Ethylene Response Factor Family Research Articles

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.

Computational analysis of the AP2/ERF family in crops genome

BackgroundThe Apetala 2/ethylene-responsive factor family has diverse functions that enhance development and torment resistance in the plant genome. In variation, the ethylene-responsive factor (ERF) family of TF’s genes is extensive in the crop genome. Generally, the plant-specific ethylene-responsive factor family may divided by the dehydration-responsive element-binding (DREB) subfamily. So, the AP2/ERF super-family demonstrated the repeated AP2 domain during growth. The sole AP2 domain function represents abiotic stress resistance. Also, the AP2 with B3 domain enhances during the replication of brassinosteroid.ObjectiveThe study objective is to investigate the Apetala 2/ethylene-responsive factor family in a model organism of the Arabidopsis thaliana for comparative analysis towards Solanum lycopersicum (Tomato), Brassica juncea (Indian and Chinese mustard), Zea mays L. (Maize) and Oryza sativa (Indian and Japanese Rice). So, examinations of the large AP2/ERF super-family are mandatory to explore the Apetala 2 (AP2) family, ERF family, DREB subfamily, and RAV family involved during growth and abiotic stress stimuli in crops.MethodsTherefore, perform bioinformatics and computational methods to the current knowledge of the Apetala 2/ethylene-responsive factor family and their subfamilies in the crop genome. This method may be valuable for functional analysis of particular genes and their families in the plant genome.ResultsObservation data provided evidence of the Apetala 2/ethylene-responsive factor (AP2/ERF) super-family and their sub-family present in Arabidopsis thaliana (Dicots) and compared with Solanum lycopersicum (Dicots), Brassica juncea (Dicots), Zea mays L. (Monocots) and Oryza sativa (Monocots). Also, remarks genes in Oryza sativa. This report upgraded the Apetala 2/ethylene-responsive factor (AP2/ERF) family in the crop genome. So, the analysis documented the conserved domain, motifs, and phylogenetic tree towards Dicots and Monocots species. Those outcomes will be valuable for future studies of the defensive Apetala 2/ethylene-responsive factor family in crops.ConclusionTherefore, the study concluded that the several species-specific TF genes in the Apetala 2/ethylene-responsive factor (AP2/ERF) family in Arabidopsis thaliana and compared with crop-species of Solanum lycopersicum, Brassica juncea, Zea mays L. and Oryza sativa. Those plant-specific genes regulate during growth and abiotic stress control in plants.

ERF5.1 modulates carotenoid accumulation by interacting with CCD4.1 in Lycium.

Carotenoids are important natural pigments and have medical and health functions for humans. Carotenoid cleavage dioxygenase 4 (CCD4) and ethylene responsive factor (ERF) participate in carotenoid metabolism, but their roles in Lycium have not been discovered. Here, we annotated LbCCDs from the Lycium reference genome and found that LbCCD4.1 expression was significantly correlated with the carotenoid metabolites during Lycium five fruit developmental stages. Over-expression of LbCCD4.1 in NQ's leaves resulted in a series of significantly lower contents of carotenoid metabolites, including β-carotene and β-cryptoxanthin. Moreover, LbERF5.1, a transcription factor belonging to the ERF family that was located in the nucleus, was isolated. Significant reductions in the carotenoids, especially lutein, violaxanthin and their derivatives, were observed in over-expressing ERF5.1 transgenic NQ's leaves. Over-expression or virus-induced gene silencing of LbERF5.1 in NQ's leaves induced a consistent up- or down-expression, respectively, of LbCCD4.1. Furthermore, yeast one-hybrid and dual-luciferase reporter assays showed that ERF5.1 interacted with the promoter of CCD4.1 to increase its expression, and LbERF5.1 could bind to any one of the three predicted binding sites in the promoter of LbCCD4.1. A transcriptome analysis of LbERF5.1 and LbCCD4.1 over-expressed lines showed similar global transcript expression, and geranylgeranyl diphosphate synthase, phytoene synthase, lycopene δ-cyclase cytochrome, cytochrome P450-type monooxygenase 97A, cytochrome P450-type monooxygenase 97C, and zeaxanthin epoxidase in the carotenoid biosynthesis pathway were differentially expressed. In summary, we uncovered a novel molecular mechanism of carotenoid accumulation that involved an interaction between ERF5.1 and CCD4.1, which may be used to enhance carotenoid in Lycium.

ETHYLENE-INSENSITIVE 3-LIKE 2 regulates β-carotene and ascorbic acid accumulation in tomatoes during ripening.

ETHYLENE-INSENSITIVE 3/ETHYLENE-INSENSITIVE 3-LIKEs (EIN3/EILs) are important ethylene response factors during fruit ripening. Here, we discovered that EIL2 controls carotenoid metabolism and ascorbic acid (AsA) biosynthesis in tomato (Solanum lycopersicum). In contrast to the red fruits presented in the wild type (WT) 45 d after pollination, the fruits of CRISPR/Cas9 eil2 mutants and SlEIL2 RNA interference lines (ERIs) showed yellow or orange fruits. Correlation analysis of transcriptome and metabolome data for the ERI and WT ripe fruits revealed that SlEIL2 is involved in β-carotene and AsA accumulation. ETHYLENE RESPONSE FACTORs (ERFs) are the typical components downstream of EIN3 in the ethylene response pathway. Through a comprehensive screening of ERF family members, we determined that SlEIL2 directly regulates the expression of 4 SlERFs. Two of these, SlERF.H30 and SlERF.G6, encode proteins that participate in the regulation of LYCOPENE-β-CYCLASE 2 (SlLCYB2), encoding an enzyme that mediates the conversion of lycopene to carotene in fruits. In addition, SlEIL2 transcriptionally repressed L-GALACTOSE 1-PHOSPHATE PHOSPHATASE 3 (SlGPP3) and MYO-INOSITOL OXYGENASE 1 (SlMIOX1) expression, which resulted in a 1.62-fold increase of AsA via both the L-galactose and myoinositol pathways. Overall, we demonstrated that SlEIL2 functions in controlling β-carotene and AsA levels, providing a potential strategy for genetic engineering to improve the nutritional value and quality of tomato fruit.

Genome-Wide Identification of the ERF Transcription Factor Family for Structure Analysis, Expression Pattern, and Response to Drought Stress in Populus alba × Populus glandulosa.

The Ethylene Responsive Factor (ERF) transcription factor family is important for regulating plant growth and stress responses. Although the expression patterns of ERF family members have been reported in many plant species, their role in Populus alba × Populus glandulosa, an important model plant for forest research, remains unclear. In this study, we identified 209 PagERF transcription factors by analyzing the P. alba × P. glandulosa genome. We analyzed their amino acid sequences, molecular weight, theoretical pI (Isoelectric point), instability index, aliphatic index, grand average of hydropathicity, and subcellular localization. Most PagERFs were predicted to localize in the nucleus, with only a few PagERFs localized in the cytoplasm and nucleus. Phylogenetic analysis divided the PagERF proteins into ten groups, Class I to X, with those belonging to the same group containing similar motifs. Cis-acting elements associated with plant hormones, abiotic stress responses, and MYB binding sites were analyzed in the promoters of PagERF genes. We used transcriptome data to analyze the expression patterns of PagERF genes in different tissues of P. alba × P. glandulosa, including axillary buds, young leaves, functional leaves, cambium, xylem, and roots, and the results indicated that PagERF genes are expressed in all tissues of P. alba × P. glandulosa, especially in roots. Quantitative verification results were consistent with transcriptome data. When P. alba × P. glandulosa seedlings were treated with 6% polyethylene glycol 6000 (PEG6000), the results of RT-qRCR showed that nine PagERF genes responded to drought stress in various tissues. This study provides a new perspective on the roles of PagERF family members in regulating plant growth and development, and responses to stress in P. alba × P. glandulosa. Our study provides a theoretical basis for ERF family research in the future.

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.

Identification of AP2/ERF transcription factors in Tetrastigma hemsleyanum revealed the specific roles of ERF46 under cold stress.

Tetrastigma hemsleyanum (T. hemsleyanum) is a traditional medicinal plant that is widely used in China. Cultivated T. hemsleyanum usually encounters cold stress, limiting its growth and quality at key developmental stages. APETALA2 (AP2)/ethylene-responsive factor (ERF) transcription factors (TFs) comprise one of the largest gene superfamilies in plants and are widely involved in biotic and abiotic stresses. To reveal the roles of AP2/ERF TFs during T. hemsleyanum development, 70 AP2/ERF TFs were identified in T. hemsleyanum. Among them, 18 and 2 TFs were classified into the AP2 and RAV families, respectively. The other 50 TFs belonged to the ERF family and were further divided into the ERF and (dehydration reaction element binding factor) DREB subfamilies. The ERF subfamily contained 46 TFs, while the DREB subfamily contained 4 TFs. Phylogenetic analysis indicated that AP2/ERF TFs could be classified into five groups, in which 10 conserved motifs were confirmed. Several motifs were group- or subgroup-specific, implying that they were significant for the functions of the AP2/ERF TFs of these clades. In addition, 70 AP2/ERF TFs from the five groups were used for an expression pattern analysis under three low-temperature levels, namely, –4, 0, and 4°C. The majority of these AP2/ERF TFs exhibited a positive response to cold stress conditions. Specifically, ThERF5, ThERF31, ThERF46, and ThERF55 demonstrated a more sensitive response to cold stress. Moreover, AP2/ERF TFs exhibited specific expression patterns under cold stress. Transient overexpression and RNA interference indicated that ThERF46 has a specific tolerance to cold stress. These new insights provide the basis for further studies on the roles of AP2/ERF TFs in cold stress tolerance in T. hemsleyanum.

Transcriptome Level Analysis of Genes of Exogenous Ethylene Applied under Phosphorus Stress in Chinese Fir.

Chinese fir (Cunninghamia lanceolata (Lamb.) Hook) is a widely grown gymnosperm in China. Phosphorus (P) is an indispensable nutrient for the growth of Chinese fir. Inorganic phosphate (Pi) deficiency exists in soils of many Chinese fir planting area regions, and the trees themselves have limited efficiency in utilizing P from the soil. Ethylene is important in regulation responses to nutrient deficiencies. However, little is known about how ethylene signals participate in Pi stress in Chinese fir. A total of six different treatments were performed to reveal the transcript levels of Chinese fir under Pi, ethephon (an ethylene-releasing compound), and CoCl2 (cobalt chloride, an ethylene biosynthesis inhibitor) treatments. We assembled a full-length reference transcriptome containing 22,243 unigenes as a reference for UMI RNA-seq (Digital RNA-seq). There were 586 Differentially Expressed Genes (DEGs) in the Pi starvation (NP) group, while DEGs from additional ethephon or CoCl2 in NP were 708 and 292, respectively. Among the DEGs in each treatment, there were 83 TFs in these treatment groups. MYB (v-myb avian myeloblastosis viral oncogene homolog) family was the most abundant transcription factors (TFs). Three ERF (Ethylene response factor) family genes were identified when only ethylene content was imposed as a variable. Enrichment analysis indicated that the ascorbate and aldarate metabolism pathway plays a key role in resistance to Pi deficiency. This study provides insights for further elucidating the regulatory mechanism of Pi deficiency in Chinese fir.

BnERF114.A1, a Rapeseed Gene Encoding APETALA2/ETHYLENE RESPONSE FACTOR, Regulates Plant Architecture through Auxin Accumulation in the Apex in Arabidopsis.

Plant architecture is crucial for rapeseed breeding. Here, we demonstrate the involvement of BnERF114.A1, a transcription factor for ETHYLENE RESPONSE FACTOR (ERF), in the regulation of plant architecture in Brassica napus. BnERF114.A1 is a member of the ERF family group X-a, encoding a putative 252-amino acid (aa) protein, which harbours the AP2/ERF domain and the conserved CMX-1 motif. BnERF114.A1 is localised to the nucleus and presents transcriptional activity, with the functional region located at 142–252 aa of the C-terminus. GUS staining revealed high BnERF114.A1 expression in leaf primordia, shoot apical meristem, leaf marginal meristem, and reproductive organs. Ectopic BnERF114.A1 expression in Arabidopsis reduced plant height, increased branch and silique number per plant, and improved seed yield per plant. Furthermore, in Arabidopsis, BnERF114.A1 overexpression inhibited indole-3-acetic acid (IAA) efflux, thus promoting auxin accumulation in the apex and arresting apical dominance. Therefore, BnERF114.A1 probably plays an important role in auxin-dependent plant architecture regulation.

Prediction of characteristics of interactions of miRNA with mRNA of GRAS, ERF, C2H2 genes of A. thaliana, O. sativa and Z. mays

Modern methods of increasing plant productivity are based on knowledge of the molecular basis of plant growth and development. The key regulators of these processes are transcription factors (TF) that control the expression of the genome at all stages of ontogenesis. The expression of significant part of the TF genes depends on miRNAs, therefore, it is necessary to find out which miRNAs and to what extent they can affect TFs. In the present work, using the MirTarget program, we identified miRNAs capable of affecting the expression of GRAS (gibberellic-acid insensitive, repressor of gai and scarecrow), ERF (ethylene responsive factor), C2H2 (cysteine and histidine residues in their secondary structures) TF genes of A. thaliana, O. sativa, and Z. mays. The obtained characteristics of the interaction of miRNA with mRNA TF genes revealed effective associations of miRNA and TF of GRAS, ERF, and C2H2 families. During the interaction of 428 ath-miRNAs with mRNAs of 37 GRAS family genes of A. thaliana, only 11 target genes were found for eight ath-miRNAs. Of the 60 genes of the GRAS family of O. sativa, 18 genes were targets only for 16 osa-miRNAs out of 738 osa-miRNAs. Of 325 zma-miRNAs and 86 genes of the GRAS family of Z. mays, only 14 genes were targets for eight zma-miRNAs. Of 428 ath-miRNAs of A. thaliana with 123 mRNA genes of the ERF family, 25 target genes were identified for eight ath-miRNAs. Out of 738 osa-miRNAs only 13 osa-miRNAs efficiently bind to mRNA of 16 ERF genes. Of 325 zma-miRNAs with mRNAs of 186 genes of the ERF family of Z. mays, only two genes were targets for two zma-miRNAs. Of 428 ath-miRNAs and 87 genes of C2H2 family of A. thaliana, only 17 genes were targets of nine ath-miRNAs. Ten osa-miRNAs could interact with mRNA of 17 genes of the C2H2 family of O. sativa. An important role of osa-miRNA in the regulation of rice growth and development is suggested by influencing the C2H2 family TF genes of O. sativa. The ath-miR5021–5p, ath-miR5658–5p, osa-miR2102–5p, and osa-miR5075–3p interacted with several TF target genes of the C2H2, ERF, and GRAS families and had conserved binding sites that encode conserved oligopeptides.

Maize Ethylene Response Factor ZmERF061 Is Required for Resistance to Exserohilum turcicum.

Plants have evolved a series of sophisticated defense mechanisms to help them from harm. Ethylene Response Factor (ERF) plays pivotal roles in plant immune reactions, however, its underlying mechanism in maize with a defensive function to Exserohilum turcicum (E. turcicum) remains poorly understood. Here, we isolated and characterized a novel ERF transcription factor, designated ZmERF061, from maize. Phylogenetic analysis revealed that ZmERF061 is a member of B3 group in the ERF family. qRT-PCR assays showed that the expression of ZmERF061 is significantly induced by E. turcicum inoculation and hormone treatments with salicylic acid (SA) and methyl jasmonate (MeJA). ZmERF061 was proved to function as a nucleus-localized transcription activator and specifically bind to the GCC-box element. zmerf061 mutant lines resulted in enhanced susceptibility to E. turcicum via decreasing the expression of ZmPR10.1 and ZmPR10.2 and the activity of antioxidant defense system. zmerf061 mutant lines increased the expression of the SA signaling-related gene ZmPR1a and decreased the expression of the jasmonic acid (JA) signaling-related gene ZmLox1 after infection with E. turcicum. In addition, ZmERF061 could interact with ZmMPK6-1. These results suggested that ZmERF061 plays an important role in response to E. turcicum and may be useful in genetic engineering breeding.

Genome-wide identification and functional analysis of the ERF2 gene family in response to disease resistance against Stemphylium lycopersici in tomato

BackgroundAPETALA2/ethylene responsive factor (AP2/ERF) transcription factors are a plant-specific family of transcription factors and one of the largest families of transcription factors. Ethylene response factors (ERF) regulate plant growth, development, and responses to biotic and abiotic stress. In a previous study, the ERF2 gene was significantly upregulated in both resistant and susceptible tomato cultivars in response to Stemphylium lycopersici. The main purpose of this study was to systematically analyze the ERF family and to explore the mechanism of ERF2 in tomato plants resisting pathogen infection by the Virus-induced Gene Silencing technique.ResultsIn this experiment, 134 ERF genes were explored and subjected to bioinformatic analysis and divided into twelve groups. The spatiotemporal expression characteristics of ERF transcription factor gene family in tomato were diverse. Combined with RNA-seq, we found that the expression of 18 ERF transcription factors increased after inoculation with S. lycopersici. In ERF2-silenced plants, the susceptible phenotype was observed after inoculation with S. lycopersici. The hypersensitive response and ROS production were decreased in the ERF2-silenced plants. Physiological analyses showed that the superoxide dismutase, peroxidase and catalase activities were lower in ERF2-silenced plants than in control plants, and the SA and JA contents were lower in ERF2-silenced plants than in control plants after inoculation with S. lycopersici. Furthermore, the results indicated that ERF2 may directly or indirectly regulate Pto, PR1b1 and PR-P2 expression and enhance tomato resistance.ConclusionsIn this study, we identified and analyzed members of the tomato ERF family by bioinformatics methods and classified, described and analyzed these genes. Subsequently, we used VIGS technology to significantly reduce the expression of ERF2 in tomatoes. The results showed that ERF2 had a positive effect on tomato resistance to S. lycopersici. Interestingly, ERF2 played a key role in multiple SA, JA and ROS signaling pathways to confer resistance to invasion by S. lycopersici. In addition, ERF2 may directly or indirectly regulate Pto, PR1b1 and PR-P2 expression and enhance tomato resistance to S. lycopersici. In summary, this study provides gene resources for breeding for disease resistance in tomato.

Genome-Wide Analysis and the Expression Pattern of the ERF Gene Family in Hypericum perforatum.

Hypericum perforatum is a well-known medicinal herb currently used as a remedy for depression as it contains many high levels of secondary metabolites. The ethylene response factor (ERF) family encodes transcriptional regulators with multiple functions that play a vital role in the diverse developmental and physiological processes of plants, which can protect plants from various stresses by regulating the expression of genes. Although the function of several ERF genes from other plants has been further confirmed, H. perforatum is the first sequenced species in Malpighiales, and no information regarding the ERFs has been reported thus far. In this study, a total of 101 ERF genes were identified from H. perforatum. A systematic and thorough bioinformatic analysis of the ERF family was performed using the genomic database of H. perforatum. According to the phylogenetic tree analysis, HpERFs were further classified into 11 subfamilies. Gene ontology (GO) analysis suggested that most of the HpERFs likely participate in the biological processes of plants. The cis-elements were mainly divided into five categories, associated with the regulation of gene transcription, response to various stresses, and plant development. Further analysis of the expression patterns showed that the stress-responsive HpERFs responded to different treatments. This work systematically analyzed HpERFs using the genome sequences of H. perforatum. Our results provide a theoretical basis for further investigation of the function of stress-related ERFs in H. perforatum.

Expression analysis of transcription factor ERF gene family of Panax ginseng

Ethylene responsive factor(ERF), one of the largest families of transcriptional factors in plants, plays a key role in se-condary metabolism of herbal plants. To analyze the expression of ERF family genes, the heat map clustering method was used by analyzing the ginseng transcriptomes of different parts and different growth years. The contents of ginsenosides Rg_1, Re and Rb_1 in various concentrations of MeJA-treated ginseng adventitious roots were determined by UPLC-MS/MS method. The expression of key genes of ginsenoside biosynthesis(DDS, CYP716A47, CYP716A53v2) and ERF family genes in MeJA-treated ginseng adventitious roots were determined by using real-time quantitative PCR. Pearson correlation was adopted to analyze the gene expression pattern of DDS, CYP716A47, CYP716A53v2 gene and ERF family. The results showed that the content of ginseng diol ginsenoside Rb_1 in ginseng adventitious roots treated with different concentrations of MeJA increased, and the content of ginseng triol ginsenoside Rg_1 and Re decreased. It is consistent with the increase of DDS and CYP716A47 expression and the decrease of CYP716A53v2 gene expression. The expression of ERF003, ERF118 and ERF012 genes was significantly positively correlated with CYP716A53v2, but negatively correlated with DDS. While the expression of ERF1B was significantly negatively correlated with CYP716A47.It is proved that ERF003, ERF118 and ERF012 were likely to inhibit the expression of DDS and promote the expression of CYP716A53v2, and ERF1B was likely to inhibit CYP716A47. This work could provide theoretical basis of ERF functional verification of regulating the biosynthesis of ginsenosides.

Overexpression of the Watermelon Ethylene Response Factor ClERF069 in Transgenic Tomato Resulted in Delayed Fruit Ripening

Watermelon fruit undergoes distinct development stages with dramatic changes during fruit ripening. To date, the molecular mechanics of watermelon ripening remain unclear. Genetic and transcriptome evidences suggested that the ethylene response factor (ERF) gene ClERF069 may be an important candidate factor affecting watermelon fruit ripening. To dissect the roles of ClERF069 in fruit ripening, structure and phylogenetic analysis were performed using the amplified full-length sequence. Normal-ripening watermelon 97103, non-ripening watermelon PI296341-FR and the RIL population were used to analyze ClERF069 expression dynamics and the correlation with fruit ripening indexs. The results indicated that ClERF069 belongs to ERF family group VI and show high homology (83% identity) to melon ERF069-like protein. ClERF069 expression in watermelon flesh was negatively correlated with fruit lycopene content and sugar content during fruit ripening progress. Further transgenic evidences indicated that overexpression of 35S:ClERF069 in tomato noticeably delayed the ripening process up to 5.2 days. Lycopene, β-carotenoid accumulation patterns were altered and ethylene production patterns in transgenic fruits was significantly delayed during fruit ripening. Taken together, watermelon ethylene response factor ClERF069 was concluded to be a negative regulator of fruit ripening.

Molecular characterization of the ERF family in susceptible poplar infected by virulent Melampsora larici-populina

Melampsora larici-populina causes serious poplar foliar diseases called rust worldwide. The ethylene response factor (ERF) family, which is one of the largest groups of transcription factors (TFs) families, is considered to play a crucial role in regulating networks related to plant disease resistance. To investigate the transcript profiles of ERF genes involved in poplar rust resistance, Populus nigra × P. deltoides (Pnd), which is susceptible to the E4 race of M. larici-populina, was selected for this study. Digital gene expression (DGE) libraries of mixed time points of E4-inoculated (rust+) or E4-free (rust-) Pnd leaves were examined, yielding 143 Pnd-ERF genes of which 41 Pnd-ERF genes had altered transcript levels. RT-qPCR results indicated that transcript levels of these Pnd-ERF genes were mostly reduced or unchanged when plants were at 4 dpi. Then transcriptome sequencing for rust + or rust- Pnd leaves at 4 dpi was performed and interaction network of Pnd-ERFs and their target genes were predicted and analyzed. Based on the results of the transcriptome sequencing, transcript levels of 93 Pnd-ERF genes in rust + Pnd leaves were significantly different from those in rust- Pnd leaves, including 21 upregulated and 72 downregulated genes. In addition, 199 target genes were predicted to be regulated by 24 Pnd-ERFs in 216 interactions. Among them, only 31 target genes were activated, whereas 70 target genes were unchanged and the remaining 98 target genes were repressed. GO enrichment analysis indicated that most disease resistance-related target genes of Pnd-ERFs from rust + Pnd leaves were downregulated compared with their transcript levels in rust- Pnd leaves. Nevertheless, a negative regulator of plant innate immunity, the histone deacetylase 1 gene, which was targeted by a transcriptional repressor, Pnd-ERF40, was dramatically upregulated at 4 dpi. The inactivation of most Pnd-ERF genes and their disease resistance-related target genes, along with the activation of the negative regulator of plant innate immunity, at 4 dpi might account for the susceptibility of Pnd. Therefore, the inactivation of the Pnd-ERF gene network at the crucial time point should work on the breaking down of the resistance of Pnd.

Corrigendum: AP2/ERF Family Transcription Factors ORA59 and RAP2.3 Interact in the Nucleus and Function Together in Ethylene Response

[This corrects the article DOI: 10.3389/fpls.2018.01675.].

Ethylene Response Factor (ERF) Family Proteins in Abiotic Stresses and CRISPR–Cas9 Genome Editing of ERFs for Multiple Abiotic Stress Tolerance in Crop Plants: A Review

Abiotic stresses such as extreme heat, cold, drought, and salt have brought alteration in plant growth and development, threatening crop yield and quality leading to global food insecurity. Many factors plays crucial role in regulating various plant growth and developmental processes during abiotic stresses. Ethylene response factors (ERFs) are AP2/ERF superfamily proteins belonging to the largest family of transcription factors known to participate during multiple abiotic stress tolerance such as salt, drought, heat, and cold with well-conserved DNA-binding domain. Several extensive studies were conducted on many ERF family proteins in plant species through over-expression and transgenics. However, studies on ERF family proteins with negative regulatory functions are very few. In this review article, we have summarized the mechanism and role of recently studied AP2/ERF-type transcription factors in different abiotic stress responses. We have comprehensively discussed the application of advanced ground-breaking genome engineering tool, CRISPR/Cas9, to edit specific ERFs. We have also highlighted our on-going and published R&D efforts on multiplex CRISPR/Cas9 genome editing of negative regulatory genes for multiple abiotic stress responses in plant and crop models. The overall aim of this review is to highlight the importance of CRISPR/Cas9 and ERFs in developing sustainable multiple abiotic stress tolerance in crop plants.

TERF1 Regulates Nuclear Gene Expression Through Chloroplast Retrograde Signals

Ethylene is an important phytohormone that regulates many important biological processes in plant. ERF (ethylene response factor) proteins are key transcription factors that activate the ethylene signaling pathway. However, our knowledge about the mechanism of the ERF transcription factors in regulating nuclear genes expression is limited. Retrograde signaling pathway in chloroplast is a novel kind of mechanism that regulates nuclear gene expression by different signals in plastid. Based on our former research we analyzed the components related with retrograde signaling from plastid in the transgenic tobacco overexpressing TERF1, a member of ERF family transcription factors, to elucidate the interaction between ethylene signaling pathway and different retrograde signaling pathway in plastid under normal growth condition. Results show that TERF1 regulates different retrograde signals in plastid and thus regulates different nuclear genes expression under normal growth condition. We propose a new mechanism that links ethylene signaling pathway and retrograde signaling pathway as well as new potential of TERF1 in regulating nuclear genes expression at post-transcriptional level.

AP2/ERF Family Transcription Factors ORA59 and RAP2.3 Interact in the Nucleus and Function Together in Ethylene Responses.

The gaseous plant hormone ethylene is a key signaling molecule regulating plant growth, development, and defense against pathogens. Octadecanoid-responsive arabidopsis 59 (ORA59) is an ethylene response factor (ERF) transcription factor and has been suggested to integrate ethylene and jasmonic acid signaling and regulate resistance to necrotrophic pathogens. Here we screened for ORA59 interactors using the yeast two-hybrid system to elucidate the molecular function of ORA59. This led to the identification of RELATED TO AP2.3 (RAP2.3), another ERF transcription factor belonging to the group VII ERF family. In binding assays, ORA59 and RAP2.3 interacted in the nucleus and showed ethylene-dependent nuclear localization. ORA59 played a positive role in ethylene-regulated responses, including the triple response, featured by short, thick hypocotyl and root, and exaggerated apical hook in dark-grown seedlings, and resistance to the necrotrophic pathogen Pectobacterium carotovorum, as shown by the increased and decreased ethylene sensitivity and disease resistance in ORA59-overexpressing (ORA59OE) and null mutant (ora59) plants, respectively. In genetic crosses, ORA59OE rap2.3 crossed lines lost ORA59-mediated positive effects and behaved like rap2.3 mutant. These results suggest that ORA59 physically interacts with RAP2.3 and that this interaction is important for the regulatory roles of ORA59 in ethylene responses.