Comprehensive Expression Analysis of the WRKY Gene Family in Phoebe bournei under Drought and Waterlogging Stresses.

In response to biotic and abiotic stresses, the WRKY gene family plays a crucial role in plant growth and development. This study focused on Phoebe bournei and involved genome-wide identification of WRKY gene family members, clarification of their molecular evolutionary characteristics, and comprehensive mapping of their expression profiles under diverse abiotic stress conditions. A total of 60 WRKY gene family members were identified, and their phylogenetic classification revealed three distinct groups. A conserved motif analysis underscored the significant conservation of motif 1 and motif 2 among the majority of PbWRKY proteins, with proteins within the same class sharing analogous gene structures. Furthermore, an examination of cis-acting elements and protein interaction networks revealed several genes implicated in abiotic stress responses in P. bournei. Transcriptomic data were utilized to analyze the expression patterns of WRKY family members under drought and waterlogged conditions, with subsequent validation by quantitative real-time PCR (RT-qPCR) experiments. Notably, PbWRKY55 exhibited significant expression modulation under drought stress; PbWRKY36 responded prominently to waterlogging stress; and PbWRKY18, PbWRKY38, and PbWRKY57 demonstrated altered expression under both drought and waterlogging stresses. This study revealed the PbWRKY candidate genes that potentially play a pivotal role in enhancing abiotic stress resilience in P. bournei. The findings have provided valuable insights and knowledge that can guide further research aimed at understanding and addressing the impacts of abiotic stress within this species.

WRKY Transcription Factor Responses and Tolerance to Abiotic Stresses in Plants.

Plants are subjected to abiotic stresses throughout their developmental period. Abiotic stresses include drought, salt, heat, cold, heavy metals, nutritional elements, and oxidative stresses. Improving plant responses to various environmental stresses is critical for plant survival and perpetuation. WRKY transcription factors have special structures (WRKY structural domains), which enable the WRKY transcription factors to have different transcriptional regulatory functions. WRKY transcription factors can not only regulate abiotic stress responses and plant growth and development by regulating phytohormone signalling pathways but also promote or suppress the expression of downstream genes by binding to the W-box [TGACCA/TGACCT] in the promoters of their target genes. In addition, WRKY transcription factors not only interact with other families of transcription factors to regulate plant defence responses to abiotic stresses but also self-regulate by recognising and binding to W-boxes in their own target genes to regulate their defence responses to abiotic stresses. However, in recent years, research reviews on the regulatory roles of WRKY transcription factors in higher plants have been scarce and shallow. In this review, we focus on the structure and classification of WRKY transcription factors, as well as the identification of their downstream target genes and molecular mechanisms involved in the response to abiotic stresses, which can improve the tolerance ability of plants under abiotic stress, and we also look forward to their future research directions, with a view of providing theoretical support for the genetic improvement of crop abiotic stress tolerance.

Arabidopsis WRKY1 promotes monocarpic senescence by integrative regulation of flowering, leaf senescence and nitrogen remobilization.

Monocarpic senescence, characterized by whole plant senescence following a single flowering phase, is widespread in seed plants, particularly in crops, determining seed harvest time and quality. However, how external and internal signals are systemically integrated into monocarpic senescence remains largely unknown. Here, we disclose that Arabidopsis thaliana transcription factor WRKY1 plays essential roles in multiple key steps of monocarpic senescence. WRKY1 expression is induced by age, SA, and nitrogen (N) deficiency. Flowering and leaf senescence are promoted in the WRKY1 overexpression lines but are delayed in the wrky1 mutants. The joint analyses of DAP-Seq and RNA-Seq uncover the direct target genes of WRKY1. Further studies show that WRKY1 coordinately regulates three processesin monocarpic senescence: 1) suppressing FLOWERING LOCUS C gene expression to initiate flowering; 2) inducing SA biosynthesis genes to promote leaf senescence; and 3) activating the N-assimilation and transport genes to trigger N remobilization. In summary, we reveal how one stress-responsive transcription factor, WRKY1, synergistically integrates flowering and leaf senescence into monocarpic senescence, providing important insights into plant lifetime regulation.

Genome-Wide Identification of the WRKY Gene Family in Four Cotton Varieties and the Positive Role of GhWRKY31 in Response to Salt and Drought Stress.

The WRKY gene family is ubiquitously distributed in plants, serving crucial functions in stress responses. Nevertheless, the structural organization and evolutionary dynamics of WRKY genes in cotton have not been fully elucidated. In this study, a total of 112, 119, 217, and 222 WRKY genes were identified in Gossypium arboreum, Gossypium raimondii, Gossypium hirsutum, and Gossypium barbadense, respectively. These 670 WRKY genes were categorized into seven distinct subgroups and unequally distributed across chromosomes. Examination of conserved motifs, domains, cis-acting elements, and gene architecture collectively highlighted the evolutionary conservation and divergence within the WRKY gene family in cotton. Analysis of synteny and collinearity further confirmed instances of expansion, duplication, and loss events among WRKY genes during cotton evolution. Furthermore, GhWRKY31 transgenic Arabidopsis exhibited heightened germination rates and longer root lengths under drought and salt stress. Silencing GhWRKY31 in cotton led to reduced levels of ABA, proline, POD, and SOD, along with downregulated expression of stress-responsive genes. Yeast one-hybrid and molecular docking assays confirmed the binding capacity of GhWRKY31 to the W box of GhABF1, GhDREB2, and GhRD29. The findings collectively offer a systematic and comprehensive insight into the evolutionary patterns of cotton WRKYs, proposing a suitable regulatory framework for developing cotton cultivars with enhanced resilience to drought and salinity stress.

Overexpression of AcWRKY31 Increases Sensitivity to Salt and Drought and Improves Tolerance to Mealybugs in Pineapple.

Pineapple is a globally significant tropical fruit, but its cultivation faces numerous challenges due to abiotic and biotic stresses, affecting its quality and quantity. WRKY transcription factors are known regulators of stress responses, however, their specific functions in pineapple are not fully understood. This study investigates the role of AcWRKY31 by overexpressing it in pineapple and Arabidopsis. Transgenic pineapple lines were obtained using Agrobacterium-mediated transformation methods and abiotic and biotic stress treatments. Transgenic AcWRKY31-OE pineapple plants showed an increased sensitivity to salt and drought stress and an increased resistance to biotic stress from pineapple mealybugs compared to that of WT plants. Similar experiments in AcWRKY31-OE, AtWRKY53-OE, and the Arabidopsis Atwrky53 mutant were performed and consistently confirmed these findings. A comparative transcriptomic analysis revealed 5357 upregulated genes in AcWRKY31-OE pineapple, with 30 genes related to disease and pathogen response. Notably, 18 of these genes contained a W-box sequence in their promoter region. A KEGG analysis of RNA-Seq data showed that upregulated DEG genes are mostly involved in translation, protein kinases, peptidases and inhibitors, membrane trafficking, folding, sorting, and degradation, while the downregulated genes are involved in metabolism, protein families, signaling, and cellular processes. RT-qPCR assays of selected genes confirmed the transcriptomic results. In summary, the AcWRKY31 gene is promising for the improvement of stress responses in pineapple, and it could be a valuable tool for plant breeders to develop stress-tolerant crops in the future.

Hyperspectral imaging of liverwort Marchantia polymorpha identifies MpWRKY10 as a key regulator defining Foliar pigmentation patterns.

Foliar pigmentation patterns vary among plant species and growth conditions. In this study, we utilize hyperspectral imaging to assess foliar pigmentation in the bryophyte Marchantia polymorpha under nutrient stress and identify associated genetic factors. Using singular value decomposition (SVD) for feature selection, we quantitate color variations induced by deficiencies in phosphate, nitrate, magnesium, calcium, and iron. Pseudo-colored thallus images show that disrupting MpWRKY10 causes irregular pigmentation with auronidin accumulation. Transcriptomic profiling shows that MpWRKY10 regulates phenylpropanoid pathway enzymes and R2R3-MYB transcription factors during phosphate deficiency, with MpMYB14 upregulation preceding pigment accumulation. MpWRKY10 is downregulated in older, pigmented thalli under phosphate deficiency but maintained in young thalli, where it suppresses pigmentation genes. This downregulation is absent in pigmented thalli due to aging. Comparative transcriptome analysis suggests similar WRKY and MYB roles in nutrient response and pigmentation in red-leaf lettuce, alluding to conserved genetic factors controlling foliar pigmentation patterns under nutrient deficiency.

Transcription factors involved in plant responses to cadmium-induced oxidative stress.

Cadmium (Cd) is a heavy metal highly toxic to living organisms. Cd pollution of soils has become a serious problem worldwide, posing a severe threat to crop production and human health. When plants are poisoned by Cd, their growth and development are inhibited, chloroplasts are severely damaged, and respiration and photosynthesis are negatively affected. Therefore, elucidating the molecular mechanisms that underlie Cd tolerance in plants is important. Transcription factors can bind to specific plant cis-acting genes. Transcription factors are frequently reported to be involved in various signaling pathways involved in plant growth and development. Their role in the resistance to environmental stress factors, particularly Cd, should not be underestimated. The roles of several transcription factor families in the regulation of plant resistance to Cd stress have been widely demonstrated. In this review, we summarize the mechanisms of five major transcription factor families-WRKY, ERF, MYB, bHLH, and bZIP-in plant resistance to Cd stress to provide useful information for using molecular techniques to solve Cd pollution problems in the future.

WRKY transcription factors modulate flowering time in four Arachis species: a bioinformatics analysis.

BACKGROUND: WRKY proteins are important transcription factors (TFs) in plants, involved in growth and development and responses to environmental changes. Although WRKY TFs have been studied at the genome level in Arachis genus, including oil crop and turfgrass, their regulatory networks in controlling flowering time remain unclear. The aim of this study was to predict the molecular mechanisms of WRKY TFs regulation flowering time in Arachis genus at the genome level using bioinformatics approaches. RESULTS: The flowering-time genes of Arachis genus were retrieved from the flowering-time gene database. The regulatory networks between WRKY TFs and downstream genes in Arachis genus were predicted using bioinformatics tools. The results showed that WRKY TFs were involved in aging, autonomous, circadian clock, hormone, photoperiod, sugar, temperature, and vernalization pathways to modulate flowering time in Arachis duranensis, Arachis ipaensis, Arachis monticola, and Arachis hypogaea cv. Tifrunner. The WRKY TF binding sites in homologous flowering-time genes exhibited asymmetric evolutionary pattern, indicating that the WRKY TFs interact with other transcription factors to modulate flowering time in the four Arachis species. Protein interaction network analysis showed that WRKY TFs interacted with FRUITFULL and APETALA2 to modulate flowering time in the four Arachis species. WRKY TFs implicated in regulating flowering time had low expression levels, whereas their interaction proteins had varying expression patterns in 22 tissues of A. hypogaea cv. Tifrunner. These results indicate that WRKY TFs exhibit antagonistic or synergistic interactions with the associated proteins. CONCLUSIONS: This study reveals complex regulatory networks through which WRKY TFs modulate flowering time in the four Arachis species using bioinformatics approaches.

Transcription factor StWRKY1 is involved in monoterpene biosynthesis induced by light intensity in Schizonepeta tenuifolia Briq.

Menthone-type monoterpenes are the main active ingredients of Schizonepeta tenuifolia Briq. Previous studies have indicated that light intensity influences the synthesis of menthone-type monoterpenes in S. tenuifolia, but the mechanism remains unclear. WRKY transcription factors play a crucial role in plant metabolism, yet their regulatory mechanisms in S. tenuifolia are not well understood. In this study, transcriptome data of S. tenuifolia leaves under different light intensities were analyzed, identifying 57 candidate transcription factors that influence monoterpene synthesis. Among these, 7 members of the StWRKY gene family were identified and mapped onto chromosomes using bioinformatics methods. The physicochemical properties of the proteins encoded by these StWRKY genes, their gene structures, and cis-acting elements were also studied. Comparative genomics and phylogenetic analyses revealed that Sch000013479 is closely related to AaWRKY1, AtWRKY41, and AtWRKY53, and it was designated as StWRKY1. Upon silencing and overexpressing the StWRKY1 transcription factor in S. tenuifolia leaves, changes in the expression of key genes in the menthone-type monoterpene synthesis pathway were observed. Specifically, when StWRKY1 was effectively silenced, the content of (-)-pulegone significantly decreased. These results enhance our understanding of the impact of StWRKYs on monoterpene synthesis in S. tenuifolia and lay the groundwork for further exploration of the regulatory mechanisms involved in the biosynthesis of menthone-type monoterpenes.

ZmWRKY30 modulates drought tolerance in maize by influencing myo-inositol and reactive oxygen species homeostasis.

Maize (Zea mays L.) is an important food crop with a wide range of uses in both industry and agriculture. Drought stress during its growth cycle can greatly reduce maize crop yield and quality. However, the molecular mechanisms underlying maize responses to drought stress remain unclear. In this work, a WRKY transcription factor-encoding gene, ZmWRKY30, from drought-treated maize leaves was screened out and characterized. ZmWRKY30 gene expression was induced by dehydration treatments. The ZmWRKY30 protein localized to the nucleus and displayed transactivation activity in yeast. Compared with wild-type (WT) plants, Arabidopsis lines overexpressing ZmWRKY30 exhibited a significantly enhanced drought stress tolerance, as evidenced by the improved survival rate, increased antioxidant enzyme activity by superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), elevated proline content, and reduced lipid peroxidation recorded after drought stress treatment. In contrast, the mutator (Mu)-interrupted ZmWRKY30 homozygous mutant (zmwrky30) was more sensitive to drought stress than its null segregant (NS), characterized by the decreased survival rate, reduced antioxidant enzyme activity (SOD, POD, and CAT) and proline content, as well as increased malondialdehyde accumulation. RNA-Seq analysis further revealed that, under drought conditions, the knockout of the ZmWRKY30 gene in maize affected the expression of genes involved in reactive oxygen species (ROS), proline, and myo-inositol metabolism. Meanwhile, the zmwrky30 mutant exhibited significant downregulation of myo-inositol content in leaves under drought stress. Combined, our results suggest that ZmWRKY30 positively regulates maize responses to water scarcity. This work provides potential target genes for the breeding of drought-tolerant maize.

Tuber transcriptome analysis reveals a novel WRKY transcription factor StWRKY70 potentially involved in potato pigmentation.

Tuber flesh pigmentation, conferred by the presence of secondary metabolite anthocyanins, is one of many key agronomic traits for potato tubers. Although several genes of potato anthocyanin biosynthesis have been reported, transcription factors (TFs) contributing to tuber flesh pigmentation are still not fully understood. In this study, transcriptomic profiling of diploid potato accessions with or without tuber flesh pigmentation was conducted and genes of the anthocyanin biosynthesis pathway were found significantly enriched within the 1435 differentially expressed genes (DEGs). Weighted Gene Co-expression Network Analysis (WGCNA) and connectivity analysis pinpointed a subset of 173 genes closely related to the key biosynthetic gene StDFR. Of the eight transcription factors in the subset, group III WRKY StWRKY70, was chosen for showing high connectivity to StDFR and ten other anthocyanin biosynthetic genes and homology to known WRKYs of anthocyanin pathway. The transient activation assay showed StWRKY70 predominantly stimulated the expression of StDFR and StANS as well as the accumulation of anthocyanins by enhancing the function of the MYB transcription factor StAN1. Furthermore, the interaction between StWRKY70 and StAN1 was verified by Y2H and BiFC. Our analysis discovered a new transcriptional activator StWRKY70 which potentially involved in tuber flesh pigmentation, thus may lay the foundation for deciphering how the WRKY-MYB-bHLH-WD40 (WRKY-MBW) complex regulate the accumulation of anthocyanins and provide new strategies to breed for more nutritious potato varieties with enhanced tuber flesh anthocyanins.

WRKY47 transcription factor modulates leaf senescence through regulating PCD-associated genes in Arabidopsis.

Transcription factors play crucial roles in almost all physiological processes including leaf senescence. Cell death is a typical symptom appearing in senescing leaves, which is also classified as developmental programmed cell death (PCD). However, the link between PCD and leaf senescence still remains unclear. Here, we found a WRKY transcription factor WRKY47 positively modulates age-dependent leaf senescence in Arabidopsis (Arabidopsis thaliana). WRKY47 was expressed preferentially in senescing leaves. A subcellular localization assay indicated that WRKY47 was exclusively localized in nuclei. Overexpression of WRKY47 showed precocious leaf senescence, with less chlorophyll content and higher electrolyte leakage, but loss-of-function mutants of WRKY47 delayed this biological process. Through qRT-PCR and dual luciferase reporter assays, we found that WRKY47 could activate the expression of senescence-associated genes (SAGs) and PCD-associated genes to regulate leaf senescence. Furthermore, through electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-qPCR, WRKY47 was found to bind to W-box fragments in promoter regions of BFN1 (Bifunctional Nuclease 1) and MC6 (Metacaspase 6) directly. In general, our research revealed that WRKY47 regulates age-dependent leaf senescence by activating the transcription of two PCD-associated genes.

OsWRKY12 negatively regulates the drought-stress tolerance and secondary cell wall biosynthesis by targeting different downstream transcription factor genes in rice.

With the increasing occurrence of global warming, drought is becoming a major constraint for plant growth and crop yield. Plant cell walls experience continuous changes during the growth, development, and in responding to stressful conditions. The plant WRKYs play pivotal roles in regulating the secondary cell wall (SCW) biosynthesis and helping plant defend against abiotic stresses. qRT-PCR evidence showed that OsWRKY12 was affected by drought and ABA treatments. Over-expression of OsWRKY12 decreased the drought tolerance of the rice transgenics at the germination stage and the seedling stage. The transcription levels of drought-stress-associated genes as well as those genes participating in the ABA biosynthesis and signaling were significantly different compared to the wild type (WT). Our results also showed that less lignin and cellulose were deposited in the OsWRKY12-overexpressors, and heterogenous expression of OsWRKY12 in atwrky12 could lower the increased lignin and cellulose contents, as well as the improved PEG-stress tolerance, to a similar level as the WT. qRT-PCR results indicated that the transcription levels of all the genes related to lignin and cellulose biosynthesis were significantly decreased in the rice transgenics than the WT. Further evidence from yeast one-hybrid assay and the dual-luciferase reporter system suggested that OsWRKY12 could bind to promoters of OsABI5 (the critical component of the ABA signaling pathway) and OsSWN3/OsSWN7 (the key positive regulators in the rice SCW thickening), and hence repressing their expression. In conclusion, OsWRKY12 mediates the crosstalk between SCW biosynthesis and plant stress tolerance by binding to the promoters of different downstream genes.

A comprehensive analysis of the WRKY family in soybean and functional analysis of GmWRKY164-GmGSL7c in resistance to soybean mosaic virus.

BACKGROUND: Soybean mosaic disease caused by soybean mosaic virus (SMV) is one of the most devastating and widespread diseases in soybean producing areas worldwide. The WRKY transcription factors (TFs) are widely involved in plant development and stress responses. However, the roles of the GmWRKY TFs in resistance to SMV are largely unclear. RESULTS: Here, 185 GmWRKYs were characterized in soybean (Glycine max), among which 60 GmWRKY genes were differentially expressed during SMV infection according to the transcriptome data. The transcriptome data and RT-qPCR results showed that the expression of GmWRKY164 decreased after imidazole treatment and had higher expression levels in the incompatible combination between soybean cultivar variety Jidou 7 and SMV strain N3. Remarkably, the silencing of GmWRKY164 reduced callose deposition and enhanced virus spread during SMV infection. In addition, the transcript levels of the GmGSL7c were dramatically lower upon the silencing of GmWRKY164. Furthermore, EMSA and ChIP-qPCR revealed that GmWRKY164 can directly bind to the promoter of GmGSL7c, which contains the W-box element. CONCLUSION: Our findings suggest that GmWRKY164 plays a positive role in resistance to SMV infection by regulating the expression of GmGSL7c, resulting in the deposition of callose and the inhibition of viral movement, which provides guidance for future studies in understanding virus-resistance mechanisms in soybean.

The WRKY gene family in the halophyte Limonium bicolor: identification, expression analysis, and regulation of salt stress tolerance.

KEY MESSAGE: 63 L. bicolor WRKY genes were identified and their informatics was analyzed. The results suggested that the LbWRKY genes involved in the development and salt secretion of salt glands in L. bicolor. Salt stress, as a universal abiotic stress, severely inhibits the growth and development of plants. WRKY transcription factors play a vital role in plant growth and development, as well as in response to various stresses. Nevertheless, little is known of systematic genome-wide analysis of the WRKY genes in Limonium bicolor, a model recretohalophyte. In this study, 63 L. bicolor WRKY genes were identified (LbWRKY1-63), which were unevenly distributed across seven chromosomes and one scaffold. Based on the structural and phylogenetic characteristics, 63 LbWRKYs are divided into three main groups. Cis-elements in the LbWRKY promoters were related to growth and development, phytohormone responses, and stress responses. Colinearity analysis showed strong colinearity between LbWRKYs and GmWRKYs from soybean (Glycine max). Therefore, LbWRKY genes maybe have similar functions to GmWRKY genes. Expression analysis showed that 28 LbWRKY genes are highly expressed in roots, 9 in stems, 26 in leaves, and 12 in flowers and most LbWRKY genes responded to NaCl, ABA, and PEG6000. Silencing LbWRKY10 reduced salt gland density and salt secretion ability of leaves, and the salt tolerance of the species. Consistent with this, genes associated with salt gland development were markedly down-regulated in the LbWRKY10-silenced lines. Our findings suggested that the LbWRKY genes involved in the development and salt secretion of salt glands in L. bicolor. Our research provides new insights into the functions of the WRKY family in halophytes.

Unlocking nature's secrets: The pivotal role of WRKY transcription factors in plant flowering and fruit development.

The WRKY transcription factor family is a key player in the regulatory mechanisms of flowering plants, significantly influencing both their biotic and abiotic response systems as well as being vital to numerous physiological and biological functions. Over the past two decades, the functionality of WRKY proteins has been the subject of extensive research in over 50 plant species, with a strong focus on their roles in responding to various stresses. Despite this extensive research, there remains a notable gap in comprehensive studies aimed at understanding how specific WRKY genes directly influence the timing of flowering and fruit development. This review offers an up-to-date look at WRKY family genes and provides insights into the key genes of WRKY to control flowering, enhance fruit ripening and secondary metabolism synthesis, and maintain fruit quality of various plants, including annuals, perennials, medicinal, and crop plants. The WRKY transcription factors serve as critical regulators within the transcriptional regulatory network, playing a crucial role in the precise enhancement of flowering processes. It is also involved in the up-regulation of fruit ripening was strongly demonstrated by combined transcriptomics and metabolomic investigation. Therefore, we speculated that the WRKY family is known to be a key regulator of flowering and fruiting in plants. This detailed insight will enable the identification of the series of molecular occurrences featuring WRKY proteins throughout the stages of flowering and fruiting.

A Functional InDel in the WRKY10 Promoter Controls the Degree of Flesh Red Pigmentation in Apple.

MYB transcription factors have been linked to anthocyanin synthesis and various color phenotypes in plants. In apple, MYB10 confers a red-flesh phenotype due to a minisatellite insertion in its R6 promoter, but R6:MYB10 genotypes exhibit various degrees of red pigmentation in the flesh, suggesting the involvement of other genetic factors. Here, it is shown that MdWRKY10, a transcription factor identified via DNA pull-down trapping, binds to the promoter of MdMYB10 and activates its transcription. MdWRKY10 specifically interacts with the WDR protein MdTTG1 to join the apple MYB-bHLH-WDR (MBW) complex, which significantly enhances its transcriptional activation activity. A 163-bp InDel detected in the promoter region of the alleles of MdWRKY10 in a hybrid population of identical heterozygous genotypes regarding R6 by structural variation analysis, contains a typical W-box element that MdWRKY10 binds to for transactivation. This leads to increased transcript levels of MdWRKY10 and MdMYB10 and enhanced anthocyanin synthesis in the flesh, largely accounting for the various degrees of flesh red pigmentation in the R6 background. These findings reveal a novel regulatory role of the WRKY-containing protein complex in the formation of red flesh apple phenotypes and provide broader insights into the molecular mechanism governing anthocyanin synthesis in plants.

Genome-wide profiling of WRKY genes involved in flavonoid biosynthesis in Erigeron breviscapus.

The transcription factors of WRKY genes play essential roles in plant growth, stress responses, and metabolite biosynthesis. Erigeron breviscapus, a traditional Chinese herb, is abundant in flavonoids and has been used for centuries to treat cardiovascular and cerebrovascular diseases. However, the WRKY transcription factors that regulate flavonoid biosynthesis in E. breviscapus remain unknown. In this study, a total of 75 EbWRKY transcription factors were predicted through comprehensive genome-wide characterization of E. breviscapus and the chromosomal localization of each EbWRKY gene was investigated. RNA sequencing revealed transient responses of 74 predicted EbWRKY genes to exogenous abscisic acid (ABA), salicylic acid (SA), and gibberellin 3 (GA3) after 4 h of treatment. In contrast, the expression of key structural genes involved in flavonoid biosynthesis increased after 4 h in GA3 treatment. However, the content of flavonoid metabolites in leaves significantly increased at 12 h. The qRT-PCR results showed that the expression patterns of EbWRKY11, EbWRKY30, EbWRKY31, EbWRKY36, and EbWRKY44 transcription factors exhibited a high degree of similarity to the 11 structural genes involved in flavonoid biosynthesis. Protein-DNA interactions were performed between the key genes involved in scutellarin biosynthesis and candidate WRKYs. The result showed that F7GAT interacts with EbWRKY11, EbWRKY36, and EbWRKY44, while EbF6H has a self-activation function. This study provides comprehensive information on the regulatory control network of flavonoid accumulation mechanisms, offering valuable insights for breeding E. breviscapus varieties with enhanced scutellarin content.

Comparative Transcriptome Analysis Revealed Candidate Gene Modules Involved in Salt Stress Response in Sweet Basil and Overexpression of ObWRKY16 and ObPAL2 Enhanced Salt Tolerance of Transgenic Arabidopsis.

Sweet basil (Ocimum basilicum L.) is an important aromatic plant with high edibility and economic value, widely distributed in many regions of the tropics including the south of China. In recent years, environmental problems, especially soil salinization, have seriously restricted the planting and spread of sweet basil. However, the molecular mechanism of the salt stress response in sweet basil is still largely unknown. In this study, seed germination, seedling growth, and chlorophyll synthesis in sweet basil were inhibited under salt stress conditions. Through comparative transcriptome analysis, the gene modules involved in the metabolic processes, oxidative response, phytohormone signaling, cytoskeleton, and photosynthesis were screened out. In addition, the landscape of transcription factors during salt treatment in sweet basil was displayed as well. Moreover, the overexpression of the WRKY transcription factor-encoding gene, ObWRKY16, and the phenylalanine ammonia-lyase-encoding gene, ObPAL2, enhanced the seed germination, seedling growth, and survival rate, respectively, of transgenic Arabidopsis, suggesting that they might be important candidates for the creation of salt-tolerant sweet basil cultivars. Our data enrich the study on salt responses in sweet basil and provide essential gene resources for genetic improvements in sweet basil in the future.

PtWRKY2, a WRKY transcription factor from Pinellia ternata confers heat tolerance in Arabidopsis.

High temperatures are a major stress factor that limit the growth of Pinellia ternata. WRKY proteins widely distribute in plants with the important roles in plant growth and stress responses. However, WRKY genes have not been identified in P. ternata thus far. In this study, five PtWRKYs with four functional subgroups were identified in P. ternata. One group III WRKY transcription factor, PtWRKY2, was strongly induced by high temperatures, whereas the other four PtWRKYs were suppressed. Analysis of transcription factor characteristics revealed that PtWRKY2 localized to the nucleus and specifically bound to W-box elements without transcriptional activation activity. Overexpression of PtWRKY2 increased the heat tolerance of Arabidopsis, as shown by the higher percentage of seed germination and survival rate, and the longer root length of transgenic lines under high temperatures compared to the wild-type. Moreover, PtWRKY2 overexpression significantly decreased reactive oxygen species accumulation by increasing the catalase, superoxide dismutase, and peroxidase activities. Furthermore, the selected heat shock-associated genes, including five transcription factors (HSFA1A, HSFA7A, bZIP28, DREB2A, and DREB2B), two heat shock proteins (HSP70 and HSP17.4), and three antioxidant enzymes (POD34, CAT1, and SOD1), were all upregulated in transgenic Arabidopsis. The study identifies that PtWRKY2 functions as a key transcriptional regulator in the heat tolerance of P. ternata, which might provide new insights into the genetic improvement of P. ternata.