Overexpression of <Emphasis Type="Italic">AtLEA3-3</Emphasis> confers resistance to cold stress in <Emphasis Type="Italic">Escherichia coli</Emphasis> and provides enhanced osmotic stress tolerance and ABA sensitivity in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Previous studies have shown that the late embryogenesis abundant (LEA) group 3 proteins significantly respond to changes in environmental conditions. However, reports that demonstrate their biological role, especially in Arabidopsis, are notably limited. This study examines the functional roles of the Arabidopsis LEA group 3 proteins AtLEA3-3 and AtLEA3-4 in abiotic stress and ABA treatments. Expression of AtLEA3-3 and AtLEA3-4 is upregulated by ABA, high salinity, and osmotic stress. Results on the ectopic expression of AtLEA3-3 and AtLEA3-4 in E. coli suggest that both proteins play important roles in resistance to cold stress. Overexpression of AtLEA3-3 in Arabidopsis (AtLEA3-3-OE) confers salt and osmotic stress tolerance that is characterized during germination and early seedling establishment. However, AtLEA3-3-OE lines show sensitivity to ABA treatment during early seedling development. These results suggest that accumulation of AtLEA3-3 mRNA and/or proteins may help heterologous ABA reinitiate second dormancy during seedling establishment. Analysis of yellow fluorescent fusion proteins localization shows that AtLEA3-3 and AtLEA3-4 are mainly distributed in the ER and that AtLEA3-3 also localizes in the nucleus, and in response to salt, mannitol, cold, or BFA treatments, the localization of AtLEA3-3 and AtLEA3-4 is altered and becomes more condensed. Protein translocalization may be a positive and effective strategy for responding to abiotic stresses. Taken together, these results suggest that AtLEA3-3 has an important function during seed germination and seedling development of Arabidopsis under abiotic stress conditions.     

Overexpression of CaDSR6 increases tolerance to drought and salt stresses in transgenic Arabidopsis plants

The partial CaDSR6 (Capsicum annuum Drought Stress Responsive 6) cDNA was previously identified as a drought-induced gene in hot pepper root tissues. However, the cellular role of CaDSR6 with regard to drought stress tolerance was unknown. In this report, full-length CaDSR6 cDNA was isolated. The deduced CaDSR6 protein was composed of 234 amino acids and contained an approximately 30 amino acid-long Asp-rich domain in its central region. This Asp-rich domain was highly conserved in all plant DSR6 homologs identified and shared a sequence identity with the N-terminal regions of yeast p23^fyp and human hTCTP, which contain Rab protein binding sites. Transgenic Arabidopsis plants overexpressing CaDSR6 (35S:CaDSR6-sGFP) were tolerant to high salinity, as identified by more vigorous root growth and higher levels of total chlorophyll than wild type plants. CaDSR6-overexpressors were also more tolerant to drought stress compared to wild type plants. The 35S:CaDSR6-sGFP leaves retained their water content and chlorophyll more efficiently than wild type leaves in response to dehydration stress. The expression of drought-induced marker genes, such as RD20, RD22, RD26, RD29A, RD29B, RAB18, KIN2, ABF3, and ABI5, was markedly increased in CaDSR6-overexpressing plants relative to wild type plants under both normal and drought conditions. These results suggest that overexpression of CaDSR6 is associated with increased levels of stress-induced genes, which, in turn, conferred a drought tolerant phenotype in transgenic Arabidopsis plants. Overall, our data suggest that CaDSR6 plays a positive role in the response to drought and salt stresses.     

The wheat gene <Emphasis Type="Italic">TaST</Emphasis> can increase the salt tolerance of transgenic <Emphasis Type="Italic">Arabidopsis</Emphasis>

On the basis of the results of gene chip analysis of the salt-tolerant wheat mutant RH8706-49 under conditions of salt stress, we identified and cloned an unknown salt-induced gene TaST (Triticum aestivum salt-tolerant). Real-time quantitative PCR analysis showed that the expression of the gene was induced by salt stress. Transgenic Arabidopsis plants overexpressing the TaST gene showed higher salt tolerance than the wild-type controls. Subcellular localization studies revealed that the protein encoded by this gene was in the nucleus. In comparison with wild-type controls, transgenic Arabidopsis plants accumulated more Ca²⁺, soluble sugar, and proline and less Na⁺ under salt stress. Real-time quantitative PCR analysis showed that Arabidopsis plants overexpressing TaST also showed increased expression of many stress-related genes. All these findings indicated that TaST can enhance the salt tolerance of transgenic Arabidopsis plants.     

Arabidopsis STAYGREEN-LIKE (SGRL) promotes abiotic stress-induced leaf yellowing during vegetative growth

During leaf senescence in Arabidopsis, STAYGREEN 1 (SGR1) and SGR2 regulate chlorophyll degradation positively and negatively, respectively. SGR-LIKE (SGRL) is also expressed in pre-senescing leaves, but its function remains largely unknown. Here we show that under abiotic stress, Arabidopsis plants overexpressing SGRL exhibit early leaf yellowing and sgrl-1 mutants exhibit persistent green color of leaves. Under salt stress, SGR1 and SGRL act synergistically for rapid Chl degradation prior to senescence. Furthermore, SGRL forms homo- and heterodimers with SGR1 and SGR2 in vivo, and interacts with LHCII and chlorophyll catabolic enzymes. The role of SGRL under abiotic stress is discussed.     

GsSRK, a G-type lectin S-receptor-like serine/threonine protein kinase,is a positive regulator of plant tolerance to salt stress

Receptor-like protein kinases (RLKs) play vital roles in sensing outside signals, yet little is known about RLKs functions and roles in stress signal perception and transduction in plants, especially in wild soybean. Through the microarray analysis, GsSRK was identified as an alkaline (NaHCO₃)-responsive gene, and was subsequently isolated from Glycine soja by homologous cloning. GsSRK encodes a 93.22 kDa protein with a highly conserved serine/threonine protein kinase catalytic domain, a G-type lectin region, and an S-locus region. Real-time PCR results showed that the expression levels of GsSRK were largely induced by ABA, salt, and drought stresses. Over expression of GsSRK in Arabidopsis promoted seed germination, as well as primary root and rosette leaf growth during the early stages of salt stress. Compared to the wild type Arabidopsis, GsSRK overexpressors exhibited enhanced salt tolerance and higher yields under salt stress, with higher chlorophyll content, lower ion leakage, higher plant height, and more siliques at the adult developmental stage. Our studies suggest that GsSRK plays a crucial role in plant response to salt stress.     

Cytokinin oxidase/dehydrogenase overexpression modifies antioxidant defense against heat,drought and their combination in Nicotiana tabacum plants

Cytokinins (CKs) as well as the antioxidant enzyme system (AES) play important roles in plant stress responses. The expression and activity of antioxidant enzymes (AE) were determined in drought, heat and combination of both stresses, comparing the response of tobacco plants overexpressing the main cytokinin degrading enzyme, cytokinin oxidase/dehydrogenase, under the control of root-specific WRKY6 promoter (W6:CKX1 plants) or constitutive promoter (35S:CKX1 plants) and the corresponding wild-type (WT). Expression levels as well as activities of cytosolic ascorbate peroxidase, catalase 3, and cytosolic superoxide dismutase were low under optimal conditions and increased after heat and combined stress in all genotypes. Unlike catalase 3, two other peroxisomal enzymes, catalase 1 and catalase 2, were transcribed extensively under control conditions. Heat stress, in contrast to drought or combined stress, increased catalase 1 and reduced catalase 2 expression in WT and W6:CKX1 plants. In 35S:CKX1, catalase 1 expression was enhanced by heat or drought, but not under combined stress conditions. Mitochondrial superoxide dismutase expression was generally higher in 35S:CKX1 plants than in WT. Genes encoding for chloroplastic AEs, stromatal ascorbate peroxidase, thylakoidal ascorbate peroxidase and chloroplastic superoxide dismutase, were strongly transcribed under control conditions. All stresses down-regulated their expression in WT and W6:CKX1, whereas more stress-tolerant 35S:CKX1 plants maintained high expression during drought and heat. The achieved data show that the effect of down-regulation of CK levels on AES may be mediated by altered habit, resulting in improved stress tolerance, which is associated with diminished stress impact on photosynthesis, and changes in source/sink relations.     

ABA Inducible Rice Protein Phosphatase 2C Confers ABA Insensitivity and Abiotic Stress Tolerance in Arabidopsis

Arabidopsis PP2C belonging to group A have been extensively worked out and known to negatively regulate ABA signaling. However, rice (Oryza sativa) orthologs of Arabidopsis group A PP2C are scarcely characterized functionally. We have identified a group A PP2C from rice (OsPP108), which is highly inducible under ABA, salt and drought stresses and localized predominantly in the nucleus. Genetic analysis revealed that Arabidopsis plants overexpressing OsPP108 are highly insensitive to ABA and tolerant to high salt and mannitol stresses during seed germination, root growth and overall seedling growth. At adult stage, OsPP108 overexpression leads to high tolerance to salt, mannitol and drought stresses with far better physiological parameters such as water loss, fresh weight, chlorophyll content and photosynthetic potential (Fv/Fm) in transgenic Arabidopsis plants. Expression profile of various stress marker genes in OsPP108 overexpressing plants revealed interplay of ABA dependent and independent pathway for abiotic stress tolerance. Overall, this study has identified a potential rice group A PP2C, which regulates ABA signaling negatively and abiotic stress signaling positively. Transgenic rice plants overexpressing this gene might provide an answer to the problem of low crop yield and productivity during adverse environmental conditions.     

A salt-inducible chloroplastic monodehydroascorbate reductase from halophyte Avicennia marina confers salt stress tolerance on transgenic plants

Plant growth and productivity are adversely affected by various abiotic stress factors. In our previous study, we used Avicennia marina, a halophytic mangrove, as a model plant system for isolating genes functioning in salt stress tolerance. A large scale random EST sequencing from a salt stressed leaf tissue cDNA library of one month old A. marina plants resulted in identification of a clone showing maximum homology to Monodehydroascorbate reductase (Am-MDAR). MDAR plays a key role in regeneration of ascorbate from monodehydroascorbate for ROS scavenging. In this paper, we report the cellular localization and the ability to confer salt stress tolerance in transgenic tobacco of this salt inducible Am-MDAR. A transit peptide at the N-terminal region of Am-MDAR suggested that it encodes a chloroplastic isoform. The chloroplastic localization was confirmed by stable transformation and expression of the Am-MDAR-GFP fusion protein in tobacco. Transgenic tobacco plants overexpressing Am-MDAR survived better under conditions of salt stress compared to untransformed control plants. Assays of enzymes involved in ascorbate–glutathione cycle revealed an enhanced activity of MDAR and ascorbate peroxidase whereas the activity of dehyroascorbate reductase was reduced under salt stressed and unstressed conditions in Am-MDAR transgenic lines. The transgenic lines showed an enhanced redox state of ascorbate and reduced levels of malondialdehyde indicating its enhanced tolerance to oxidative stress. The results of our studies could be used as a starting point for genetic engineering of economically important plants tolerant to salt stress.     

Overexpression of maize mitogen-activated protein kinase gene, ZmSIMK1 in Arabidopsis increases tolerance to salt stress

Mitogen-activated protein kinase (MAPK) cascades play a remarkably crucial role in plants. It has been studied intensively in model plants Arabidopsis, tobacco and rice. However, the function of MAPKs in maize (Zea mays L.) has not been well documented. ZmSIMK1 (Zea mays salt-induced mitogen-activated protein kinase 1) is a previously identified MAPK gene in maize. In this research, we charactered ZmSIMK1 and showed that ZmSIMK1 was involved in Arabidopsis salt stress. The genomic organization of ZmSIMK1 gene and its expression in maize have been analyzed. In order to investigate the function of ZmSIMK1, we generated transgenic Arabidopsis constitutively overexpressing ZmSIMK1. Ectopic expression of ZmSIMK1 in Arabidopsis resulted in increased resistance against salt stress. Importantly, ZmSIMK1-overexpressing Arabidopsis exhibited constitutive expression of stress-responsive marker genes, RD29A and P5CS1. Furthermore, RD29A and P5CS1 were upregulated under salt stress. These results suggest that ZmSIMK1 may play an important role in plant salt stress.     

Comparative analysis of GT14/GT14-like gene family in Arabidopsis, Oryza, Populus, Sorghum and Vitis

Glycosyltransferase family14 (GT14) belongs to the glycosyltransferase (GT) superfamily that plays important roles in the biosynthesis of cell walls, the most abundant source of cellulosic biomass for bioethanol production. It has been hypothesized that DUF266 proteins are a new class of GTs related to GT14. In this study, we identified 62 GT14 and 106 DUF266 genes (named GT14-like herein) in Arabidopsis, Oryza, Populus, Sorghum and Vitis. Our phylogenetic analysis separated GT14 and GT14-like genes into two distinct clades, which were further divided into eight and five groups, respectively. Similarities in protein domain, 3D structure and gene expression were uncovered between the two phylogenetic clades, supporting the hypothesis that GT14 and GT14-like genes belong to one family. Therefore, we proposed a new family name, GT14/GT14-like family that combines both subfamilies. Variation in gene expression and protein subcellular localization within the GT14-like subfamily were greater than those within the GT14 subfamily. One-half of the Arabidopsis and Populus GT14/GT14-like genes were found to be preferentially expressed in stem/xylem, indicating that they are likely involved in cell wall biosynthesis. This study provided new insights into the evolution and functional diversification of the GT14/GT14-like family genes.     

Ectopic expression of ADP ribosylation factor 1 (SaARF1) from smooth cordgrass (Spartina alterniflora Loisel) confers drought and salt tolerance in transgenic rice and Arabidopsis

Salinity and drought are two very important abiotic stressors that negatively impact the growth and yield of all sensitive crop plants. Genes from halophytes have been shown to be useful to engineer crop plants that can survive under adverse soil and water conditions. The present report establishes, for the first time, the physiological role of a class one ADP ribosylation factor gene (SaARF1) from the halophyte Spartina alterniflora (smooth cordgrass) in imparting salinity and drought stress tolerance when expressed in both monocot (rice) and dicot (Arabidopsis) systems. The Arabidopsis and rice plants overexpressing ARF1 are many-fold more tolerant to salt and drought than wild-type (WT) plants. The transgenics exhibited improved growth and productivity relative to WT through tissue tolerance by maintaining higher relative water content and membrane stability, and higher photosynthetic yield by retaining higher chlorophyll concentration and fluorescence under stress conditions compared to WT. These findings indicated that genes from halophyte resources can be useful to engineer and improve salt and drought stress tolerance in both monocot and dicot plants.