WRKY Domain Research Articles

Genome-wide identification and characterization of the Populus WRKY transcription factor family and analysis of their expression in response to biotic and abiotic stresses.

WRKY proteins are a large family of regulators involved in various developmental and physiological processes, especially in coping with diverse biotic and abiotic stresses. In this study, 100 putative PtrWRKY genes encoded the proteins contained in the complete WRKY domain in Populus. Phylogenetic analysis revealed that the members of this superfamily among poplar, Arabidopsis, and other species were divided into three groups with several subgroups based on the structures of the WRKY protein sequences. Various cis-acting elements related to stress and defence responses were found in the promoter regions of PtrWRKY genes by promoter analysis. High-throughput transcriptomic analyses identified that 61 of the PtrWRKY genes were induced by biotic and abiotic treatments, such as Marssonina brunnea, salicylic acid (SA), methyl jasmonate (MeJA), wounding, cold, and salinity. Among these PtrWRKY genes, transcripts of 46 selected genes were observed in different tissues, including roots, stems, and leaves. Quantitative RT-PCR analysis further confirmed the induced expression of 18 PtrWRKY genes by one or more stress treatments. The overexpression of an SA-inducible gene, PtrWRKY89, accelerated expression of PR protein genes and improved resistance to pathogens in transgenic poplar, suggesting that PtrWRKY89 is a regulator of an SA-dependent defence-signalling pathway in poplar. Taken together, our results provided significant information for improving the resistance and stress tolerance of woody plants.

Phylogenetic analysis of barley WRKY proteins and characterization of HvWRKY1 and -2 as repressors of the pathogen-inducible gene HvGER4c.

The WRKY transcription factors belong to an evolutionarily conserved superprotein family predominantly present in the plant kingdom. WRKY proteins of barley are not yet fully annotated and most of them are not functionally characterized. We performed a genome-wide identification of WRKY members based on the recently accessible barley draft genome sequence and full-length cDNA datasets. As a result, 34 novel putative proteins have been identified which extend the existing list for barley WRKYs to 94. Phylogenetic analysis of the WRKY domains allowed ranking into three groups (I, II, III), with an expansion in group III in monocots. Two members of subgroup IIa, the wound and pathogen-inducible HvWRKY1 and HvWRKY2, are known as negative defense regulators. Here, we demonstrate that both transcription factors repress the activity of the powdery mildew-induced promoter of HvGER4c, a germin-like defense-related protein. The repression did not require the negative defense regulator MLO nor was it affected by the presence of the R protein MLA12. Moreover, the expression of the Arabidopsis ortholog AtWRKY40 in barley compromised basal resistance to powdery mildew, providing evidence for functional conservation of sequence-related WRKY proteins across monocots and dicots.

Genome-wide analysis of the WRKY gene family in cotton.

WRKY proteins are major transcription factors involved in regulating plant growth and development. Although many studies have focused on the functional identification of WRKY genes, our knowledge concerning many areas of WRKY gene biology is limited. For example, in cotton, the phylogenetic characteristics, global expression patterns, molecular mechanisms regulating expression, and target genes/pathways of WRKY genes are poorly characterized. Therefore, in this study, we present a genome-wide analysis of the WRKY gene family in cotton (Gossypium raimondii and Gossypium hirsutum). We identified 116 WRKY genes in G. raimondii from the completed genome sequence, and we cloned 102 WRKY genes in G. hirsutum. Chromosomal location analysis indicated that WRKY genes in G. raimondii evolved mainly from segmental duplication followed by tandem amplifications. Phylogenetic analysis of alga, bryophyte, lycophyta, monocot and eudicot WRKY domains revealed family member expansion with increasing complexity of the plant body. Microarray, expression profiling and qRT-PCR data revealed that WRKY genes in G. hirsutum may regulate the development of fibers, anthers, tissues (roots, stems, leaves and embryos), and are involved in the response to stresses. Expression analysis showed that most group II and III GhWRKY genes are highly expressed under diverse stresses. Group I members, representing the ancestral form, seem to be insensitive to abiotic stress, with low expression divergence. Our results indicate that cotton WRKY genes might have evolved by adaptive duplication, leading to sensitivity to diverse stresses. This study provides fundamental information to inform further analysis and understanding of WRKY gene functions in cotton species.

Cloning and characterization of FcWRKY40, A WRKY transcription factor from Fortunella crassifolia linked to oxidative stress tolerance

The WRKY transcription factors play a key role in regulating stress responses, but the functions of most WRKYs from the non-model plants remain poorly understood. Herein, we report the isolation and functional characterization of a Group IIa WRKY gene (FcWRKY40) in Meiwa kumquat (Fortunella crassifolia). FcWRKY40 encodes a putative protein of 257 amino acids containing a coiled-coil domain and a WRKY domain. FcWRKY40 was localized in the nuclei. Transcript levels of FcWRKY40 were upregulated by salt, salicylic acid, cold and abscisic acid, but repressed by dehydration. Transgenic tobacco plants overexpressing FcWRKY40 were shown to be more tolerant to the oxidative stresses than the wild type (WT), as indicated by better morphological phenotypes of leaf or whole plant, less severe cell death, and lower MDA and H2O2 levels. In addition, mRNA abundances of two genes related to H2O2 scavenging, peroxidase (POD) and catalase (CAT), were higher in the transgenic plants than in the WT before and after oxidative stress treatment. The promoters of POD and CAT genes contained W-box elements. Both yeast one-hybrid and transient expression assay demonstrated that FcWRKY40 could interact with the two promoters. Taken together, our data suggest that FcWRKY40 functions in the oxidative stress tolerance, possibly by alleviating H2O2 accumulation via regulation of the antioxidant genes.

Microarray analysis of laser-microdissected tissues indicates the biosynthesis of suberin in the outer part of roots during formation of a barrier to radial oxygen loss in rice (Oryza sativa).

Internal aeration is crucial for root growth in waterlogged soil. A barrier to radial oxygen loss (ROL) can enhance long-distance oxygen transport via the aerenchyma to the root tip; a higher oxygen concentration at the apex enables root growth into anoxic soil. The ROL barrier is formed within the outer part of roots (OPR). Suberin and/or lignin deposited in cell walls are thought to contribute to the barrier, but it is unclear which compound is the main constituent. This study describes gene expression profiles during ROL barrier formation in rice roots to determine the relative responses of suberin and/or lignin biosyntheses for the barrier. OPR tissues were isolated by laser microdissection and their transcripts were analysed by microarray. A total of 128 genes were significantly up- or downregulated in the OPR during the barrier formation. Genes associated with suberin biosynthesis were strongly upregulated, whereas genes associated with lignin biosynthesis were not. By an ab initio analysis of the promoters of the upregulated genes, the putative cis-elements that could be associated with transcription factors, WRKY, AP2/ERF, NAC, bZIP, MYB, CBT/DREB, and MADS, were elucidated. They were particularly associated with the expression of transcription factor genes containing WRKY, AP2, and MYB domains. A semiquantitative reverse-transcription PCR analysis of genes associated with suberin biosynthesis (WRKY, CYP, and GPAT) confirmed that they were highly expressed during ROL barrier formation. Overall, these results suggest that suberin is a major constituent of the ROL barrier in roots of rice.

Molecular characterization of 26 cotton WRKY genes that are expressed differentially in tissues and are induced in seedlings under high salinity and osmotic stress

WRKY proteins that contain highly conserved WRKYGQK sequence and zinc-finger-like motif belong to a large transcription factor family in plants. In this study, 26 genes encoding putative WRKY proteins were identified in cotton (Gossypium hirsutum). On the basis of their conserved WRKY domain sequences, all of the GhWRKY proteins can be assigned to the previously defined groups I–III. Among 26 members of cotton WRKY family, five with two WRKY domains belong to group I, sixteen contribute to group II, and five containing C2HC zinc-finger pattern fall into group III. Phylogenetic analysis of GhWRKY proteins with Arabidopsis WRKYs revealed that they were indeed classified into three groups, and also their relative distance can be used as clues to confer their putative functions. Quantitative RT-PCR analysis showed that the isolated GhWRKY genes were expressed differentially in different tissues (such as leaves, roots, cotyledons, petals, anthers, or/and fibers), suggesting they may have an impact on cotton development. Under NaCl, mannitol and abscisic acid treatments, expressions of the GhWRKY genes are up-regulated in roots, cotyledons and/or hypocotyls of cotton seedlings, suggesting that the isolated GhWRKY genes may take part in plant response to abiotic stress during cotton development. Thus, our results may provide a valuable reference data set as the basis for further studying the roles of these WRKY genes in cotton.

Genome-wide identification of WRKY family genes and their response to cold stress in Vitis vinifera.

BackgroundWRKY transcription factors are one of the largest families of transcriptional regulators in plants. WRKY genes are not only found to play significant roles in biotic and abiotic stress response, but also regulate growth and development. Grapevine (Vitis vinifera) production is largely limited by stressful climate conditions such as cold stress and the role of WRKY genes in the survival of grapevine under these conditions remains unknown.ResultsWe identified a total of 59 VvWRKYs from the V. vinifera genome, belonging to four subgroups according to conserved WRKY domains and zinc-finger structure. The majority of VvWRKYs were expressed in more than one tissue among the 7 tissues examined which included young leaves, mature leaves, tendril, stem apex, root, young fruits and ripe fruits. Publicly available microarray data suggested that a subset of VvWRKYs was activated in response to diverse stresses. Quantitative real-time PCR (qRT-PCR) results demonstrated that the expression levels of 36 VvWRKYs are changed following cold exposure. Comparative analysis was performed on data from publicly available microarray experiments, previous global transcriptome analysis studies, and qRT-PCR. We identified 15 VvWRKYs in at least two of these databases which may relate to cold stress. Among them, the transcription of three genes can be induced by exogenous ABA application, suggesting that they can be involved in an ABA-dependent signaling pathway in response to cold stress.ConclusionsWe identified 59 VvWRKYs from the V. vinifera genome and 15 of them showed cold stress-induced expression patterns. These genes represented candidate genes for future functional analysis of VvWRKYs involved in the low temperature-related signal pathways in grape.

Optimization of expression and purification of recombinant Salvia miltiorrhiza WRKY1 protein in Escherichia coli

WRKY transcription factor is one of the Zinc finger proteins which contains a highly conserved WRKY domain and is a family of the plant-specific transcription factor. The plasmid pET28a-SmWRKY1 harboring Salvia miltiorrhiza WRKY1 (SmWRKY1) gene was successfully transformed and expressed in Escherichia coli BL21 (DE3). The conditions on protein expression of SmWRKY1 in E. coli, including induction duration, temperature, IPTG concentration and the E. coli concentration were optimized. The results showed that the highest protein expression of SmWRKY1 was obtained at 24 hours after the E. coli was cultured in the presence of 0.2 mol x L(-1) IPTG at 20 degrees C with A600 values of 1.0-1.5. This recombinant histidine-tagged protein was expressed at 2.454 g x L(-1) as inclusion body, which was first extracted using urea, and then purified by Ni2+ affinity chromatography and identified by SDS-PAGE. The expression of SmWRKY1 in E. coli was further confirmed by western blotting analysis.

Genome-wide analysis of the WRKY transcription factor gene family in Gossypium raimondii and the expression of orthologs in cultivated tetraploid cotton

WRKY proteins are members of a family of transcription factors in higher plants that function in plant responses to various physiological processes. We identified 120 candidate WRKY genes from Gossypium raimondii with corresponding expressed sequence tags in at least one of four cotton species, Gossypium hirsutum, Gossypium barbadense, Gossypium arboreum, and G. raimondii. These WRKY members were anchored on 13 chromosomes in G. raimondii with uneven distribution. Phylogenetic analysis showed that WRKY candidate genes can be classified into three groups, with 20 members in group I, 88 in group II, and 12 in group III. The 88 genes in group II were further classified into five subgroups, groups IIa–e, containing 7, 16, 37, 15, and 13 members, respectively. We characterized diversity in amino acid residues in the WRKY domain and/or other zinc finger motif regions in the WRKY proteins. The expression patterns of WRKY genes revealed their important roles in diverse functions in cotton developmental stages of vegetative and reproductive growth and stress response. Structural and expression analyses show that WRKY proteins are a class of important regulators of growth and development and play key roles in response to stresses in cotton.

Genome-Wide Evolutionary Characterization and Expression Analyses of WRKY Family Genes in Brachypodium distachyon

Members of plant WRKY gene family are ancient transcription factors that function in plant growth and development and respond to biotic and abiotic stresses. In our present study, we have investigated WRKY family genes in Brachypodium distachyon, a new model plant of family Poaceae. We identified a total of 86 WRKY genes from B. distachyon and explored their chromosomal distribution and evolution, domain alignment, promoter cis-elements, and expression profiles. Combining the analysis of phylogenetic tree of BdWRKY genes and the result of expression profiling, results showed that most of clustered gene pairs had higher similarities in the WRKY domain, suggesting that they might be functionally redundant. Neighbour-joining analysis of 301 WRKY domains from Oryza sativa, Arabidopsis thaliana, and B. distachyon suggested that BdWRKY domains are evolutionarily more closely related to O. sativa WRKY domains than those of A. thaliana. Moreover, tissue-specific expression profile of BdWRKY genes and their responses to phytohormones and several biotic or abiotic stresses were analysed by quantitative real-time PCR. The results showed that the expression of BdWRKY genes was rapidly regulated by stresses and phytohormones, and there was a strong correlation between promoter cis-elements and the phytohormones-induced BdWRKY gene expression.

Functional characterisation of a WRKY transcription factor of wheat and its expression analysis during leaf rust pathogenesis.

WRKY proteins are a large family of plant-specific transcription factors associated with regulation of biotic and abiotic stress responses, but how they respond to cereal rust pathogens has never been explored at the molecular level. Full-length cDNA of TaWRKY1B was obtained from a wheat cultivar HD2329 derivative containing leaf rust resistance gene Lr28 based on domain characteristics. The unique feature of this WRKY transcription factor gene was the close proximity of the DNA-binding domain and consensus DNA element W-Box within the open reading frame. Infection with a virulent race of leaf rust fungus resulted in 146-fold induction of the gene in resistant plants, but only 12-fold in the susceptible plants as compared with mock-inoculated controls. Docking models of 74 amino acids DNA-binding domain and 26bp W-Box element showed that the WRKY domain, located on the β1 strand, only interacts with the W-Box at positions corresponding to W125, R126, K127 and Y128 amino acids. A truncated recombinant protein of 9.0 kD, encompassing the DNA-binding domain also showed binding specificity to the 32bp W-Box element in electrophoretic mobility shift assays. The protein-DNA ensemble was also characterised using high-resolution atomic force microscopic imaging. The results contribute to an understanding of the molecular structure and function of a previously uncharacterised WRKY transcription factor in wheat that can be manipulated to improve biotic stress tolerance.

The WRKY Transcription Factor Genes in Lotus japonicus.

WRKY transcription factor genes play critical roles in plant growth and development, as well as stress responses. WRKY genes have been examined in various higher plants, but they have not been characterized in Lotus japonicus. The recent release of the L. japonicus whole genome sequence provides an opportunity for a genome wide analysis of WRKY genes in this species. In this study, we identified 61 WRKY genes in the L. japonicus genome. Based on the WRKY protein structure, L. japonicus WRKY (LjWRKY) genes can be classified into three groups (I–III). Investigations of gene copy number and gene clusters indicate that only one gene duplication event occurred on chromosome 4 and no clustered genes were detected on chromosomes 3 or 6. Researchers previously believed that group II and III WRKY domains were derived from the C-terminal WRKY domain of group I. Our results suggest that some WRKY genes in group II originated from the N-terminal domain of group I WRKY genes. Additional evidence to support this hypothesis was obtained by Medicago truncatula WRKY (MtWRKY) protein motif analysis. We found that LjWRKY and MtWRKY group III genes are under purifying selection, suggesting that WRKY genes will become increasingly structured and functionally conserved.

501Genome-wide Analysis of WRKY Transcription Factors in <I>Solanum tuberosum</I>

The WRKY transcription factor family is widely involved in regulating plant development and defense response against various biotic and abiotic stresses.The Hidden Markov Model Profile(HMM) of WRKY domain was used to identify WRKY transcription factors in potato genome with HMMER 3.0.All together 81 potato WRKY transcription factors were identified and then classified into three main groups,with 5 subgroups in Group Ⅱ,namely Ⅱ-a,Ⅱ-b,Ⅱ-c,Ⅱ-d,and Ⅱ-e.The constructed phylogenetic tree of WRKY genes among potato,Arabidopsis and rice demonstrated that the distinct gene expansion events occurred in the genome of potato and rice,respectively,while the gene expansion of Group Ⅱ-e was unique in potato genome.The potato WRKY transcription factors showed similar motif compositions within each group,but different ones among different groups,though several conserved motifs were shared by all.Furthermore,28 potato WRKY genes showed distinct expression patterns in response to stresses of wounding,drought,salt,and virus invasion.The expressions of several potato WRKY genes were significantly up/down-regulated,implying that these members might participate in regulating the defense response against various biotic and abiotic stresses.

An abiotic stress-responsive WRKY gene is transiently induced in response to cold and drought stresses in Poncirus trifoliata

A partial cDNA sequence sharing significant homology with WRKY transcription factors (TFs) was previously amplified from Poncirus trifoliata by reverse transcription polymerase chain reaction (RT-PCR) using degenerate primers. In this study, the full-length cDNA sequence of this cDNA, PtrWRKY3, was obtained by RACE method from P. trifoliata. The length of PtrWRKY3 cDNA was 1908 bp and contained open reading frame coding for a 512-amino-acid polypeptide. The ortholog of this gene, CpWRKY3, was also cloned and sequenced from Citrus paradisi (grapefruit). The gene consists of 1550 bp cDNA encoding a protein of 516 amino acids. Both genes have double WRKY domains with Cys2His2 signature motif and showed 98% amino acid sequence identity with each other. They were tightly clustered together and showed a close phylogenetic relation with putative WRKY TFs from different woody perennial plants. The expression of the WRKY3 gene was analyzed quantitatively by northern blot and real-time RT-PCR in response to cold and drought stresses in both cold-hardy Poncirus and one of the cold-sensitive Citrus species, C. paradisi. While the expression of the WRKY3 gene was initially induced transiently in response to cold in cold-hardy Poncirus, no significant changes in the expression of this gene were observed in cold-sensitive grapefruit plants. On the other hand, the expression of the WRKY3 gene was transiently induced by drought stress in both Poncirus and grapefruit plants. These results indicated that the WRKY3 gene from Poncirus and Citrus is conserved and its response to drought stress was similar in both species. However, its expression in response to cold was different in cold-hardy and cold-sensitive species indicating that it is transiently induced early in cold acclimation and may be involved in cold tolerance in Poncirus.

Functional Analysis of a Novel Chrysanthemum WRKY Transcription Factor Gene Involved in Salt Tolerance

Plant-specific WRKY transcription factors (TFs) are involved in stress responses such as cold, high salinity, or drought as well as abscisic acid (ABA) signaling. However, their roles in abiotic stresses are still not well known in chrysanthemum. Here, we isolated a novel WRKY gene, DgWRKY1, from chrysanthemum (Dendranthema grandiflorum). DgWRKY1 contains one WRKY domain and one C2H2 zinc-finger motif (C-X4-C-X23-H-X-H), and was localized in the nucleus. Expression of DgWRKY1 was up-regulated by drought, salinity, and ABA. The DgWRKY1-overexpression tobacco plants were more tolerant to salt, and seed gerrmination was more sensitive to ABA, than the wild-type (WT). The transgenic lines exhibited less accumulation of hydrogen peroxide (H2O2) and malondialdehyde (MDA) under salt stress, and less antioxidant enzyme activity, including peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT), than the WT under both control conditions and salt stress. In addition, there was greater up-regulation of the ROS-related enzyme genes (NtSOD, NtPOD, and NtCAT) in transgenic lines under normal or salt conditions. These findings suggest that DgWRKY1 plays a positive regulatory role in salt stress response.

Characterization of HbWRKY1, a WRKY transcription factor from Hevea brasiliensis that negatively regulates HbSRPP

Small rubber particle protein (SRPP) is a major component of Hevea brasiliensis (H. brasiliensis) latex, which is involved in natural rubber (NR) biosynthesis. However, little information is available on the regulation of SRPP gene (HbSRPP) expression. To study the transcriptional regulation of HbSRPP, the yeast one-hybrid experiment was performed to screen the latex cDNA library using the HbSRPP promoter as bait. One cDNA that encodes the WRKY transcription factor, designated as HbWRKY1, was isolated from H. brasiliensis. HbWRKY1 contains a 1437 bp open reading frame that encodes 478 amino acids. The deduced HbWRKY1 protein was predicted to possess two conserved WRKY domains and a C2H2 zinc-finger motif. HbWRKY1 was expressed at different levels, with the highest transcription in the flower, followed by the bark, latex, and leaf. Furthermore, the co-expression of pHbSRP::GUS with CaMV35S::HbWRKY1 significantly decreased the GUS activity in transgenic tobacco, indicating that HbWRKY1 significantly suppressed the HbSRPP promoter. These results suggested that HbWRKY1 maybe a negative transcription regulator of HbSRPP involved in NR biosynthesis in H. brasiliensis.

Identification of a drought- and cold-stress inducible WRKY gene in the cold-hardy Citrus relative Poncirus trifoliata

WRKY transcription factors (TFs) are involved in regulation of abiotic and biotic stress responsive genes and play a role in abiotic and biotic stress tolerance in plants. Although commercial Citrus species are sensitive to cold and drought stresses, an interfertile Citrus relative, Poncirus trifoliata, is resistant to cold and more drought tolerant than most commercial Citrus cultivars. An expressed sequence tag clone sharing homology with WRKY-type TFs was previously isolated from a cDNA library constructed using two-day cold-acclimated P. trifoliata cv. Rubidoux seedlings. In this study, a full-length sequence of this WRKY gene was cloned and isolated from 2-day cold-acclimated P. trifoliata plants using the rapid amplification of cDNA ends (RACE) method. The full-length PtrWRKY1 cDNA is 1807 bp and encodes a polypeptide of 405 amino acids containing a single WRKY domain with a Cys2His2 motif, which is a signature of WRKY TFs. A homologous open reading frame (ORF) sequence was isolated from Citrus grandis, CgWRKY1, sharing 98% identity at the nucleotide level with PtrWRKY1. The expression of PtrWRKY1 and CgWRKY1 was investigated in response to cold and drought stresses by northern blot analysis. While the expression of both genes was induced in response to drought, only PtrWRKY1 expression was induced in response to cold. This differential expression in cold-hardy Poncirus and cold-sensitive pummelo suggests that PtrWRKY1 gene may play a role in adaptation and tolerance to cold stress in Poncirus.

Genome-wide analysis of the WRKY gene family in physic nut (Jatropha curcas L.)

The WRKY proteins, which contain highly conserved WRKYGQK amino acid sequences and zinc-finger-like motifs, constitute a large family of transcription factors in plants. They participate in diverse physiological and developmental processes. WRKY genes have been identified and characterized in a number of plant species. We identified a total of 58 WRKY genes (JcWRKY) in the genome of the physic nut (Jatropha curcas L.). On the basis of their conserved WRKY domain sequences, all of the JcWRKY proteins could be assigned to one of the previously defined groups, I–III. Phylogenetic analysis of JcWRKY genes with Arabidopsis and rice WRKY genes, and separately with castor bean WRKY genes, revealed no evidence of recent gene duplication in JcWRKY gene family. Analysis of transcript abundance of JcWRKY gene products were tested in different tissues under normal growth condition. In addition, 47 WRKY genes responded to at least one abiotic stress (drought, salinity, phosphate starvation and nitrogen starvation) in individual tissues (leaf, root and/or shoot cortex). Our study provides a useful reference data set as the basis for cloning and functional analysis of physic nut WRKY genes.

Discovery of WRKY transcription factors through transcriptome analysis and characterization of a novel methyl jasmonate-inducible PqWRKY1 gene from Panax quinquefolius

Transcription factors (TFs) are important regulating factors that can mediate many life processes. However, no TF genes have previously been reported in Panax quinquefolius (American ginseng), with the exception of a few expressed sequence tags. In this study, 753 unigenes (unique sequences) have been annotated in the plant transcription factor database PlnTFDB (version 3.0) by mining a 454 transcriptome dataset of P. quinquefolius. After classification of the unigenes, 45 unigenes were discovered to be annotated as WRKY transcripts in the public databases. Furthermore, PqWRKY1, one of the WRKY family TF genes that respond to methyl jasmonate, was isolated according to the sequences in the 454 transcriptome dataset. The cDNA of PqWRKY1 (P. quinquefolius WRKY1) encodes a putative protein of 358 amino acids, including a WRKY domain and a zinc finger motif. A subcellular localization assay demonstrated that the protein localizes to the nucleus and has strong transcriptional activation activity in transgenic yeast. In comparison to control lines, the PqWRKY1 transgenic Arabidopsis line exhibited insensitive phenotypes when exposed to high salt or mannitol. Correspondingly, in transgenic Arabidopsis lines, the expression levels of some genes involved in the anti-stress process were relatively higher than those in the control lines. Additionally, genes involved in triterpene biosynthesis were expressed onefold to fivefold higher in the transgenic line compared to the control line. These results suggest that the transcription factor PqWRKY1 is a positive regulator related to osmotic stress and triterpene ginsenoside biosynthesis in P. quinquefolius.

WRKY Transcription Factors in Wheat and Their Induction by Biotic and Abiotic Stress

WRKY proteins constitute a large family of transcription factors in higher plants, and are involved in the regulation of development, senescence and stress resistance. However, little is known with respect to the constitution and function of WRKY genes in bread wheat (Triticum aestivum). We identified a set of 92 wheat WRKY genes from publicly available sequence databases, and constructed a phylogeny map based on the conserved WRKY domain peptide sequences. The wheat sequences were classified into the standard set of WRKY groups and sub-groups established from plant model genomes. Our results show the WRKY orthologues of wheat and rice are more similar compared to their Arabidopsis counterparts. Therefore, we argue some expansion of the WRKY family occurred following the divergence of these three species. The induction of sequence variation in a subset of 18 TaWRKY genes via somatic hybridization was studied by comparing the sequences from cv. JN177 and its derivative cv. SR3. We also analyzed the responses of these 18 genes to imposed salinity, PEG, ABA and SA using quantitative real time PCR. Nine of the 18 TaWRKY genes were up-regulated by exogenous SA. Six genes were up-regulated by salinity or PEG treatment, and their expression was also induced by exogenous ABA. Interestingly, induction by salinity or PEG was impaired when ABA biosynthesis was inhibited. The indication was that most of the TaWRKY genes involved in the abiotic stress response acted in an ABA-dependent manner.