis demonstrated that 24 SjERFs were highly positively correlated with the contents of various alkaloids and expression levels of CEP biosynthetic genes. This study provides the first systematic identification and analysis of ERF transcription factors in the S.japonica genome, laying the foundation for the future functional research of SjERFs transcription factors.

Potassium indole-3-butyric acid affects rice's adaptability to salt stress by regulating carbon metabolism, transcription factor genes expression, and biosynthesis of secondary metabolites.

Soil salinity pollution is increasing worldwide, seriously affecting plant growth and crop production. Existing reports on how potassium indole-3-butyric acid (IBAK) regulates rice salt stress adaptation by affecting rice carbon metabolism, transcription factor (TF) genes expression, and biosynthesis of secondary metabolites still have limitations. In this study, an IBAK solution at 40 mg L-1 was sprayed on rice leaves at the seedling stage. The results showed that the IBAK application could promote shoot and root growth, decrease sucrose and fructose content, increase starch content, and enhance acid invertase (AI) and neutral invertase (NI) activity under salt stress, indicating altered carbon allocation. Furthermore, the expression of TF genes belonging to the ethylene responsive factor (ERF), WRKY, and basic helix-loop-helix (bHLH) families was influenced by IBAK. Many key genes (OsSSIIc, OsSHM1, and OsPPDKB) and metabolites (2-oxoglutaric acid, fumaric acid, and succinic acid) were upregulated in the carbon metabolism pathway. In addition, this study highlighted the role of IBAK in regulating the biosynthesis of secondary metabolites pathway, potentially contributing to rice stress adaptability. The results of this study can provide new sustainable development solutions for agricultural production.

Jasmonic acid plays an important role in mediating retrograde signaling under mitochondrial translational stress to balance plant growth and defense

Proper mitochondrial function is crucial to plant growth and development. Inhibition of mitochondrial translation leads to mitochondrial proteotoxic stress, which triggers a protective transcriptional response that regulates nuclear gene expression, commonly referred to as the mitochondrial unfolded protein response (UPRmt). Although the UPRmt has been extensively studied in yeast and mammals, very little is known about the UPRmt in plants. Here, we show that mitochondrial translational stress inhibits plant growth and development by inducing jasmonic acid (JA) biosynthesis and signaling. The inhibitory effect of mitochondrial translational stress on plant growth was alleviated in the JA-signaling-defective mutants coi1-2, myc2, and myc234. Genetic analysis indicated that Arabidopsis mitochondrial ribosomal protein L1 (MRPL1), a key factor in the UPRmt, regulates plant growth in a CORONATINE-INSENSITIVE 1 (COI1)-dependent manner. Moreover, under mitochondrial translational stress, MYC2 shows direct binding to G boxes in the ETHYLENE RESPONSE FACTOR 109 (ERF109) promoter. The induction of ERF109 expression enhances hydrogen peroxide production, which acts as a feedback loop to inhibit root growth. In addition, mutation of MRPL1 increases JA accumulation, reduces plant growth, and enhances biotic stress resistance. Overall, our findings reveal that JA plays an important role in mediating retrograde signaling under mitochondrial translational stress to balance plant growth and defense.

Integrated gene expression and network analysis identify drought-response genes and pathways in Solanum: a computational study

At times of rapid global climatic changes, refining drought tolerance in tomatoes is a demanding concern. Drought-resistant wild tomato relatives offer a valued resource for studying drought response mechanisms and leveraging them for breeding. To understand drought-response mechanisms in Solanum, we performed an integrated gene expression, transcription factor (TF), and network-based analysis in wild (Solanum pimpinellifolium L.) and cultivated (Solanum lycopersicum L.) tomato species with differing drought tolerance. This integrated analysis unveiled distinct gene expression patterns under drought stress in both species. Ethylene-responsive factors (ERFs) were notably the most significantly expressed transcription factors in both varieties. Additionally, species-specific TF expression was observed, such as Ethylene Insensitive3-like (EIL) and MIKC_MADS in the wild type, and Zinc Finger homeodomain (ZF-HD) in the cultivated variety. Differential expression pattern in ROS rummaging and heat shock proteins was observed in the top DEGs of both species. Network analysis highlighted shared genes involved in crucial pathways like jasmonic acid biosynthesis, the protein kinase pathway, and reactive oxygen species foraging in both species. Notably, the wild variety showed altered expression of genes associated with jasmonic acid/ethylene biosynthesis, antioxidant response, heat shock proteins, and cell wall remodeling. These observations offer insights into the differences in drought response in both species and suggest the role of jasmonic acid/ethylene biosynthesis, antioxidant response and ROS scavenging as significant factors contributing to improved drought tolerance in S. pimpinellifolium.

Mechanistic Insights and Genetic Approaches for Enhancing Submergence Tolerance in Rice

Rice, a pivotal staple crop, serves as a dietary foundation for approximately half of the global population. Its cultivation, traditionally reliant on submerged conditions, faces significant challenges due to sudden flooding events. The majority of rice varieties are highly sensitive to prolonged submersion beyond a mere seven days, resulting in substantial yield losses. As global climate patterns shift, the occurrence and intensity of flooding are predictable to increase, exacerbating this issue. Therefore, development of rice varieties having enhanced submergence tolerance is critical to sustaining productivity and ensuring food security in flood-prone regions. Understanding the physiological and biochemical mechanisms underlying submergence tolerance is crucial in this context. Key physiological mechanisms include the regulation of carbohydrate reserves for energy during low oxygen availability, ethylene-responsive factors, especially the Sub1A gene to, regulate growth and stress responses, curbing excessive elongation to conserve energy and enhance survival. Additionally, specialized tissues such as aerenchyma and adventitious roots improve oxygen transport and nutrient uptake under flooded conditions. Genetic approaches, including Marker-Assisted Selection (MAS), have been instrumental in the development of submergence-tolerant rice varieties. The integration of the Sub1 QTL, especially the Sub1A gene, into high-yielding rice varieties has led to significant advancements in breeding submergence-tolerant cultivars. These genetic improvements provide a promising solution to mitigate the detrimental effects of flooding on rice production, ensuring the stability and sustainability of this essential crop in the face of changing climate conditions. This review discusses various mechanisms adapted by rice and genetic advancements aimed at enhancing submergence tolerance in rice to secure food production for future generations.

Transcriptome Analysis Identifies Key Genes Involved in Response and Recovery to High Heat Stress Induced by Fire in Schima superba.

Fire-resistant tree species play a crucial role in forest fire prevention, utilizing several physiological and molecular mechanisms to respond to extreme heat stress. Many transcription factors (TFs) and genes are known to be involved in the regulatory network of heat stress response in plants. However, their roles in response to high temperatures induced by fire remain less understood. In this study, we investigated Schima superba, a fire-resistant tree, to elucidate these mechanisms. Leaves of S. superba seedlings were exposed to fire stimulation for 10 s, 30 s, and 1 min, followed by a 24-h recovery period. Fifteen transcriptomes were assembled to identify key molecular and biological pathways affected by high temperatures. Differentially expressed genes (DEGs) analysis revealed essential candidate genes and TFs involved in the heat stress response, including members of the ethylene-responsive factors, WRKY, MYB, bHLH, and Nin-like families. Genes related to heat shock proteins/factors, lipid metabolism, antioxidant enzymes, dehydration responses, and hormone signal transduction were differentially expressed after heat stress and recovery, underscoring their roles in cellular process and recovery after heat stress. This study advances our understanding of plant response and defense strategies against extreme abiotic stresses.

Hypoxia represses pattern-triggered immune responses in Arabidopsis.

Biotic and abiotic stresses frequently co-occur in nature, yet relatively little is known about how plants co-ordinate the response to combined stresses. Protein degradation by the ubiquitin/proteasome system is central to the regulation of multiple independent stress response pathways in plants. The Arg/N-degron pathway is a subset of the ubiquitin/proteasome system that targets proteins based on their N termini and has been specifically implicated in the responses to biotic and abiotic stresses, including hypoxia, via accumulation of group VII ETHYLENE RESPONSE FACTOR (ERF-VII) transcription factors that orchestrate the onset of the hypoxia response program. Here, we investigated the role of the Arabidopsis (Arabidopsis thaliana) Arg/N-degron pathway in mediating the crosstalk between combined abiotic and biotic stresses using hypoxia treatments and the flg22 elicitor of pattern-triggered immunity (PTI), respectively. We uncovered a link between the plant transcriptional responses to hypoxia and flg22. Combined hypoxia and flg22 treatments showed that hypoxia represses the flg22 transcriptional program, as well as the expression of pattern recognition receptors, mitogen-activated protein kinase (MAPK) signalling and callose deposition during PTI through mechanisms that are mostly independent from the ERF-VIIs. These findings improve our understanding of the trade-offs between plant responses to combined abiotic and biotic stresses in the context of our efforts to increase crop resilience to global climate change. Our results also show that the well-known repressive effect of hypoxia on innate immunity in animals also applies to plants.

The StPti5 ethylene response factor acts as a susceptibility factor by negatively regulating the potato immune response to pathogens.

Ethylene response factors (ERFs) have been associated with biotic stress in Arabidopsis, while their function in non-model plants is still poorly understood. Here we investigated the role of potato ERF StPti5 in plant immunity. We show that StPti5 acts as a susceptibility factor. It negatively regulates potato immunity against potato virus Y and Ralstonia solanacearum, pathogens with completely different modes of action, and thereby has a different role than its orthologue in tomato. Remarkably, StPti5 is destabilised in healthy plants via the autophagy pathway and accumulates exclusively in the nucleus upon infection. We demonstrate that StEIN3 and StEIL1 directly bind the StPti5 promoter and activate its expression, while synergistic activity of the ethylene and salicylic acid pathways is required for regulated StPti expression. To gain further insight into the mode of StPti5 action in attenuating potato defence responses, we investigated transcriptional changes in salicylic acid deficient potato lines with silenced StPti5 expression. We show that StPti5 regulates the expression of other ERFs and downregulates the ubiquitin-proteasome pathway as well as several proteases involved in directed proteolysis. This study adds a novel element to the complex puzzle of immune regulation, by deciphering a two-level regulation of ERF transcription factor activity in response to pathogens.

Auxin responsive factor MdARF17 promotes ethylene synthesis in apple fruits by activating MdERF003 expression.

Auxin (AUX) promotion of apple fruit ripening is ethylene-dependent, and AUX-MdARF17-MdERF003 plays a role in AUX-promoted ethylene synthesis in apple. Phytohormones play important roles in plant growth and fleshy fruit ripening, and the phytohormone auxin (AUX) can either promote or inhibit the ripening of fleshy fruits. Although AUX can influence ethylene (ETH) synthesis in apple (Malus domestica) fruits by affecting ETH system II, this mechanism remains to be explored. Here, we identified an ETH response factor (ERF) family transcription factor, MdERF003, whose expression could be activated by naphthalene acetic acid. The transient silencing of MdERF003 inhibited ETH synthesis in fruits, and MdERF003 could bind to the MdACS1 promoter. To explore the upstream target genes of MdERF003, we screened the MdARF family members by yeast one-hybrid assays of the MdERF003 promoter, and found that the transcription factor MdARF17, which showed AUX-promoted expression, could bind to the MdERF003 promoter and promote its expression. Finally, we silenced MdERF003 in apple fruits overexpressing MdARF17 and found that MdERF003 plays a role in MdARF17-promoted ETH synthesis in apple. Thus, AUX-MdARF17-MdERF003 promotes ETH synthesis in apple fruits.

Konjac Glucomannan Oligosaccharides (KGMOS) Confers Innate Immunity against Phytophthora nicotianae in Tobacco

In this study, KGMOS (DP, 2-13), derived from KGM (Konjac glucomannan), was applied to elucidate plant immunity in a Nicotiana benthamiana Phytophthora nicotianae model. Application of KGMOS (25–100 mg/L) notably inhibited P. nicotianae, resulting in reduced disease indices and a significant accumulation of defense molecules such as H2O2 and callose. Transcriptomic analysis revealed that genes shared between KGMOS-treated and control plants are involved in signaling pathways, transcription regulation, hydrogen peroxide catabolism, and oxidative stress response. This suggests that KGMOS triggers H2O2 accumulation, callose deposition, and activation of the salicylic acid (SA) and jasmonic acid/ethylene (JA/ET) pathways after pathogen inoculation. Upregulated defense-response genes in the KGMOS group included SA-related late blight-resistant, pathogenesis-related (PR), and JA/ET-related ethylene response factor (ERF) genes. Heatmap analysis showed more upregulated defense genes (PR and NPR) related to SA in the KGMOS-treated group than in controls. RT-qPCR validation revealed significant upregulation of SA and JA/ET pathway genes in KGMOS-treated plants. Higher SA content in these plants suggests enhanced disease resistance. This study concludes that KGMOS pre-treatment induced resistance against P. nicotianae, especially at a lower concentration (25 mg/L). These findings could offer valuable insights for the future application of KGMOS in controlling plant diseases for sustainable agriculture and postharvest management.

A novel AP2/ERF transcription factor, NtERF10, positively regulates plant height in tobacco

Ethylene response factors have been shown to be involved in the effects of plant developmental processes and to regulate stress tolerance. The aim of this study was to recognize the regulatory mechanisms of ethylene response factors on tobacco plant height. In this study, a gene-edited mutant (ERF10-KO) and wild type (WT) were utilized as experimental materials. Transcriptome and metabolome analyses were used to investigate the regulatory mechanism of NtERF10 gene editing on plant height in tobacco. Here, through the analysis of differentially expressed genes (DEGs), 2051 genes were upregulated and 1965 genes were downregulated. We characterized the different ERF10-KO and WT plant heights and identified key genes for photosynthesis, the plant hormone signal transduction pathway and the terpene biosynthesis pathway. NtERF10 was found to affect the growth and development of tobacco by regulating the expression levels of the PSAA, PSBA, GLY17 and GGP3 genes. Amino acid metabolism was analyzed by combining analyses of differentially expressed genes (DEGs) and differentially accumulated metabolites (DAMs). In addition, we found that members of the bHLH, NAC, MYB, and WRKY transcription factor families have vital roles in regulating plant height. This study not only provides important insights into the positive regulation of the ethylene response factor NtERF10 on plant height during plant growth and development but also provides new research ideas for tobacco molecular breeding.

Comprehensive expression analysis of ERF transcription factors during chilling acclimation in Saintpaulia

AbstractSaintpaulia (Saintpaulia ionantha), a popular indoor ornamental potted plant, is native to the highlands of Kenya and Tanzania where temperatures rarely fall below 4 °C. Chilling injury during cultivation and transportation is a major commercial problem in Saintpaulia. In this study, we investigated chilling acclimation in Saintpaulia ‘Kilauea’. Plants grown at 20 °C (14 h light/10 h dark) displayed rapid and severe chilling injury after 24-h exposure to 4 °C. However, chilling injury at 4 °C could be dramatically reduced by pre-treating the plants at 10 °C but not at 6 °C. From whole genome analysis, 161 ethylene-responsive factors (ERFs) were identified and classified into 12 clades according to existing reports. Among these ERFs, 43, 8, and 4 ERFs were upregulated at 12, 24, and 48 h after 10 °C treatment, respectively. Most of these ERFs had GCC box and/or DRE/CRT core motifs-like sequences in their upstream regions. Finally, we compared the expression of ERFs between the treatments for 24 h at 10 °C, an effective temperature for chilling acclimation, and 6 °C, an ineffective temperature. The results showed that the expression of all six ERFs we investigated was increased by the 10 °C treatment, but not or only barely increased by the 6 °C treatment. This study suggests that Saintpaulia, a subtropical plant, can acclimate to low temperatures and that ERF upregulation is involved in chilling acclimation.

Transcriptome and weighted gene co-expression network analyses reveal key genes and pathways involved in early fruit ripening in Citrus sinensis

BackgroundThe fruit ripening period is an important target trait in fruit tree crop breeding programs. Thus, citrus tree breeders seek to develop extreme early ripening cultivars that allow optimization of citrus maturation periods. In this study, we explored the regulatory network involved in fruit ripening in Citrus sinensis using the ‘Newhall’ navel orange variety and its early-ripening mutant, ‘Gannanzao’. This research will provide a basis for further research on important signaling pathways, gene functions and variety breeding of Citrus sinensis related to fruit ripening period.ResultsPhysiological analyses suggested that early fruit ripening in ‘Gannanzao’ is regulated by early accumulation of abscisic acid (ABA), persistently high levels of jasmonic acid (JA), and higher sucrose content in the pericarp. Pericarp samples from ‘Gannanzao’ and ‘Newhall’ navel oranges were sampled for RNA sequencing analysis at 180, 200, and 220 days after flowering; 1430 differentially expressed genes (DEGs) were identified. Functional enrichment analysis indicated that these DEGs were mainly enriched in the plant hormone signal transduction and sugar metabolism pathways, as well as other pathways related to fruit ripening. Important DEGs associated with fruit ripening in ‘Gannanzao’ included genes involved in ABA and JA metabolism and signal transduction, as well as sugar metabolism. Weighted gene co-expression network analysis showed that the deep pink module had the strongest correlations with ABA content, JA content, and early ripening. Based on gene functionality and gene expression analyses of 37 genes in this module, two candidate hub genes and two ethylene response factor 13 (ERF13) genes (Cs_ont_5g000690 and Cs_ont_5g000700) were identified as key genes regulated by ABA and JA signaling. These findings will help to clarify the mechanisms that underlie early citrus fruit ripening and will lead to the development of excellent genetic resources for further breeding of extreme early-ripening varieties.ConclusionsThrough analyses of the ‘Newhall’ navel orange cultivar and its early-ripening mutant ‘Gannanzao’, we identified genes involved in ABA and JA metabolism, signal transduction, and sugar metabolism that were related to fruit ripening. Among these, two ERF13 genes were inferred to be key genes in the regulation of fruit ripening. These findings provide insights into the genetic architecture related to early fruit ripening in C. sinensis.

An AP2/ERF member LlERF012 confers thermotolerance via activation of HSF pathway in lily.

Heat stress transcription factors (HSFs) are core factors of plants in response to heat stress (HS), but their regulatory network is complicated and remains elusive in a large part, especially HSFBs. In this study, we reported that the LlERF012-LlHSFA1 module participates in heat stress response (HSR) by directly regulating HSF pathway in lily (Lilium longiflorum). LlHSFB1 was confirmed as a positive regulator in lily thermotolerance and a heat-inducible AP2/ERF member LlERF012 (Ethylene Response Factor 012) was further identified to be a direct trans-activator of LlHSFB1. Overexpression of LlERF012 elevated the thermotolerance of transgenic Arabidopsis and lily, but silencing LlERF012 reduced thermotolerance in lily. Further analysis showed LlERF012 interacted with LlHSFA1, which led to enhanced transactivation activity and DNA-binding capability of LlERF012. In addition, LlERF012 also directly activated the expression of LlHSFA1 by binding its promoter. As expected, we found that LlERF012 bound the promoters of LlHSFA2, LlHSFA3A, and LlHSFA3B to stimulate their expression, and LlERF012-LlHSFA1 interaction enhanced these activation effects. Overall, our data suggested that LlERF012 was a key factor for lily thermotolerance and the LlERF012-LlHSFA1 interaction synergistically regulated the activity of the HSF pathway including the class A and B members, which might be of great significance for coordinating the functions of different HSFs.

Genome-wide identification of the EIN3/EIL gene family in Ginkgo biloba and functional study of a GbEIL in the ethylene signaling pathway

ETHYLENE-INSENSITIVE3 (EIN3) or EIN3-Like (EIL) proteins, play critical roles in integrating ethylene signaling and physiological regulation in plants by modulating the expression of various downstream genes, such as ethylene-response factors (ERFs). However, little is known about the characteristics of EIN3/EILs in the gymnosperm Ginkgo biloba. In the present study, a genome-wide comparative analysis of Ginkgo EIN3/EIL gene family was performed with those from an array of species, including bryophytes (Physcomitrella patens), gymnosperms (Cycas panzhihuaensis), and angiosperms (Arabidopsis thaliana, Gossypium raimondii, Gossypium hirsutum, Oryza sativa, and Brachypodium distachyon). Within the constructed phylogenetic tree for the 53 EIN3/EILs identified, 5 GbEILs from G. biloba, 2 PpEILs from P. patens, and 3 CpEILs from C. panzhihuaensis were assigned to one cluster, suggesting that their derivation occurred after the split of their ancestors and angiosperms. Although considerable divergence accumulated in amino acid sequences along with the evolutionary process, the specific EIN3_DNA-binding domains were evolutionarily conserved among the 53 EIN3/EILs. Collinearity analysis indicated that whole-genome or segmental duplication and subsequent purifying selection might have prompted the generation and evolution of EIN3/EIL multigene families. Based on the expression patterns of five GbEILs at the four developmental stages of Ginkgo ovules, one GbEIL gene (Gb_03292) was further investigated for its role in mediating ethylene signaling. The functional activity of Gb_03292 was closely related to ethylene signaling, as it complemented the triple response via ectopic expression in ein3eil1 double mutant Arabidopsis. Additionally, GbEIL likely modulates the expression of a Ginkgo ERF (Gb_15517) by directly binding to its promoter. These results demonstrated that the GbEIL gene could have participated in mediating ethylene signal transduction during ovule development in G. biloba. The present study also provides insights into the conservation of ethylene signaling across the gymnosperm G. biloba and angiosperm species.

Jasmonate/ethylene- and NaWRKY6/3-regulated Alternaria resistance depends on ethylene response factor 1B-like in Nicotiana attenuata.

Biosynthesis of the phytoalexins scopoletin and scopolin in Nicotiana species is regulated by upstream signals including jasmonate (JA), ethylene (ET), and NaWRKY3 in response to the necrotrophic fungus Alternaria alternata, which causes brown spot disease. However, how these signals are coordinated to regulate these phytoalexins remains unknown. By analyzing RNA sequencing data and RNAi, we identified NaERF1B-like (NaERF1B-L) as a key player in Nicotiana attenuata during A. alternata infection by regulating the transcripts of Feruloyl-CoA 6'-hydroxylase 1 (NaF6'H1), encoding a key enzyme for scopoletin biosynthesis, and NaVS1-like (NaVS1-L), a putative biosynthetic gene of the phytoalexin solavetivone. We further demonstrated that the synergistic induction of these two genes by JA and ET signaling is mediated by NaERF1B-L. Additionally, we found that the two closely related proteins, NaWRKY6 and NaWRKY3, physically interact to enhance NaERF1B-L expression by directly binding to and activating the NaERF1B-L promoter. Collectively, our current results demonstrate that NaERF1B-L plays a positive role in resistance to A. alternata by modulating phytoalexin biosynthesis through the integration of JA/ET and NaWRKY6/3 signaling. Our findings reveal a fine-tuned transcriptional regulatory hierarchy mediated by NaERF1B-L for brown spot disease resistance in wild tobacco.