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.     

Expression of the Kale WRKY Gene BoWRKY10 in Transgenic Tobacco Confers Drought Stress Tolerance

Russian Journal of Plant Physiology - WRKYs play important roles in plant growth, defense regulation, and the stress response. However, the mechanisms through which WRKYs are involved in drought...     

Expression of a Late Embryogenesis Abundant Protein Gene,HVA1, from Barley Confers Tolerance to Water Deficit and Salt Stress in Transgenic Rice

A late embryogenesis abundant (LEA) protein gene, HVA1, from barley (Hordeum vulgare L.) was introduced into rice suspension cells using the Biolistic-mediated transformation method, and a large number of independent transgenic rice (Oryza sativa L.) plants were generated. Expression of the barley HVA1 gene regulated by the rice actin 1 gene promoter led to high-level, constitutive accumulation of the HVA1 protein in both leaves and roots of transgenic rice plants. Second-generation transgenic rice plants showed significantly increased tolerance to water deficit and salinity. Transgenic rice plants maintained higher growth rates than nontransformed control plants under stress conditions. The increased tolerance was also reflected by delayed development of damage symptoms caused by stress and by improved recovery upon the removal of stress conditions. We also found that the extent of increased stress tolerance correlated with the level of the HVA1 protein accumulated in the transgenic rice plants. Using a transgenic approach, this study provides direct evidence supporting the hypothesis that LEA proteins play an important role in the protection of plants under water-or salt-stress conditions. Thus, LEA genes hold considerable potential for use as molecular tools for genetic crop improvement toward stress tolerance.     

The SbMT-2 Gene from a Halophyte Confers Abiotic Stress Tolerance and Modulates ROS Scavenging in Transgenic Tobacco

Heavy metals are common pollutants of the coastal saline area and Salicornia brachiata an extreme halophyte is frequently exposed to various abiotic stresses including heavy metals. The SbMT-2 gene was cloned and transformed to tobacco for the functional validation. Transgenic tobacco lines (L2, L4, L6 and L13) showed significantly enhanced salt (NaCl), osmotic (PEG) and metals (Zn⁺⁺, Cu⁺⁺ and Cd⁺⁺) tolerance compared to WT plants. Transgenic lines did not show any morphological variation and had enhanced growth parameters viz. shoot length, root length, fresh weight and dry weight. High seed germination percentage, chlorophyll content, relative water content, electrolytic leakage and membrane stability index confirmed that transgenic lines performed better under salt (NaCl), osmotic (PEG) and metals (Zn⁺⁺, Cu⁺⁺ and Cd⁺⁺) stress conditions compared to WT plants. Proline, H₂O₂ and lipid peroxidation (MDA) analyses suggested the role of SbMT-2 in cellular homeostasis and H₂O₂ detoxification. Furthermore in vivo localization of H₂O₂ and O₂ ⁻; and elevated expression of key antioxidant enzyme encoding genes, SOD, POD and APX evident the possible role of SbMT-2 in ROS scavenging/detoxification mechanism. Transgenic lines showed accumulation of Cu⁺⁺ and Cd⁺⁺ in root while Zn⁺⁺ in stem under stress condition. Under control (unstressed) condition, Zn⁺⁺ was accumulated more in root but accumulation of Zn⁺⁺ in stem under stress condition suggested that SbMT-2 may involve in the selective translocation of Zn⁺⁺ from root to stem. This observation was further supported by the up-regulation of zinc transporter encoding genes NtZIP1 and NtHMA-A under metal ion stress condition. The study suggested that SbMT-2 modulates ROS scavenging and is a potential candidate to be used for phytoremediation and imparting stress tolerance.     

Expression of a Modified Cry1Ie Gene in E. coli and in Transgenic Tobacco Confers Resistance to Corn Borer

Gene transfer technology provides an alternativeapproach to breed insect-resistant crops. Insect-resistantgenes from plants or microbes could be introduced intoplants and the expressed insecticidal protein in plantcells could kill the target insects. Transgenic plantsexpressing a corresponding insecticidal crystal protein genefrom Bacillus thuringiensis (Bt) have been developed sincethe early 1980s [1,2]. Analysis of Bt gene sequencesrevealed that they contain numerous motifs seldom foundin p…     

The Choline Oxidase Gene codA Confers Salt Tolerance to Transgenic Eucalyptus globulus in a Semi-Confined Condition

The performance of tree species is influenced by environmental factors and growth stages. To evaluate the practical performance of transgenic tree species, it is insufficient to grow small, young trees under controlled conditions, such as in a growth chamber. Three transgenic Eucalyptus globulus lines, carrying the choline oxidase gene, were investigated for their salt tolerance and expression of the transgene at the young plantlet stage in a special netted-house. To clarify the characteristics at the young as well during the later stages, salt tolerance and the properties of the transgenic lines at large juvenile and adult stages were evaluated in the special netted-house. All transgenic lines showed high glycinebetaine content, particularly in young leaves. Trees of the transgenic line 107-1 showed low damage because of salinity stress based on the results from the chlorophyll analysis and malondialdehyde content, and they survived the high-salt-shock treatment at the large juvenile and adult stages. Only this line showed salt tolerance at all stages in the special netted-house. In this evaluation in the special netted-house, the tolerant line among young plantlets might perform better at all stages. Since evaluation in these special netted-house mimics field evaluation, line 107-1 is a potential tolerant line.     

Arabidopsis DELLA Protein Degradation Is Controlled by a Type-One Protein Phosphatase,TOPP4

Gibberellins (GAs) are a class of important phytohormones regulating a variety of physiological processes during normal plant growth and development. One of the major events during GA-mediated growth is the degradation of DELLA proteins, key negative regulators of GA signaling pathway. The stability of DELLA proteins is thought to be controlled by protein phosphorylation and dephosphorylation. Up to date, no phosphatase involved in this process has been identified. We have identified a dwarfed dominant-negative Arabidopsis mutant, named topp4-1. Reduced expression of TOPP4 using an artificial microRNA strategy also resulted in a dwarfed phenotype. Genetic and biochemical analyses indicated that TOPP4 regulates GA signal transduction mainly via promoting DELLA protein degradation. The severely dwarfed topp4-1 phenotypes were partially rescued by the DELLA deficient mutants rga-t2 and gai-t6, suggesting that the DELLA proteins RGA and GAI are required for the biological function of TOPP4. Both RGA and GAI were greatly accumulated in topp4-1 but significantly decreased in 35S-TOPP4 transgenic plants compared to wild-type plants. Further analyses demonstrated that TOPP4 is able to directly bind and dephosphorylate RGA and GAI, confirming that the TOPP4-controlled phosphorylation status of DELLAs is associated with their stability. These studies provide direct evidence for a crucial role of protein dephosphorylation mediated by TOPP4 in the GA signaling pathway.     

Co-transfer of LEA and bZip Genes from Tamarix Confers Additive Salt and Osmotic Stress Tolerance in Transgenic Tobacco

In this study, the additive effects of a late embryogenesis abundant (LEA) gene and a basic leucine zipper (bZIP) gene on salt and osmotic stress in Tamarix plants were analyzed. The constructs containing one or both of the LEA and bZIP genes were transformed into tobacco. Northern blot analysis showed the genes were overexpressed under the control of the CaMV 35S promoter in both dual and single gene-transgenic tobacco lines. The effects of salt and osmotic stress in transgenic tobacco plant were investigated. Following exposure to NaCl, mannitol, and PEG6000 stress, dual gene-transgenic lines showed higher seed generation and growth rates than single gene-transgenic lines and the wild-type. In response to NaCl stress, the dual gene-transgenic lines showed lower malondialdehyde and higher leaf chlorophyll content than single gene-transgenic lines and the wild-type. These results suggested that the co-expression of LEA and bZIP resulted in an additive enhancement of stress tolerance in dual gene-transgenic tobacco.     

Overexpression of Rice Sphingosine-1-Phoshpate Lyase Gene OsSPL1 in Transgenic Tobacco Reduces Salt and Oxidative Stress Tolerance

Sphingolipids, including sphingosine-1-phosphate (S1P), have been shown to function as signaling mediators to regulate diverse aspects of plant growth, development, and stress response. In this study, we performed functional analysis of a rice (Oryza sativa) S1P lyase gene OsSPL1 in transgenic tobacco plants and explored its possible involvement in abiotic stress response. Overexpression of OsSPL1 in transgenic tobacco resulted in enhanced sensitivity to exogenous abscisic acid (ABA), and decreased tolerance to salt and oxidative stress, when compared with the wild type. Furthermore, the expression levels of some selected stress-related genes in OsSPL1-overexpressing plants were reduced after application of salt or oxidative stress, indicating that the altered responsiveness of stress-related genes may be responsible for the reduced tolerance in OsSPL1-overexpressing tobacco plants under salt and oxidative stress. Our results suggest that rice OsSPL1 plays an important role in abiotic stress responses.     

Island Cotton Gbve1 Gene Encoding A Receptor-Like Protein Confers Resistance to Both Defoliating and Non-Defoliating Isolates of Verticillium dahliae

Verticillium wilt caused by soilborne fungus Verticillium dahliae could significantly reduce cotton yield. Here, we cloned a tomato Ve homologous gene, Gbve1, from an island cotton cultivar that is resistant to Verticillium wilt. We found that the Gbve1 gene was induced by V. dahliae and by phytohormones salicylic acid, jasmonic acid, and ethylene, but not by abscisic acid. The induction of Gbve1 in resistant cotton was quicker and stronger than in Verticillium-susceptible upland cotton following V. dahliae inoculation. Gbve1 promoter-driving GUS activity was found exclusively in the vascular bundles of roots and stems of transgenic Arabidopsis. Virus-induced silencing of endogenous genes in resistant cotton via targeting a fragment of the Gbve1 gene compromised cotton resistance to V. dahliae. Furthermore, we transformed the Gbve1 gene into Arabidopsis and upland cotton through Agrobacterium-mediated transformation. Overexpression of the Gbve1 gene endowed transgenic Arabidopsis and upland cotton with resistance to high aggressive defoliating and non-defoliating isolates of V. dahliae. And HR-mimic cell death was observed in the transgenic Arabidopsis. Our results demonstrate that the Gbve1 gene is responsible for resistance to V. dahliae in island cotton and can be used for breeding cotton varieties that are resistant to Verticillium wilt.     

Resistance to Potato Virus X Infection in Transgenic Tobacco Plants with Coat Protein Gene of Virus

A major commercial cultivar of tobacco was transformed via Agrobacterium mediated procedure. Tobacco leaves started to form shoots on shoot inducing medium containing kanamycin after infected by Agrobacterium containing the plasmid with PVX CP gene. Regenerated plants were obtained in two weeks on hormone-free MS medium containing kanamycin. The transgenic tobacco plants were identified with nopaline detection,enzyme-linked immunosorbent assay and western blot analysis, symptom appearance was significantly delayed and virus accumulation was either absent or reduced in PVX CP gene transformed plants. Progenies of transgenic tobacco plants also gained resistance to PVX infection to a certain degree. These experiments demonstrate that CP protection is effective against PVX.     

SERPINA2 Is a Novel Gene with a Divergent Function from SERPINA1

Serine protease inhibitors (SERPINs) are a superfamily of highly conserved proteins that play a key role in controlling the activity of proteases in diverse biological processes. The SERPIN cluster located at the 14q32.1 region includes the gene coding for SERPINA1, and a highly homologous sequence, SERPINA2, which was originally thought to be a pseudogene. We have previously shown that SERPINA2 is expressed in different tissues, namely leukocytes and testes, suggesting that it is a functional SERPIN. To investigate the function of SERPINA2, we used HeLa cells stably transduced with the different variants of SERPINA2 and SERPINA1 (M1, S and Z) and leukocytes as the in vivo model. We identified SERPINA2 as a 52 kDa intracellular glycoprotein, which is localized at the endoplasmic reticulum (ER), independently of the variant analyzed. SERPINA2 is not significantly regulated by proteasome, proposing that ER localization is not due to misfolding. Specific features of SERPINA2 include the absence of insoluble aggregates and the insignificant response to cell stress, suggesting that it is a non-polymerogenic protein with divergent activity of SERPINA1. Using phylogenetic analysis, we propose an origin of SERPINA2 in the crown of primates, and we unveiled the overall conservation of SERPINA2 and A1. Nonetheless, few SERPINA2 residues seem to have evolved faster, contributing to the emergence of a new advantageous function, possibly as a chymotrypsin-like SERPIN. Herein, we present evidences that SERPINA2 is an active gene, coding for an ER-resident protein, which may act as substrate or adjuvant of ER-chaperones.