Constitutive overexpression of the calcium sensor CBL5 confers osmotic or drought stress tolerance in Arabidopsis

Calcium serves as a critical messenger in many adaptation and developmental processes. Cellular calcium signals are detected and transmitted by sensor molecules such as calcium-binding proteins. In plants, the calcineurin B-like protein (CBL) family represents a unique group of calcium sensors and plays a key role in decoding calcium transients by specifically interacting with and regulating a family of CBL-interacting protein kinases (CIPKs). In this study, we report the role of Arabidopsis CBL5 gene in high salt or drought tolerance. CBL5 gene is expressed significantly in green tissues, but not in roots. CBL5 was not induced by abiotic stress conditions such as high salt, drought or low temperature. To determine whether the CBL5 gene plays a role in stress response pathways, we ectopically expressed the CBL5 protein in transgenic Arabidopsis plants (35S-CBL5) and examined plant responses to abiotic stresses. CBL5-overexpressing plants displayed enhanced tolerance to high salt or drought stress. CBL5 overexpression also rendered plants more resistant to high salt or hyperosmotic stress during early development (i.e., seed germination) but did not alter their response to abiscisic acid (ABA). Furthermore, overexpression of CBL5 alters the gene expression of stress gene markers, such as RD29A, RD29B and Kin1 etc. These results suggest that CBL5 may function as a positive regulator of salt or drought responses in plants.     

An osmotin from the resurrection plant Tripogon loliiformis (TlOsm) confers tolerance to multiple abiotic stresses in transgenic rice

Osmotin is a key protein associated with abiotic and biotic stress response in plants. In this study, an osmotin from the resurrection plant Tripogon loliiformis (TlOsm) was characterized and functionally analyzed under abiotic stress conditions in T. loliiformis as well as in transgenic Nicotiana tabacum (tobacco) and Oryza sativa (rice) plants. Real‐time PCR analysis on mixed elicitor cDNA libraries from T. loliiformis showed that TlOsm was upregulated a 1000‐fold during the early stages of osmotic stresses (cold, drought, and salinity) in both shoots and roots but downregulated in shoots during heat stress. There was no change in TlOsm gene expression in roots of heat‐stressed plants and during plant development. The plasma membrane localization of TlOsm was showed in fluorescent‐tagged TlOsm tobacco plants using confocal laser scanning microscopic analysis. Transgenic rice plants expressing TlOsm were assessed for enhanced tolerance to salinity, drought and cold stresses. Constitutively expressed TlOsm in transgenic rice plants showed increased tolerance to cold, drought and salinity stress when compared with the wild‐type and vector control counterparts. This was evidenced by maintained growth, retained higher water content and membrane integrity, and improved survival rate of TlOsm‐expressing plants. The results thus indicate the involvement of TlOsm in plant response to multiple abiotic stresses, possibly through the signaling pathway, and highlight its potential applications for engineering crops with improved tolerance to cold, drought and salinity stress.     

Identification of a drought responsive gene encoding a nuclear protein involved in drought and freezing stress tolerance in Arabidopsis

Plants have developed adaptive strategies to survive under different abiotic stressors. To identify new components involved in abiotic stress tolerance, we screened unannotated expressed sequence tags (ESTs) and evaluated their cold or drought response in Arabidopsis. We identified a drought response gene (DRG) encoding a 39.5-kDa polypeptide. This protein was expressed specifically in siliques and was induced by drought stress in most tissues. When a DRG-GFP construct was introduced into Arabidopsis protoplasts, GFP signals were detected only in the nucleus. The drg mutant plant was more sensitive to mannitol-induced osmotic stress in agar plates and to drought or freezing stress in soil than the wild-type. Activating the DRG restored the normal sensitivity of drg mutants to abiotic stressors. No differences in drought or freezing tolerance were observed between the wild-type and transgenic plants overexpressing the DRG. When DRG was expressed in a cold-sensitive Escherichia coli strain BX04, the transformed bacteria grew faster than the untransformed BXO4 cells under cold stress. These results demonstrate that DRG is a nuclear protein induced by abiotic stresses and it is required for drought and freezing tolerance in Arabidopsis.     

Chitinase Gene Expression in Response to Environmental Stresses in Arabidopsis thaliana: Chitinase Inhibitor Allosamidin Enhances Stress Tolerance

The expression levels of three chitinase genes in Arabidopsis thaliana, AtChiA (class III), AtChiB (class I), and AtChiV (class IV), were examined under various stress conditions by semi-quantitative RT-PCR. Under normal growth conditions, the AtChiB and AtChiV genes were expressed in most organs of Arabidopsis plants at all growth stages, whereas the AtChiA gene was not expressed at all. The class III AtChiA gene was expressed exclusively when the plants were exposed to environmental stresses, especially to salt and wound stresses. Treatment of Arabidopsis plants with allosamidin, which inhibits class III chitinases, did not affect the growth rate. Surprisingly, however, the plants treated with allosamidin were more tolerant of abiotic stresses (cold, freezing, heat, and strong light) than the control plants. It also appeared that allosamidin enhances AtChiA and AtChiB expression under heat and strong light stresses. Allosamidin is likely to enhance abiotic stress tolerance, probably through crosstalk between the two signaling pathways for biotic and abiotic stress responses.     

Characterization and expression analysis of the <Emphasis Type="Italic">SNF2</Emphasis> family genes in response to phytohormones and abiotic stresses in rice

The function of SNF2 ATPases, the major catalytic subunits of chromatin remodeling complexes, in plants is not sufficiently understood. Here we identified 39 putative SNF2 genes of rice (Oryza sativa L.) by homology analyses and analyzed the expression profiles of eight of them in response to phytohormones and abiotic stresses. Our results indicated that expression of the SNF2 genes was affected by auxin, gibberellin, cytokinin, abscisic acid, ethylene, and some abiotic stresses such as heat, chilling, darkness, drought and salinity. It suggests that, like Arabidopsis SNF2s, rice SNF2 proteins may function in phytohormone signaling pathways and/or be associated with the resistance to abiotic stresses, but in distinct manners from their Arabidopsis orthologs. Some SNF2 proteins in rice may be involved in cross-talk of the signaling pathways between phytohormones and abiotic stresses.     

水稻NAM基因家族的全基因组鉴定及表达分析

植物特异性转录因子NAM家族从属于NAC转录因子超家族,在植株生长发育、生理代谢以及应对各种胁迫反应中均发挥重要作用。该研究采用生物信息学方法鉴定水稻基因组中的NAM基因,分析其时空表达模式、亚细胞定位以及蛋白相互作用,并采用实时定量qRT PCR方法分析不同外源激素(如SA、ABA和MeJA)以及非生物胁迫(包括干旱、盐和冷)处理下各NAM基因的表达特征,为进一步探索NAM基因在非生物胁迫中的功能和应激机制以及激素调控途径奠定基础。结果显示：(1)从水稻基因组中共鉴定出48个NAM基因,进化分析将其分为5个亚家族;NAM基因在水稻基因组中存在9对片段复制事件。(2)组织表达分析显示,NAM基因在水稻不同组织及发育时期表现特异性表达,特别是叶鞘、茎和节的生长过程中高表达,且大多数是核定位,并存在多种蛋白互作。(3)实时定量qRT PCR表达分析显示,10个NAM基因在不同组织中均特异表达;大部分NAM基因在盐和干旱胁迫下表达上调,而在冷胁迫下表达降低;SA、ABA和MeJA处理均可显著改变各NAM基因的表达水平。研究表明,NAM基因在水稻生长发育、激素应答和非生物胁迫响应中具有重要作用。     

Identification of MsHsp23 gene using annealing control primer system

To identify potential candidates for acquiring stress tolerance, a new annealing control primer (ACP) system was used to identify the differentially expressed genes. Alfalfa (Medicago sativa L.) seedlings were exposed to various abiotic stresses such as cold (4°C for 6 h), heat (42°C for 6 h), salt (300 mM for 6 h), drought (withdrawing irrigation for 48 h), copper (500 μM for 6 h), cadmium (500 μM for 6 h), and arsenic (500 μM for 6 h). Primer sets 41 and 93 were differentially expressed and identified as same sequence, which represents a mitochondrial small heat-shock protein encoding gene, MsHsp23. This band was markedly increased or induced in alfalfa under heat, salt, and arsenic stresses. Differential expression of MsHsp23 was further evaluated by Northern blot analysis. Temporal expression analysis showed that mRNA pool was altered as early as 1 h of treatment. Thus, differential accumulation of MsHsp23 under heat, salt, and arsenic stresses suggests its potential involvement in diverse abiotic stress tolerance, and thereby making a target for further molecular analysis.     

Overexpression of a <Emphasis Type="Italic">Panax ginseng</Emphasis> tonoplast aquaporin alters salt tolerance,drought tolerance and cold acclimation ability in transgenic <Emphasis Type="Italic">Arabidopsis</Emphasis> plants

Water movement across cellular membranes is regulated largely by a family of water channel proteins called aquaporins (AQPs). Since several abiotic stresses such as, drought, salinity and freezing, manifest themselves via altering water status of plant cells and are linked by the fact that they all result in cellular dehydration, we overexpressed an AQP (tonoplast intrinsic protein) from Panax ginseng, PgTIP1, in transgenic Arabidopsis thaliana plants to test its role in plant’s response to drought, salinity and cold acclimation (induced freezing tolerance). Under favorable conditions, PgTIP1 overexpression significantly increased plant growth as determined by the biomass production, and leaf and root morphology. PgTIP1 overexpression had beneficial effect on salt-stress tolerance as indicated by superior growth status and seed germination of transgenic plants under salt stress; shoots of salt-stressed transgenic plants also accumulated greater amounts of Na⁺ compared to wild-type plants. Whereas PgTIP1 overexpression diminished the water-deficit tolerance of plants grown in shallow (10 cm deep) pots, the transgenic plants were significantly more tolerant to water stress when grown in 45 cm deep pots. The rationale for this contrasting response, apparently, comes from the differences in the root morphology and leaf water channel activity (speed of dehydration/rehydration) between the transgenic and wild-type plants. Plants overexpressed with PgTIP1 exhibited lower (relative to wild-type control) cold acclimation ability; however, this response was independent of cold-regulated gene expression. Our results demonstrate a significant function of PgTIP1 in growth and development of plant cells, and suggest that the water movement across tonoplast (via AQP) represents a rate-limiting factor for plant vigor under favorable growth conditions and also significantly affect responses of plant to drought, salt and cold stresses.