Stress-related Cis-acting Elements Research Articles

Genome-Wide Analysis of Myeloblastosis-Related Genes in Brassica napus L. and Positive Modulation of Osmotic Tolerance by BnMRD107.

Myeloblastosis (MYB)-related transcription factors comprise a large subfamily of the MYB family. They play significant roles in plant development and in stress responses. However, MYB-related proteins have not been comprehensively investigated in rapeseed (Brassica napus L.). In the present study, a genome-wide analysis of MYB-related transcription factors was performed in rapeseed. We identified 251 Brassica napus MYB (BnMYB)-related members, which were divided phylogenetically into five clades. Evolutionary analysis suggested that whole genome duplication and segmental duplication events have played a significant role in the expansion of BnMYB-related gene family. Selective pressure of BnMYB-related genes was estimated using the Ka/Ks ratio, which indicated that BnMYB-related genes underwent strong purifying selection during evolution. In silico analysis showed that various development-associated, phytohormone-responsive, and stress-related cis-acting regulatory elements were enriched in the promoter regions of BnMYB-related genes. Furthermore, MYB-related genes with tissue or organ-specific, stress-responsive expression patterns were identified in B. napus based on temporospatial and abiotic stress expression profiles. Among the stress-responsive MYB-related genes, BnMRD107 was strongly induced by drought stress, and was therefore selected for functional study. Rapeseed seedlings overexpressing BnMRD107 showed improved resistance to osmotic stress. Our findings not only lay a foundation for further functional characterization of BnMYB-related genes, but also provide valuable clues to determine candidate genes for future genetic improvement of B. napus.

A salicylic acid inducible mulberry WRKY transcription factor, MiWRKY53 is involved in plant defence response.

MiWRKY53 is expressed in response to various stresses and hormones. Although it is localized in the nucleus, it shows no transcriptional activation. Role of SA-mediated plant defence response is demonstrated. WRKY transcription factors are one the largest gene families in plants involved in almost every process in plants including development, physiological processes, and stress response. Salicylic acid (SA) is key regulator of biotic stress against various pathogens in plants acting via its multiple mechanisms to induce defence response. Herein, we have identified and functionally validated WRKY53 from mulberry (Morus indica var. K2). MiWRKY53 expressed differentially in response to different stress and hormonal treatments. MiWRKY53 belongs to group III of WKRY gene family, localized in nucleus, and lacks transcriptional activation activity in yeast. Hormone responsive behaviour of MiWRKY53 Arabidopsis overexpression (OE) transgenics preferentially was noted in root growth assay in response to Salicylic acid (SA). Arabidopsis overexpression plants also displayed alteration in leaf phenotype having wider leaves than the wild-type plants. PR-1 transcripts were higher in MiWRKY53 Arabidopsis OE plants and they displayed resistance towards biotrophic pathogen Pseudomonas syringae PstDC3000. MiWRKY53 Mulberry OE transgenics also depicted SA-responsive behaviour. Several hormones and stress-related cis-acting elements were also identified in the 1.2-Kb upstream regulatory region (URR) of MiWRKY53. Functional characterization of full-length promoter region revealed that it is induced by SA and further analysis of deletion constructs helped in the identification of minimal promoter responsible for its inducibility by SA. Altogether, the findings from this study point towards the SA preferential behaviour of MiWRKY53 and its function as regulator of plant defence response through SA-mediated mechanisms.

Molecular Characterization, Expression Pattern and Function Analysis of Glycine-Rich Protein Genes Under Stresses in Chinese Cabbage (Brassica rapa L. ssp. pekinensis).

Plant Glycine-rich proteins (GRP), a superfamily with a glycine-rich domain, play an important role in various stresses such as high or low temperature stress and drought stress. GRP genes have been studied in many plants, but seldom in Chinese cabbage (Brassica rapa L. ssp. pekinensis). In this study, a total of 64 GRP genes were identified in Chinese cabbage by homology comparative analysis. The physical and chemical characteristics predicted by ProtParam tool revealed that 62.5% of BrGRPs were alkaline, 53.1% were stable, and 79.7% were hydrophilic. Conserved domain analysis by MEME and TBtools showed that 64 BrGRPs contained 20 of the same conserved motifs, based on which BrGRPs were classified into five main classes and four subclasses in class IV to clarify their evolutionary relationship. Our results demonstrated that The BrGRP genes were located on ten chromosomes and in three different subgenomes of Chinese cabbage, and 43 pairs of orthologous GRP genes were found between Chinese cabbage and Arabidopsis. According to the transcriptome data, 64 BrGRP genes showed abnormal expression under high temperature stress, 52 under low temperature stress, 39 under drought stress, and 23 responses to soft rot. A large number of stress-related cis-acting elements, such as DRE, MYC, MYB, and ABRE were found in their promoter regions by PlantCare, which corresponded with differential expressions. Two BrGRP genes-w546 (Bra030284) and w1409 (Bra014000), both belonging to the subfamily Subclass IVa RBP-GRP (RNA binding protein-glycine rich protein), were up-regulated under 150 mmol⋅L–1 NaCl stress in Chinese cabbage. However, the overexpressed w546 gene could significantly inhibit seed germination, while w1409 significantly accelerated seed germination under 100 mmol⋅L–1 NaCl or 300 mmol⋅L−1 mannitol stresses. In short, most BrGRP genes showed abnormal expression under adversity stress, and some were involved in multiple stress responses, suggesting a potential capacity to resist multiple biotic and abiotic stresses, which is worthy of further study. Our study provides a systematic investigation of the molecular characteristics and expression patterns of BrGRP genes and promotes for further work on improving stress resistance of Chinese cabbage.

Identification and expression analysis of WRKY gene family under drought stress in peanut (Arachis hypogaea L.).

WRKY transcription factors play crucial roles in regulation mechanism leading to the adaption of plants to the complex environment. In this study, AhWRKY family was comprehensively analyzed using bioinformatic approaches in combination with transcriptome sequencing data of the drought-tolerant peanut variety ‘L422’. A total of 158 AhWRKY genes were identified and named according to their distribution on the chromosomes. Based on the structural features and phylogenetic analysis of AhWRKY proteins, the AhWRKY family members were classified into three (3) groups, of which group II included five (5) subgroups. Results of structure and conserved motifs analysis for the AhWRKY genes confirmed the accuracy of the clustering analysis. In addition, 12 tandem and 136 segmental duplication genes were identified. The results indicated that segmental duplication events were the main driving force in the evolution of AhWRKY family. Collinearity analysis found that 32 gene pairs existed between Arachis hypogaea and two diploid wild ancestors (Arachis duranensis and Arachis ipaensis), which provided valuable clues for phylogenetic characteristics of AhWRKY family. Furthermore, 19 stress-related cis-acting elements were found in the promoter regions. During the study of gene expression level of AhWRKY family members in response to drought stress, 73 differentially expressed AhWRKY genes were obtained to have been influenced by drought stress. These results provide fundamental insights for further study of WRKY genes in peanut drought resistance.

Characterization of the AP2/ERF Transcription Factor Family and Expression Profiling of DREB Subfamily under Cold and Osmotic Stresses in Ammopiptanthus nanus.

APETALA2/ethylene-responsive factor (AP2/ERF) is one of the largest transcription factor (TF) families in plants, which play important roles in regulating plant growth, development, and response to environmental stresses. Ammopiptanthus nanus, an unusual evergreen broad-leaved shrub in the arid region in the northern temperate zone, demonstrates a strong tolerance to low temperature and drought stresses, and AP2/ERF transcription factors may contribute to the stress tolerance of A. nanus. In the current study, 174 AP2/ERF family members were identified from the A. nanus genome, and they were divided into five subfamilies, including 92 ERF members, 55 dehydration-responsive element binding (DREB) members, 24 AP2 members, 2 RAV members, and 1 Soloist member. Compared with the other leguminous plants, A. nanus has more members of the DREB subfamily and the B1 group of the ERF subfamily, and gene expansion in the AP2/ERF family is primarily driven by tandem and segmental duplications. Promoter analysis showed that many stress-related cis-acting elements existed in promoter regions of the DREB genes, implying that MYB, ICE1, and WRKY transcription factors regulate the expression of DREB genes in A. nanus. Expression profiling revealed that the majority of DREB members were responsive to osmotic and cold stresses, and several DREB genes such as EVM0023336.1 and EVM0013392.1 were highly induced by cold stress, which may play important roles in cold response in A. nanus. This study provided important data for understanding the evolution and functions of AP2/ERF and DREB transcription factors in A. nanus.

Function analysis of ZmNAC33, a positive regulator in drought stress response in Arabidopsis

Drought significantly affects plant growth and has devastating effects on crop production, NAC transcription factors respond to abiotic stresses by activating gene expression. In this study, a maize NAC transcription factor, ZmNAC33, was cloned and characterized its function in Arabidopsis. Transient transformation in Arabidopsis leaves mesophyll protoplasts and trans-activation assays in yeast showed that ZmNAC33 was localized in the nucleus and had transactivation activity. qRT-PCR analysis showed that ZmNAC33 in maize was induced by drought, high salinity and abscisic acid (ABA) stress. Promoter analysis identified multiple stress-related cis-acting elements in the promoter region of ZmNAC33. In ZmNAC33 transgenic Arabidopsis, germination rates were higher than in wild type plants under ABA and osmotic stress at the germination stage, and overexpression lines exhibited higher survival rates and higher antioxidant enzyme activities compared with wild type under drought stress. These results indicate that ZmNAC33 actes as a positive regulator in drought tolerance in plants.

A Stress-Responsive NAC Transcription Factor from Tiger Lily (LlNAC2) Interacts with LlDREB1 and LlZHFD4 and Enhances Various Abiotic Stress Tolerance in Arabidopsis.

Our previous studies have indicated that a partial NAC domain protein gene is strongly up-regulated by cold stress (4 °C) in tiger lily (Lilium lancifolium). In this study, we cloned the full-length of this NAC gene, LlNAC2, to further investigate the function of LlNAC2 in response to various abiotic stresses and the possible involvement in stress tolerance of the tiger lily plant. LlNAC2 was noticeably induced by cold, drought, salt stresses, and abscisic acid (ABA) treatment. Promoter analysis showed that various stress-related cis-acting regulatory elements were located in the promoter of LlNAC2; and the promoter was sufficient to enhance activity of GUS protein under cold, salt stresses and ABA treatment. DREB1 (dehydration-responsive binding protein1) from tiger lily (LlDREB1) was proved to be able to bind to the promoter of LlNAC2 by yeast one-hybrid (Y1H) assay. LlNAC2 was shown to physically interact with LlDREB1 and zinc finger-homeodomain ZFHD4 from the tiger lily (LlZFHD4) by bimolecular fluorescence complementation (BiFC) assay. Overexpressing LlNAC2 in Arabidopsis thaliana showed ABA hypersensitivity and enhanced tolerance to cold, drought, and salt stresses. These findings indicated LlNAC2 is involved in both DREB/CBF-COR and ABA signaling pathways to regulate stress tolerance of the tiger lily.

A MYB-Related Transcription Factor from Lilium lancifolium L. (LlMYB3) Is Involved in Anthocyanin Biosynthesis Pathway and Enhances Multiple Abiotic Stress Tolerance in Arabidopsis thaliana.

Most commercial cultivars of lily are sensitive to abiotic stresses. However, tiger lily (Lilium lancifolium L.), one of the most widely distributed wild lilies in Asia, has strong abiotic stresses resistance. Thus, it is indispensable to identify stress-responsive candidate genes in tiger lily for the stress resistance improvement of plants. In this study, a MYB related homolog (LlMYB3) from tiger lily was functionally characterized as a positive regulator in plant stress tolerance. LlMYB3 is a nuclear protein with transcriptional activation activity at C-terminus. The expression of LlMYB3 gene was induced by multiple stress treatments. Several stress-related cis-acting regulatory elements (MYBRS, MYCRS, LTRE and DRE/CRT) were located within the promoter of LlMYB3; however, the promoter activity was not induced sufficiently by various stresses treatments. Overexpressing LlMYB3 in Arabidopsis thaliana L. transgenic plants showed ABA hypersensitivity and enhanced tolerance to cold, drought, and salt stresses. Furthermore, we found LlMYB3 highly co-expressed with LlCHS2 gene under cold treatment; yeast one-hybrid (Y1H) assays demonstrated LlMYB3 was able to bind to the promoter of LlCHS2. These findings suggest that the stress-responsive LlMYB3 may be involved in anthocyanin biosynthesis pathway to regulate stress tolerance of tiger lily.

Comparative phylogenetic analysis of aquaporins provides insight into the gene family expansion and evolution in plants and their role in drought tolerant and susceptible chickpea cultivars

Aquaporins (AQPs) are water channel proteins that play a significant role in drought stress. Although the AQPs identified in multiple plant species, there is no detailed evolutionary and comparative study of AQPs regarding chickpea plant. The current study involved evolutionary analyses coupled with promoter and expression analyses of chickpea AQPs (CaAQPs). A total of 924 non-redundant AQPs were studied in 24 plant species including algae, mosses, lycophytes, monocots and dicots. Phylogenetic analysis demonstrated a clear divergence of eight AQP subfamilies (LIPs, SIPs, GIPs, NIPs, XIPs, PIPs, HIPs and TIPs). The comparative phylogenetic trees of AQP subfamilies among Arabidopsis, soybean, common bean, maize and chickpea demonstrated that the AQPs were highly species-specific. Interestingly, the dual NPA motif was conserved in all species. However, the ar/R selectivity filter signatures [W/T/S/N/G/A]-[V/S/L/I/A]-[S/G/A]-R (in NIPs), F-H-T-R (in PIPs), [H/N/Q/S]-[A/I/L/S/V]-[A/G]-[A/C/L/M/R/V] (in TIPs) and [V/I/L/M]-[V/I/A/F/M]-[A/S/F/C]-[N/F/L/I/A/S (in SIPs) were found in five species. Moreover, the Froger's positions (P1-P5) were found as [F/L/Y]-[S/T]-A-Y-[L/I/M/V/F] (in NIPs), [Q/E/M]-S-A-F-W (in PIPs), [A/L/S/T/V]-[A/C/N/S/T/V]-[P/R/S]-[Y/N/F]-[W/Q] (in TIPs) and [I/M/F]-[A/V]-[A/V]-Y-W (in SIPs). The MEME motif analyses showed that most of the motifs were specific to subfamily and subgroups. Tissue-specific expression profiling of CaAQPs revealed that CaTIPs and CaPIPs are highly expressed in most of the tissues, while CaNIPs and CaSIPs have low expression. In promoter analysis of CaAQPs, multiple stress-related cis-acting elements e.g. MYB, MYC, ABRE, etc. were found. Semi-quantitative RT-PCR analysis showed that CaPIP2;3 and CaNIP3;1 are positive regulator, while CaSIP1;1 and CaPIP2;1 have a negative role in drought tolerance. The findings and implications of this study are discussed in detail.

Genomewide analysis of homeobox gene family in apple (Malus domestica Borkh.) and their response to abiotic stress

Homeobox proteins (HOXs) comprise a large family in eukaryotes and share a highly conserved DNA-binding motif, the homeodomain (HD). HOXs play an important role in the regulation of plant growth, development and stress response. However, systematic analysis and expression profiling of these genes have not been reported in Malus domestica. In this study, a total of 207 HOXs of M. domestica (MdHOXs) were identified and classified into 11 distinct subfamilies, and an unclassified group according to their functional domains. The MdHOXs were localized in all 17 chromosomes with various densities and a majority of them tended to formgene clusters. Analysis of the Ka/Ks ratios suggested that the duplicated genes of MdHOXs mainly underwent purifying selection with restrictive functional divergence after the duplication events. The expression of MdHOXs has organ specific characteristics and were divided into seven different groups. Stress-related cis-acting elements were prevalent in the upstream sequence of MdHOXs by systematic analysis. To explore the response to abiotic stress, eight MdHOXs were randomly selected to investigate their expression using quantitative real-time polymerase chain reaction. Transcription levels of MdHOXs were upregulated in the leaves and roots under cold, osmotic, high salinity or exogenous ABA treatments, which suggested that they may take part in the plant response to abiotic stress. These results provided basic information of HOXs in apple and will further contribute to the functional research of MdHOXs, especially the response to abiotic stress.

Identification of a novel promoter from banana aquaporin family gene (MaTIP1;2) which responses to drought and salt-stress in transgenic Arabidopsis thaliana

Drought and salt stresses often affect plant growth and crop yields. Identification of promoters involved in drought and salt stress responses is of great significance for genetic improvement of crop resistance. Our previous studies showed that aquaporin can respond to drought and salt stresses, but its promoter has not yet been reported in plants. In the present study, cis-acting elements of MaAQP family member promoters were systematically analyzed in banana. Expression of MaTIP1; 2 was induced by drought and salt stresses but not sensitive to cold stress, waterlogging stress, or mechanical damage, and its promoter contained five stress-related cis-acting elements. The MaTIP1; 2 promoter (841 bp upstream of translation initiation site) from banana (Musa acuminata L. AAA group cv. Brazilian) was isolated through genome walking polymerase chain reaction, and found to contain a TATA Box, CAAT box, ABRE element, CCGTCC box, CGTCA motif, and TCA element. Transformation of the MaTIP1; 2 promoter into Arabidopsis to assess its function indicated that it responds to both drought and salt stress treatments. These results suggest that MaTIP1; 2 utilization may improve drought and salt stresses resistance of the transgenic plants by promoting banana aquaporin expression.

Identification and characterization of the BZR transcription factor family and its expression in response to abiotic stresses in Zea mays L.

Brassinosteroids (BRs) are plant specific steroidal hormones that play diverse roles in regulating a broad spectrum of plant growth and developmental processes, as well as, in responding to various biotic and abiotic stresses. Extensive research over the years has established stress-impact-mitigating role of BRs and associated compounds in different plants exposed to various abiotic and biotic stresses, suggesting the idea that they may act as immunomodulators, thus opening new approaches for plant resistance against hazardous environmental conditions. In this research the characterization of the transcriptional response of 11 transcription factors (TFs) belonging to BRASSINAZOLE-RESISTANT 1 (BZR1) TF family of Zea mays L. was analyzed in seedlings subjected to different stress conditions. Being important regulators of the BR synthesis, BZR TFs might have stress resistance related activities. However, no stress resistance related functional study of BZR TFs has been reported in maize so far. In silico analyses of the selected 11 TFs validated the features of their protein domains, where the highest degree of similarity observed with recognized BZR TFs of rice and Sorghum bicolor. Additionally, we investigated the organ-specific expression of 11 ZmBZR in maize seedlings. Five of them did not show any transcript accumulation, suggesting that ZmBZR expression might be regulated in a manner dependent on plant developmental stage. For the remaining six ZmBZR, their ubiquitous expression in the whole plant indicates they could function as growth regulators during maize development. More importantly, in response to various stress conditions, the spatial transcript accumulation of all ZmBZR varies along the plant. All six ZmBZR showed up-regulation against N starvation, hypoxia and salt stress. On the contrary, heat stress clearly down-regulated gene expression of all ZmBZR analysed. Consistently with the expression results, the distribution of stress-related cis-acting elements in the promoter of these genes inferred that the maize BZR TFs might play some roles in regulating the expression of the corresponding genes in response to multifarious stresses. In conclusion, these data reveal that BZR TFs have stress signaling activity in maize, in addition to their confirmed role in regulating plant physiology and morphology.

Systematic Analysis of Hsf Family Genes in the Brassica napus Genome Reveals Novel Responses to Heat, Drought and High CO2 Stresses.

Drought and heat stress are major causes of lost plant crop yield. In the future, high levels of CO2, in combination of other abiotic stress factors, will become a novel source of stress. Little is known of the mechanisms involved in the acclimation responses of plants to this combination of abiotic stress factors, though it has been demonstrated that heat shock transcription factors (Hsfs) are involved in plant response to various abiotic stresses. In this study, we performed a genome-wide identification and a systematic analysis of genes in the Hsf gene family in Brassica napus. A total of 64 genes encoding Hsf proteins were identified and classified into 3 major classes: A, B and C. We found that, unlike in other eudicots, the A9 subclass is absent in rapeseed. Further gene structure analysis revealed a loss of the only intron in the DBD domain for BnaHsf63 and -64 within class C, which is evolutionarily conserved in all Hsf genes. Transcription profile results demonstrated that most BnaHsf family genes are upregulated by both drought and heat conditions, while some are responded to a high CO2 treatment. According to the combined RNA-seq and qRT-PCR analysis, the A1E/A4A/A7 subclasses were upregulated by both drought and heat treatments. Members in class C seemed to be predominantly induced only by drought. Among BnaHsf genes, the A2/A3/B2 subclasses were regulated by all three abiotic stresses. Members in A2/B2 subclasses were upregulated by drought and heat treatments, but were downregulated under high CO2 conditions. While the A3 subclass was upregulated by all the three abiotic stresses. Various stress-related cis-acting elements, enriched in promoter regions, were correlated with the transcriptional response of BnaHsfs to these abiotic stresses. Further study of these novel groups of multifunctional BnaHsf genes will improve our understanding of plant acclimation response to abiotic stresses, and may be useful for improving the abiotic stress resistance of crop varieties.

Expression analysis and promoter methylation under osmotic and salinity stress of TaGAPC1 in wheat (Triticum aestivum L).

Cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) catalyzes a key reaction in glycolysis and encoded by a multi-gene family which showed instability expression under abiotic stress. DNA methylation is an epigenetic modification that plays an important role in gene regulation in response to abiotic stress. The comprehension of DNA methylation at promoter region of TaGAPC1 can provide insights into the transcription regulation mechanisms of plant genes under abiotic stress. In this study, we cloned TaGAPC1 genes and its promoters from two wheat genomes, then investigated the expression patterns of TaGAPC1 under osmotic and salinity stress, and analyzed the promoter sequences. Moreover, the methylation patterns of promoters under stress were confirmed. Expression analysis indicated that TaGAPC1 was induced inordinately by stresses in two wheat genotypes with contrasting drought tolerance. Several stress-related cis-acting elements (MBS, DRE, GT1 and LTR et al.) were located in its promoters. Furthermore, the osmotic and salinity stress induced the demethylation of CG and CHG nucleotide in the promoter region of Changwu134. The methylation level of CHG and CHH in promoter of Zhengyin1 was always increased under stresses, and the CG contexts remained unchanged. The cytosine loci of stress-related cis-acting elements also showed different methylation changes in this process. These results provide insights into the relationship between promoter methylation and gene expression, promoting the function investigation of GAPC.

Expression and regulation of a cold-responsive gene,CsCBFinCitrus sinensis(L.) Osbeck under low temperature, high salinity and abscisic acid

CBF (C-Repeat/Dehydration-Responsive Element Binding Factor) as a transcription factor plays important roles in stress responses and stress-related regulatory network in plants. CsCBF, a CBF encoding gene in sweet orange (Citrus sinensis (L.) Osbeck) was isolated and investigated in expression profiles under abiotic stresses. Semi-quantitative RT-PCR analysis indicated that CsCBF expression was altered obviously in response to low temperature (4°C), high salinity (250 mM NaCl) and abscisic acid (100 μM ABA). To learn more information about CsCBF gene, genomic DNA and upstream regulatory sequence were sequenced definitely and analyzed with a series of bioinformatic tools. The results showed that there were no introns in CsCBF genomic DNA, and CBF signature sequences and AP2 DNA binding domain were also highly conserved in CsCBF amino acid sequence. In addition, more than one CBF protein was encoded in sweet orange genome. Promoter prediction analysis suggested that several stress-related cis-acting elements were detected in CsCBF regulatory region, such as MYB, MYC, ABRE, Circadian, et al.. Importantly, two putative induction regions of CBF expression, ICEr1 and ICEr2, were also found in an 85-bp section of CsCBF promoter, which may contribute to understanding of CBF transcriptional activation under stress.

Drought-responsive WRKY transcription factor genes TaWRKY1 and TaWRKY33 from wheat confer drought and/or heat resistance in Arabidopsis.

BackgroundDrought stress is one of the major causes of crop loss. WRKY transcription factors, as one of the largest transcription factor families, play important roles in regulation of many plant processes, including drought stress response. However, far less information is available on drought-responsive WRKY genes in wheat (Triticum aestivum L.), one of the three staple food crops.ResultsForty eight putative drought-induced WRKY genes were identified from a comparison between de novo transcriptome sequencing data of wheat without or with drought treatment. TaWRKY1 and TaWRKY33 from WRKY Groups III and II, respectively, were selected for further investigation. Subcellular localization assays revealed that TaWRKY1 and TaWRKY33 were localized in the nuclei in wheat mesophyll protoplasts. Various abiotic stress-related cis-acting elements were observed in the promoters of TaWRKY1 and TaWRKY33. Quantitative real-time PCR (qRT-PCR) analysis showed that TaWRKY1 was slightly up-regulated by high-temperature and abscisic acid (ABA), and down-regulated by low-temperature. TaWRKY33 was involved in high responses to high-temperature, low-temperature, ABA and jasmonic acid methylester (MeJA). Overexpression of TaWRKY1 and TaWRKY33 activated several stress-related downstream genes, increased germination rates, and promoted root growth in Arabidopsis under various stresses. TaWRKY33 transgenic Arabidopsis lines showed lower rates of water loss than TaWRKY1 transgenic Arabidopsis lines and wild type plants during dehydration. Most importantly, TaWRKY33 transgenic lines exhibited enhanced tolerance to heat stress.ConclusionsThe functional roles highlight the importance of WRKYs in stress response.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-016-0806-4) contains supplementary material, which is available to authorized users.

ZmNAC55, a maize stress-responsive NAC transcription factor, confers drought resistance in transgenic Arabidopsis

Abiotic stress has been shown to significantly limit the growth and productivity of crops. NAC transcription factors play essential roles in response to various abiotic stresses. However, only little information regarding stress-related NAC genes is available in maize. Here, we cloned a maize NAC transcription factor ZmNAC55 and identified its function in drought stress. Transient expression and transactivation assay demonstrated that ZmNAC55 was localized in the nucleus and had transactivation activity. Expression analysis of ZmNAC55 in maize showed that this gene was induced by drought, high salinity and cold stresses and by abscisic acid (ABA). Promoter analysis demonstrated that multiple stress-related cis-acting elements exist in promoter region of ZmNAC55. Overexpression of ZmNAC55 in Arabidopsis led to hypersensitivity to ABA at the germination stage, but enhanced drought resistence compared to wild-type seedlings. Transcriptome analysis identified a number of differentially expressed genes between 35S::ZmNAC55 transgenic and wild-type plants, and many of which are involved in stress response, including twelve qRT–PCR confirmed well-known drought-responsive genes. These results highlight the important role of ZmNAC55 in positive regulates of drought resistence, and may have potential applications in transgenic breeding of drought-tolerant crops.

CarNAC4, a NAC-type chickpea transcription factor conferring enhanced drought and salt stress tolerances in Arabidopsis.

CarNAC4 is a typical stress-responsive NAC transcription factor and enhances drought and salt stress tolerances in transgenic Arabidopsis. Chickpea (Cicer arietinum L.) is relatively vulnerable to abiotic stress conditions, but the tolerance mechanisms for such stresses in chickpea are largely unknown. To identify stress-related factors in chickpea, we previously constructed a cDNA library of chickpea leaves exposed to drought stress conditions. A cDNA encoding a putative NAC transcription factor (CarNAC4) was identified as a putative stress-responsive gene. Our study indicated that the transcript levels of CarNAC4 were enhanced in response to several abiotic stresses and phytohormones. Promoter analysis demonstrated that multiple stress-related cis-acting elements exist in promoter region of CarNAC4. CarNAC4 is localized in the nucleus and binds to the DNA sequence containing CGT[G/A], while the C-terminal region of CarNAC4 contains a transcriptional activation domain. Over-expression of CarNAC4 in Arabidopsis plants improved tolerance to drought and salt stresses. Transgenic plants exhibited greater reduced rates of water loss and more proline accumulation than Col-0 plants under drought stress and less MDA contents than Col-0 plants under salt stress. In addition, over-expression of CarNAC4 enhanced the expression of stress-responsive genes such as RD29A, ERD10, COR15A, COR47, KIN1 and DREB2A. These results indicated that CarNAC4 functions as a transcription factor involved in the regulation of drought and salt stress response.

Molecular analysis of the chloroplast Cu/Zn-SOD gene (AhCSD2) in peanut

Superoxide dismutase (SOD, EC 1.15.1.1) plays a key role in response to drought stress, and differences in SOD activity changes among cultivars are important under drought conditions. We obtained the full-length DNA of the chloroplast Cu/Zn-SOD gene (AhCSD2) from 11 allotetraploid cultivars and 5 diploid wild species in peanut. BLAST search against the peanut genome showed that the AhCSD2 genes gCSD2-1 and gCSD2-2 are located at the tops of chromosome A03 (A genome) and B03 (B genome), respectively, and both contain 8 exons and 7 introns. Nucleotide sequence analyses indicated that gCSD2-2 sequences were identical among all the tested cultivars, while gCSD2-1 sequences showed allelic variations. The amino acid sequences deduced from gCSD2-1 and gCSD2-2 both contain a chloroplast transit peptide and are distinguished by 6 amino acid (aa) residue differences. The other 2 aa residue variations in the mature peptide regions give rise to three-dimensional structure changes of the protein deduced from the genes gCSD2-1 and gCSD2-2. Sequences analyses of cultivars and wild species showed that gCSD2-2 of Arachis hypogaea and gAipCSD2 (Arachis ipaensis) are identical, and despite the abundant polymorphic loci between gCSD2-1 of A. hypogaea and sequences from A genome wild species, the deduced amino acid sequence of AhCSD2-1 (A. hypogaea) is identical to that of AduCSD2 (Arachis duranensis), whereas AcoCSD2 (Arachis correntina) and AcaCSD2 (Arachis cardenasii) both have 2 aa differences in the transit peptide region compared with AhCSD2-1 (A. hypogaea). Based on the Peanut Genome Project, promoter prediction revealed many stress-related cis-acting elements within the potential promoter regions (pp-A and pp-B). pp-A contains more binding sites for drought-associated transcriptional factors than pp-B. We hypothesize that the marked changes in SOD activity in different cultivars under drought stress are tightly regulated by transcription factors through transcription and expression of AhCSD2 genes.

Functional analyses of the maize CKS2 gene promoter in response to abiotic stresses and hormones

In our previous research, we showed that the cyclin-dependent kinase regulatory subunit (CKS2) in maize (Zea mays L.) was induced by water deficit and cold stress. To elucidate its expression patterns under adversity, we isolated and characterized its promoter (PZmCKS2). A series of PZmCKS2-deletion derivatives, P0–P3, from the translation start code (−1,455, −999, −367, and −3 bp) was fused to the β-glucuronidase (GUS) reporter gene, and each deletion construct was analyzed by Agrobacterium-mediated steady transformation into Arabidopsis. Leaves were then subjected to dehydration, cold, abscisic acid (ABA), salicylic acid (SA), and methyl jasmonic acid (MeJA). Sequence analysis showed that several stress-related cis-acting elements (MBS, CE3, TGA element, and ABRE) were located within the promoter. Deletion analysis of the promoter, PZmCKS2, suggested that the −999 bp promoter region was required for the highest basal expression of GUS, and the −367 bp sequence was the minimal promoter for ZmCKS2 activation by low temperature, ABA, and MeJA. The cis-acting element ABRE was necessary for promoter activation by exogenous ABA.