Abiotic Stress-responsive Genes Research Articles

Identification, Expression, and Interaction Network Analyses of the CDPK Gene Family Reveal Their Involvement in the Development, Ripening, and Abiotic Stress Response in Banana.

Calcium-dependent protein kinases (CDPKs) play vital roles in the regulation of plant growth, development, and tolerance to various abiotic stresses. However, little information is available for this gene family in banana. In this study, 44 CDPKs were identified in banana and were classified into four groups based on phylogenetic, gene structure, and conserved motif analyses. The majority of MaCDPKs generally exhibited similar expression patterns in the different tissues. Transcriptome analyses revealed that many CDPKs showed strong transcript accumulation at the early stages of fruit development and postharvest ripening in both varieties. Interaction network and co-expression analysis further identified some CDPKs-mediated network that was potentially active at the early stages of fruit development. Comparative expression analysis suggested that the high levels of CDPK expression in FJ might be related to its fast ripening characteristic. CDPK expression following the abiotic stress treatments indicated a significant transcriptional response to osmotic, cold, and salt treatment, as well as differential expression profiles, between BX and FJ. The findings of this study elucidate the transcriptional control of CDPKs in development, ripening, and the abiotic stress response in banana. Some tissue-specific, development/ripening-dependent, and abiotic stress-responsive candidate MaCDPK genes were identified for further genetic improvement of banana.

Differential gene regulatory pathways and co-expression networks associated with fire blight infection in apple (Malus \xd7 domestica)

Apple cultivars with durable resistance are needed for sustainable management of fire blight, the most destructive bacterial disease of apples. Although studies have identified genetic resistance to fire blight in both wild species and cultivated apples, more research is needed to understand the molecular mechanisms underlying host–pathogen interaction and differential genotypic responses to fire blight infection. We have analyzed phenotypic and transcriptional responses of ‘Empire’ and ‘Gala’ apple cultivars to fire blight by infecting them with a highly aggressive E. amylovora strain. Disease progress, based on the percentage of visual shoot necrosis, started showing significant (p < 0.001) differences between ‘Empire’ and ‘Gala’ 4 days after infection (dai). ‘Empire’ seems to slow down bacterial progress more rapidly after this point. We further compared transcriptome profiles of ‘Empire’ and ‘Gala’ at three different time points after fire blight infection. More genes showed differential expression in ‘Gala’ at earlier stages, but the number of differentially expressed genes increased in ‘Empire’ at 3 dai. Functional classes related to defense, cell cycle, response to stress, and biotic stress were identified and a few co-expression gene networks showed particular enrichment for plant defense and abiotic stress response genes. Several of these genes also co-localized in previously identified quantitative trait locus regions for fire blight resistance on linkage groups 7 and 12, and can serve as functional candidates for future research. These results highlight different molecular mechanisms for pathogen perception and control in two apple cultivars and will contribute toward better understanding of E. amylovora-Malus pathosystem.

Genome-wide identification and characterization of abiotic-stress responsive SOD (superoxide dismutase) gene family in Brassica juncea and B. rapa

BackgroundAbiotic stresses like drought, heat, cold and salinity cause major productivity loss in the rapeseed-mustard crops (Brassica). Major efforts have been made in the past to identify genes that provide resistance against such stresses. Superoxide dismutase (SOD) proteins, member of the metallo-enzyme family play vital role in protecting plants against abiotic stresses. In the present study, genome-wide analysis of abiotic stress responsive SOD gene family has been done in B. juncea and B. rapa.ResultsA total of 29 and 18 SOD genes were identified in B. juncea and B. rapa respectively and chromosome location mapping indicated their wide distribution across genome. On the basis of domain composition, the SODs were phylogenetically classified into sub-groups which was also substantiated by the gene structure and sub-cellular locations of SOD proteins. Functional annotation of SODs was also done by Gene Ontology (GO) mapping and the result was corroborated by the identified cis-regulatory elements in the promoter region of SOD genes. Based on FPKM analysis of SRA data available for drought, heat and salt stress, we identified 14 and 10 abiotic stress responsive SOD genes in B. rapa and B. juncea respectively. The differential expression analysis under drought and heat stress of identified abiotic-stress responsive SOD genes was done through quantitative Real Time PCR.ConclusionWe identified abiotic-stress responsive genes that could help in improving the plant tolerance against abiotic stresses. This was the first study to describe the genome-wide analysis of SOD gene family in B. rapa and B. juncea, and the results will help in laying basic ground for future work of cloning and functional validation of SOD genes during abiotic stresses leading to Brassica crop improvement.

Overexpression of Cotton a DTX/MATE Gene Enhances Drought, Salt, and Cold Stress Tolerance in Transgenic Arabidopsis.

Abiotic stresses have negative effects on plants growth and development. Plants, being sessile, have developed specific adaptive strategies that allow them to rapidly detect and respond to abiotic stress factors. The detoxification efflux carriers (DTX)/multidrug and toxic compound extrusion (MATE) transporters are of significance in the translocation of abscisic acid (ABA), a phytohormone with profound role in plants under various abiotic stress conditions. The ABA signaling cascades are the core regulators of abiotic stress responses in plants, triggering major changes in gene expression and adaptive physiological responses. We therefore carried out genome-wide analysis of the DTX/MATE gene family, transformed a DTX/MATE gene in Arabidopsis and carried out functional analysis under drought, salt, and cold stress conditions. We identified 128, 70, and 72 DTX/MATE genes in Gossypium hirsutum, Gossypium arboreum, and Gossypium raimondii, respectively. The proteins encoded by the DTX/MATE genes showed varied physiochemical properties but they all were hydrophobic. The Gh_D06G0281 (DTX/MATE) over-expressing Arabidopsis lines were highly tolerant under drought, salt, and cold stress with high production of antioxidant enzymes and significantly reduced levels of oxidants. Lipid peroxidation, as measured by the level of malondialdehyde concentrations was relatively low in transgenic lines compared to wild types, an indication of reduced oxidative stress levels in the transgenic plants. Based on physiological measurements, the transgenic plants exhibited significantly higher relative leaf water content, reduced excised leaf water loss and a significant reduction in ion leakage as a measure of the cell membrane stability compared to the wild types. Abiotic stress responsive genes, ABF4, CBL1, SOS1, and RD29B were highly expressed in the transgenic lines compared to the non-transformed wild type plants. The protein encoded by the Gh_D06G0281 (DTX/MATE) gene was predicted to be located within the plasma membrane. Since signals from extracellular stimuli are transmitted through the plasma membrane most of which are conducted by plasma membrane proteins it is possible the Gh_D06G0281 (DTX/MATE) gene product could be important for this process.

Overexpression of a MYB Family Gene, OsMYB6, Increases Drought and Salinity Stress Tolerance in Transgenic Rice.

MYB transcription factors have been demonstrated to play key regulatory roles in plant growth, development and abiotic stress response. However, knowledge concerning the involvement of rice MYB genes in salinity and drought stress resistance are largely unknown. In the present study, we cloned and characterized the OsMYB6 gene, which was induced by drought and salinity stress. Subcellular localization of OsMYB6-YFP fusion protein in protoplast cells indicated that OsMYB6 was localized in the nucleus. Overexpression of OsMYB6 in rice did not suggest a negative effect on the growth and development of transgenic plants, but OsMYB6-overexpressing plants showed increased tolerance to drought and salt stress compared with wild-type plants, as are evaluated by higher proline content, higher CAT and SOD activities, lower REL and MDA content in transgenic plants under drought and salt stress conditions. In addition, the expression of abiotic stress-responsive genes were significantly higher in OsMYB6 transgenic plants than that in wild-type plants under drought and salt stress conditions. These results indicate that OsMYB6 gene functions as a stress-responsive transcription factor which plays a positive regulatory role in response to drought and salt stress resistance, and may be used as a candidate gene for molecular breeding of salt-tolerant and drought-tolerant crop varieties.

Alternative splicing coupled to nonsense-mediated mRNA decay contributes to the high-altitude adaptation of maca (Lepidium meyenii)

Alpine plants remain the least studied plant communities in terrestrial ecosystems. However, how they adapt to high-altitude environments is far from clear. Here, we used RNA-seq to investigate a typical alpine plant maca (Lepidium meyenii) to understand its high-altitude adaptation at transcriptional and post-transcriptional level. At transcriptional level, we found that maca root significantly up-regulated plant immunity genes in day-time comparing to night-time, and up-regulated abiotic (cold/osmotic) stress response genes in Nov and Dec comparing to Oct. In addition, 17 positively selected genes were identified, which could be involved in mitochondrion. At post-transcriptional level, we found that maca had species-specific characterized alternative splicing (AS) profile which could be influenced by stress environments. For example, the alternative 3′ splice site events (A3SS, 39.62%) were predominate AS events in maca, rather than intron retention (IR, 23.17%). Interestingly, besides serine/arginine-rich (SR) proteins and long non-coding RNAs (lncRNAs), a lot of components in nonsense-mediated mRNA decay (NMD) were identified under differential alternative splicing (DAS), supporting AS coupled to NMD as essential mechanisms for maca's stress responses and high-altitude adaptation. Taken together, we first attempted to unveil maca's high-altitude adaptation mechanisms based on transcriptome and post-transcriptome evidence. Our data provided valuable insights to understand the high-altitude adaptation of alpine plants.

Selection of suitable reference genes for quantitative real-time PCR gene expression analysis in Mulberry (Morus alba L.) under different abiotic stresses.

Mulberry (Morus alba L.) is the sole food source for the mulberry silkworm, Bombyx mori and therefore important for sericulture industry. Different abiotic stress conditions like drought, salt, heat and cold stress adversely affect the productivity and quality of mulberry leaves. Quantitative real time PCR (qPCR) is a reliable and widely used method to identify abiotic stress responsive genes and molecular mechanism in different plant species. Selection of suitable reference genes is important requirement for normalizing the expression of genes through qRT-PCR study. In the present study, we have selected eight candidate reference genes in mulberry for analyzing their expression stability in different abiotic stress treatments including drought, salt, heat and cold stresses. The expression stability of these reference genes was determined using geNorm, NormFinder and RefFinder statistical algorithms. The results showed that Ubiquitin and protein phosphatase 2A regulatory subunit A (PP2A) were the most stable genes across all the treatment samples. However, analysis of individual stresses revealed different expression profiles and stability of reference genes. Actin3 and PP2A were most stable in drought and salt conditions respectively. RPL3 most preferred in heat stress and Ubiquitin was most stable in cold stress. We propose the ubiquitin and PP2A are the preferred reference genes for normalization of gene expression data from abiotic stresses. In addition, Actin3 are preferred for drought stress, PP2A for salt stress, RPL3 for heat stress and Ubiquitin for cold stress studies.

De novo assembly of Agave sisalana transcriptome in response to drought stress provides insight into the tolerance mechanisms

Agave, monocotyledonous succulent plants, is endemic to arid regions of North America, exhibiting exceptional tolerance to their xeric environments. They employ various strategies to overcome environmental constraints, such as crassulacean acid metabolism, wax depositions, and protective leaf morphology. Genomic resources of Agave species have received little attention irrespective of their cultural, economic and ecological importance, which so far prevented the understanding of the molecular bases underlying their adaptations to the arid environment. In this study, we aimed to elucidate molecular mechanism(s) using transcriptome sequencing of A. sisalana. A de novo approach was applied to assemble paired-end reads. The expression study unveiled 3,095 differentially expressed unigenes between well-irrigated and drought-stressed leaf samples. Gene ontology and KEGG analysis specified a significant number of abiotic stress responsive genes and pathways involved in processes like hormonal responses, antioxidant activity, response to stress stimuli, wax biosynthesis, and ROS metabolism. We also identified transcripts belonging to several families harboring important drought-responsive genes. Our study provides the first insight into the genomic structure of A. sisalana underlying adaptations to drought stress, thus providing diverse genetic resources for drought tolerance breeding research.

Overexpression of Rice Rab7 Gene Improves Drought and Heat Tolerance and Increases Grain Yield in Rice (Oryza sativa L.)

Rab family proteins play a crucial role in plant developmental processes and tolerance to environmental stresses. The current study investigated whether rice Rab7 (OsRab7) overexpression could improve rice tolerance to drought and heat stress conditions. The OsRab7 gene was cloned and transformed into rice plants. The survival rate, relative water content, chlorophyll content, gas-exchange characteristics, soluble protein content, soluble sugar content, proline content, and activities of antioxidant enzymes (CAT, SOD, APX, POD) of the transgenic rice lines were significantly higher than that of the wild-type. In contrast, the levels of hydrogen peroxide, electrolyte leakage, and malondialdehyde of the transgenic lines were significantly reduced when compared to wild-type. Furthermore, the expression of four genes encoding reactive oxygen species (ROS)-scavenging enzymes (OsCATA, OsCATB, OsAPX2, OsSOD-Cu/Zn) and eight genes conferring abiotic stress tolerance (OsLEA3, OsRD29A, OsSNAC1, OsSNAC2, OsDREB2A, OsDREB2B, OsRAB16A, OsRAB16C) was significantly up-regulated in the transformed rice lines as compared to their expression in wild-type. OsRab7 overexpression also increased grain yield in rice. Taken together, the current study indicates that the OsRab7 gene improves grain yield and enhances drought and heat tolerance in transgenic rice by modulating osmolytes, antioxidants and abiotic stress-responsive genes expression. Therefore, OsRab7 gene could be exploited as a promising candidate for improving rice grain yield and stress tolerance.

RSM1, an Arabidopsis MYB protein, interacts with HY5/HYH to modulate seed germination and seedling development in response to abscisic acid and salinity.

MYB transcription factors are involved in many biological processes, including metabolism, development and responses to biotic and abiotic stresses. RADIALIS-LIKE SANT/MYB 1 (RSM1) belongs to a MYB-related subfamily, and previous transcriptome analysis suggests that RSM1 may play roles in plant development, stress responses and plant hormone signaling. However, the molecular mechanisms of RSM1 action in response to abiotic stresses remain obscure. We show that down-regulation or up-regulation of RSM1 expression alters the sensitivity of seed germination and cotyledon greening to abscisic acid (ABA), NaCl and mannitol in Arabidopsis. The expression of RSM1 is dynamically regulated by ABA and NaCl. Transcription factors ELONGATED HYPOCOTYL 5 (HY5) and HY5 HOMOLOG (HYH) regulate RSM1 expression via binding to the RSM1 promoter. Genetic analyses reveal that RSM1 mediates multiple functions of HY5 in responses of seed germination, post-germination development to ABA and abiotic stresses, and seedling tolerance to salinity. Pull-down and BiFC assays show that RSM1 interacts with HY5/HYH in vitro and in vivo. RSM1 and HY5/HYH may function as a regulatory module in responses to ABA and abiotic stresses. RSM1 binds to the promoter of ABA INSENSITIVE 5 (ABI5), thereby regulating its expression, while RSM1 interaction also stimulates HY5 binding to the ABI5 promoter. However, no evidence was found in the dual-luciferase transient expression assay to support that RSM enhances the activation of ABI5 expression by HY. In summary, HY5/HYH and RSM1 may converge on the ABI5 promoter and independently or somehow dependently regulate ABI5 expression and ABI5-downstream ABA and abiotic stress-responsive genes, thereby improving the adaption of plants to the environment.

BpNAC012 Positively Regulates Abiotic Stress Responses and Secondary Wall Biosynthesis.

NAC (NAM, ATAF1/2, and CUC2) transcription factors play important roles in plant biological processes and stress responses. Here, we characterized the functional roles of BpNAC012 in white birch (Betula platyphylla). We found that BpNAC012 serves as a transcriptional activator. Gain- and loss-of-function analyses revealed that the transcript level of BpNAC012 was positively associated with salt and osmotic stress tolerance. BpNAC012 activated the core sequence CGT[G/A] to induce the expression of abiotic stress-responsive downstream genes, including Δ-1-pyrroline-5-carboxylate synthetase, superoxide dismutase, and peroxidase, resulting in enhanced salt and osmotic stress tolerance in BpNAC012 overexpression transgenic birch lines. We also showed that BpNAC012 is expressed predominantly in mature stems and that RNA interference-induced suppression of BpNAC012 caused a drastic reduction in the secondary wall thickening of stem fibers. Overexpression of BpNAC012 activated the expression of secondary wall-associated downstream genes by directly binding to the secondary wall NAC-binding element sites, resulting in ectopic secondary wall deposition in the stem epidermis. Moreover, salt and osmotic stresses elicited higher expression levels of lignin biosynthetic genes and elevated lignin accumulation in BpNAC012 overexpression lines. These findings provide insight into the functions of NAC transcription factors.

Characterization of DEAD-box family of RNA helicases in tomato provides insights into their roles in biotic and abiotic stresses

In plants, RNA helicases play significant roles in growth, development and stress response. In a previous study, a three-fold upregulation of a DEAD-box RNA helicase in a tomato cultivar tolerant to Tomato leaf curl New Delhi virus (ToLCNDV) as compared to susceptible cultivar during virus infection was shown. Given this, a comprehensive study was performed to identify the members of RNA helicase family in tomato and analyze their functional properties in response to abiotic stresses, hormone treatments and ToLCNDV infection. A total of 131 genes were identified and classified into DEAD- (42), DEAH- (38), and DExD/H-box (51) RNA helicases. Expression profiling of candidate genes in response to abiotic stresses and ToLCNDV infection in contrasting tomato cultivars suggested the putative roles of SlDEAD23 and SlDEAD35 in biotic and abiotic stresses. Heterologous overexpression of these genes in yeast enhanced the tolerance of transgenic cells to salt and cold stresses. Further, virus-induced silencing of SlDEAD35 in ToLCNDV tolerant cultivar resulted in susceptibility to virus infection, thus suggesting its involvement in tolerance mechanism. Altogether, this study provides novel insights into the structure, organization and involvement of DEAD-box RNA helicase genes in biotic and abiotic stress responses in tomato.

Integrated transcriptomic-metabolomic analysis reveals cellular responses of harvested strawberry fruit subjected to short-term exposure to high levels of carbon dioxide

Harvested strawberry (Fragaria × ananassa) fruit has a short shelf life due to rapid postharvest metabolism, quick softening, mechanical damage, and fungal decay. To improve the storability of strawberry fruit, we subjected harvested fruit to short-term exposure to 30% carbon dioxide (CO2) and used transcriptomic and metabolomic analyses to identify the cellular responses induced by this treatment. Fruit was stored at 10 °C for 10 d after a 3-h exposure to 30% CO2 (treatment) or ambient air (control) at 25 °C, respectively. The CO2 treatment reduced fruit decay and softening compared to the control throughout the 10-d storage period. Transcriptomic analyses revealed that expression levels of genes encoding cell wall-degrading enzymes (expansin, pectinesterase, and β-xylosidase) decreased in response to the CO2 treatment. Within 1 d after the CO2 treatment, the expression levels of genes encoding heat-shock proteins significantly increased. Metabolite profiling revealed that glucose, quinic acid, and succinic acid increased in responses to the CO2 treatment at 1 d. Transmission electron microscopy showed that disintegration of the middle lamella in the cell wall was inhibited by the CO2 treatment. Polyuronide (insoluble pectin) content in cell walls was 30% higher, on average, in the treated fruit than those in the control fruit during the 10-d storage period. These results indicate that a short-term treatment with 30% CO2 reduces the degradation of pectin in the cell wall by reducing the activity of cell wall-degrading enzymes and induces abiotic stress-response genes in harvested strawberry fruit. Our results improve our understanding of the molecular mechanisms behind CO2-responsive genes in strawberry fruit and provide insight into ways to improve the postharvest quality of strawberries.

Constitutive expression of REL1 confers the rice response to drought stress and abscisic acid

Leaf rolling is one of the most significant symptoms of drought stress in plant. Previously, we identified a dominant negative mutant, termed rolled and erect 1 (hereafter referred to rel1-D), regulating leaf rolling and erectness in rice. However, the role of REL1 in drought response is still poorly understood. Here, our results indicated that rel1-D displayed higher tolerance to drought relative to wild type, and the activity of superoxide dismutase (SOD) and drought responsive genes were significantly up-regulated in rel1-D. Moreover, our results revealed that rel1-D was hypersensitive to ABA and the expression of ABA associated genes was significantly increased in rel1-D, suggesting that REL1 likely coordinates ABA to regulate drought response. Using the RNA-seq approach, we identified a large group of differentially expressed genes that regulate stimuli and stresses response. Consistently, we also found that constitutive expression of REL1 alters the expression of biotic and abiotic stress responsive genes by the isobaric tags for relative and absolute quantification (iTRAQ) analysis. Integrative analysis demonstrated that 8 genes/proteins identified by both RNA-seq and iTRAQ would be the potential targets in term of the REL1-mediated leaf morphology. Together, we proposed that leaf rolling and drought tolerance of rel1-D under normal condition might be caused by the endogenously perturbed homeostasis derived from continuous stressful dynamics.

Winter-dormant shoot apical meristem in poplar trees shows environmental epigenetic memory

Trees have a long lifespan and must continually adapt to environmental pressures, notably in the context of climate change. Epigenetic mechanisms are doubtless involved in phenotypic plasticity and in stress memory; however, little evidence of the role of epigenetic processes is available for trees growing in fields. Here, we analyzed the possible involvement of epigenetic mechanisms in the winter-dormant shoot apical meristem of Populus × euramericana clones in memory of the growing conditions faced during the vegetative period. We aimed to estimate the range of genetic and environmentally induced variations in global DNA methylation and to evaluate their correlation with changes in biomass production, identify differentially methylated regions (DMRs), and characterize common DMRs between experiments. We showed that the variations in global DNA methylation between conditions were genotype dependent and correlated with biomass production capacity. Microarray chip analysis allowed detection of DMRs 6 months after the stressful summer period. The 161 DMRs identified as common to three independent experiments most notably targeted abiotic stress and developmental response genes. Results are consistent with a winter-dormant shoot apical meristem epigenetic memory of stressful environmental conditions that occurred during the preceding summer period. This memory may facilitate tree acclimation.

INDETERMINATE SPIKELET1 Recruits Histone Deacetylase and a Transcriptional Repression Complex to Regulate Rice Salt Tolerance.

Perception and transduction of salt stress signals are critical for plant survival, growth, and propagation. Thus, identification of components of the salt stress-signaling pathway is important for rice (Oryza sativa) molecular breeding of salt stress resistance. Here, we report the identification of an apetala2/ethylene response factor transcription factor INDETERMINATE SPIKELET1 (IDS1) and its roles in the regulation of rice salt tolerance. By genetic screening and phenotype analysis, we demonstrated that IDS1 conferred transcriptional repression activity and acted as a negative regulator of salt tolerance in rice. To identify potential downstream target genes regulated by IDS1, we conducted chromatin immunoprecipitation (ChIP) sequencing and ChIP-quantitative PCR assays and found that IDS1 may directly associate with the GCC-box-containing motifs in the promoter regions of abiotic stress-responsive genes, including LEA1 (LATE EMBRYOGENESIS ABUNDANT PROTEIN1) and SOS1 (SALT OVERLY SENSITIVE1), which are key genes regulating rice salt tolerance. IDS1 physically interacted with the transcriptional corepressor topless-related 1 and the histone deacetylase HDA1, contributing to the repression of LEA1 and SOS1 expression. Analyses of histone H3 acetylation status and RNA polymerase II occupation on the promoters of LEA1 and SOS1 further defined the molecular foundation of the transcriptional repression activity of IDS1. Our findings illustrate an epigenetic mechanism by which IDS1 modulates salt stress signaling as well as salt tolerance in rice.

Functional characterization of transposon-tagged abiotic stress-responsive rice genes and their molecular polymorphisms among various stress-tolerant genotypes

Dissociation (Ds) insertional mutagenesis has been regarded as an efficient tool to generate insertion mutants for functional genomics and molecular breeding. However, little is known about the application of the tool on exploring biological functions of abiotic stress-related genes and their molecular breeding experience. In this study, a total of 833 Ds insertion lines have been obtained, which showed significantly higher tolerance or sensitivity to high salinity, drought or cold stress, by screening around 20,000 Ds lines. Analysis of Ds flanking sequence tags revealed that 165 rice genes were tagged by Ds insertion. Gene Ontology (GO) and gene set enrichment analysis showed that over-represented Ds-tagged genes might function in the response to exogenous stimuli. These Ds-tagged genes showed expression divergence among five high salinity and five drought tolerant rice lines under either high salinity or drought stress. Higher percentages of Ds-tagged genes were down- or up-regulated by these abiotic stresses. These Ds-tagged genes were also frequently reduced or suppressed by various phytohormones including abscisic acid and jasmonate. On the other hand, we have also detected single nucleotide polymorphisms (SNPs) and 1–10 base pairs of insertion and deletions (indels) of these Ds-tagged genes among ten high salinity/drought tolerant rice lines by comparing with the reference genome Nipponbare. Our data showed that SNPs were detected among 102 out of 165 genes and indels were identified in 39 out of 165 genes. All the data provided additional information to further explore the biological functions of these genes or to carry out molecular breeding.

The Grape VlWRKY3 Gene Promotes Abiotic and Biotic Stress Tolerance in Transgenic Arabidopsis thaliana.

WRKY transcription factors are known to play important roles in plant responses to various abiotic and biotic stresses. The grape WRKY gene, WRKY3 was previously reported to respond to salt and drought stress, as well as methyl jasmonate and ethylene treatments in Vitis labrusca × V. vinifera cv. ‘Kyoho.’ In the current study, WRKY3 from the ‘Kyoho’ grape cultivar was constitutively expressed in Arabidopsis thaliana under control of the cauliflower mosaic virus 35S promoter. The 35S::VlWRKY3 transgenic A. thaliana plants showed improved salt and drought stress tolerance during the germination, seedling and the mature plant stages. Various physiological traits related to abiotic stress responses were evaluated to gain further insight into the role of VlWRKY3, and it was found that abiotic stress caused less damage to the transgenic seedlings than to the wild-type (WT) plants. VlWRKY3 over-expression also resulted in altered expression levels of abiotic stress-responsive genes. Moreover, the 35S::VlWRKY3 transgenic A. thaliana lines showed improved resistance to Golovinomyces cichoracearum, but increased susceptibility to Botrytis cinerea, compared with the WT plants. Collectively, these results indicate that VlWRKY3 plays important roles in responses to both abiotic and biotic stress, and modification of its expression may represent a strategy to enhance stress tolerance in crops.

Identification and characterization of a novel abiotic stress responsive OsTHIC gene from rice

ABSTRACTTo better understand the mechanisms of plant abiotic stress responses and identify novel stress-related genes in rice, we performed global expression analysis in indica rice Pei'ai 64S under multiple stresses. Among numerous genes, a gene named OsTHIC was selected, which was highly induced in leaves and panicles in response to all stresses at different developmental stages, especially at the booting stage under cold stress. OsTHIC contains an open reading frame (ORF) of 1920 bp and encodes a predicted protein with 639 amino acid residues. The molecular weight and isoelectric point were predicted to be about 71.31 kD and 6.41, respectively. The results of quantitative real-time polymerase chain reaction (qRT-PCR) analysis were almost identical to those from the GeneChip Rice Genome Array. The sequence alignment showed about 99% similarity to rice phosphate methylpyrimidine synthase. Promoter sequence analysis showed that various stress response-related cis-elements were contained in the promoter region. All the results suggested that OsTHIC may be involved in rice stress responses.

Tobacco Transcription Factor NtbHLH123 Confers Tolerance to Cold Stress by Regulating the NtCBF Pathway and Reactive Oxygen Species Homeostasis.

Cold stress is a major environmental factor that impairs plant growth and development, geographic distribution, and crop productivity. The C-repeat binding factor (CBF) regulatory pathway has an essential role in response to cold stress. Here, we characterized a bHLH transcription factor from Nicotiana tabacum, NtbHLH123, in response to cold stress (4°C). Overexpression of NtbHLH123 enhanced cold tolerance in transgenic tobacco plants. Based on yeast one-hybrid, chromatin immunoprecipitation PCR, and transient expression analysis assays, NtbHLH123 binds directly to the G-box/E-box motifs in the promoter of the NtCBF genes and positively regulates their expression. Furthermore, NtbHLH123-overexpressing plants showed lower electrolyte leakage, reduced malondialdehyde contents, H2O2 and reactive oxygen species (ROS) accumulation under cold stress, which contributed to alleviating oxidative damage to the cell membrane after cold stress treatment. And NtbHLH123 increased stress tolerance by improving the expression of a number of abiotic stress-responsive genes to mediate the ROS scavenging ability and other stress tolerance pathways. Taken together, we present a model suggesting that NtbHLH123 is a transcriptional activator that functions as a positive regulator of cold tolerance by activating NtCBF, ROS scavenging-related, and stress-responsive genes.