AtNPF2.5 modulates chloride (Cl -bar) efflux from roots of Arabidopsis thaliana

The accumulation of high concentrations of chloride (Cl(-)) in leaves can adversely affect plant growth. When comparing different varieties of the same Cl(-) sensitive plant species those that exclude relatively more Cl(-) from their shoots tend to perform better under saline conditions; however, the molecular mechanisms involved in maintaining low shoot Cl(-) remain largely undefined. Recently, it was shown that the NRT1/PTR Family 2.4 protein (NPF2.4) loads Cl(-) into the root xylem, which affects the accumulation of Cl(-) in Arabidopsis shoots. Here we characterize NPF2.5, which is the closest homolog to NPF2.4 sharing 83.2% identity at the amino acid level. NPF2.5 is predominantly expressed in root cortical cells and its transcription is induced by salt. Functional characterisation of NPF2.5 via its heterologous expression in yeast (Saccharomyces cerevisiae) and Xenopus laevis oocytes indicated that NPF2.5 is likely to encode a Cl(-) permeable transporter. Arabidopsis npf2.5 T-DNA knockout mutant plants exhibited a significantly lower Cl(-) efflux from roots, and a greater Cl(-) accumulation in shoots compared to salt-treated Col-0 wild-type plants. At the same time, [Formula: see text] content in the shoot remained unaffected. Accumulation of Cl(-) in the shoot increased following (1) amiRNA-induced knockdown of NPF2.5 transcript abundance in the root, and (2) constitutive over-expression of NPF2.5. We suggest that both these findings are consistent with a role for NPF2.5 in modulating Cl(-) transport. Based on these results, we propose that NPF2.5 functions as a pathway for Cl(-) efflux from the root, contributing to exclusion of Cl(-) from the shoot of Arabidopsis.Bo Li, Jiaen Qiu, Maheswari Jayakannan, Bo Xu, Yuan Li, Gwenda M. Mayo, Mark Tester, Matthew Gilliham and Stuart J. Ro

Salicylic acid improves salinity tolerance in Arabidopsis by restoring membrane potential and preventing salt-induced K(+) loss via a GORK channel

Despite numerous reports implicating salicylic acid (SA) in plant salinity responses, the specific ionic mechanisms of SA-mediated adaptation to salt stress remain elusive. To address this issue, a non-invasive microelectrode ion flux estimation technique was used to study kinetics of NaCl-induced net ion fluxes in Arabidopsis thaliana in response to various SA concentrations and incubation times. NaCl-induced K+ efflux and H+ influx from the mature root zone were both significantly decreased in roots pretreated with 10–500 μM SA, with strongest effect being observed in the 10–50 μM SA range. Considering temporal dynamics (0–8-h SA pretreatment), the 1-h pretreatment was most effective in enhancing K+ retention in the cytosol. The pharmacological, membrane potential, and shoot K+ and Na+ accumulation data were all consistent with the model in which the SA pretreatment enhanced activity of H+-ATPase, decreased NaCl-induced membrane depolarization, and minimized NaCl-induced K+ leakage from the cell within the first hour of salt stress. In long-term treatments, SA increased shoot K+ and decreased shoot Na+ accumulation. The short-term NaCl-induced K+ efflux was smallest in the gork1-1 mutant, followed by the rbohD mutant, and was highest in the wild type. Most significantly, the SA pretreatment decreased the NaCl-induced K+ efflux from rbohD and the wild type to the level of gork1-1, whereas no effect was observed in gork1-1. These data provide the first direct evidence that the SA pretreatment ameliorates salinity stress by counteracting NaCl-induced membrane depolarization and by decreasing K+ efflux via GORK channels.Maheswari Jayakannan, Jayakumar Bose, Olga Babourina, Zed Rengel, and Sergey Shabal

The NPR1-dependent salicylic acid signalling pathway is pivotal for enhanced salt and oxidative stress tolerance in Arabidopsis

The role of endogenous salicylic acid (SA) signalling cascades in plant responses to salt and oxidative stresses is unclear. Arabidopsis SA signalling mutants, namely npr1-5 (non-expresser of pathogenesis related gene1), which lacks NPR1-dependent SA signalling, and nudt7 (nudix hydrolase7), which has both constitutively expressed NPR1-dependent and NPR1-independent SA signalling pathways, were compared with the wild type (Col-0) during salt or oxidative stresses. Growth and viability staining showed that, compared with wild type, the npr1-5 mutant was sensitive to either salt or oxidative stress, whereas the nudt7 mutant was tolerant. Acute salt stress caused the strongest membrane potential depolarization, highest sodium and proton influx, and potassium loss from npr1-5 roots in comparison with the wild type and nudt7 mutant. Though salt stress-induced hydrogen peroxide production was lowest in the npr1-5 mutant, the reactive oxygen species (ROS) stress (induced by 1mM of hydroxyl-radical-generating copper-ascorbate mix, or either 1 or 10mM hydrogen peroxide) caused a higher potassium loss from the roots of the npr1-5 mutant than the wild type and nudt7 mutant. Long-term salt exposure resulted in the highest sodium and the lowest potassium concentration in the shoots of npr1-5 mutant in comparison with the wild type and nudt7 mutant. The above results demonstrate that NPR1-dependent SA signalling is pivotal to (i) controlling Na(+) entry into the root tissue and its subsequent long-distance transport into the shoot, and (ii) preventing a potassium loss through depolarization-activated outward-rectifying potassium and ROS-activated non-selective cation channels. In conclusion, NPR1-dependent SA signalling is central to the salt and oxidative stress tolerance in Arabidopsis.Maheswari Jayakannan, Jayakumar Bose, Olga Babourina, Sergey Shabala, Amandine Massart, Charlotte Poschenrieder and Zed Renge

Adaptive mechanisms of plants against salt stress and salt shock

Salinization process occurs when soil is contaminated with salt, which consequently influences plant growth and development leading to reduction in yield of many food crops. Responding to a higher salt concentration than the normal range can result in plant developing complex physiological traits and activation of stress-related genes and metabolic pathways. Many studies have been carried out by different research groups to understand adaptive mechanism in many plant species towards salinity stress. However, different methods of sodium chloride (NaCl) applications definitely give different responses and adaptive mechanisms towards the increase in salinity. Gradual increase in NaCl application causes the plant to have salt stress or osmotic stress, while single step and high concentration of NaCl may result in salt shock or osmotic shock. Osmotic shock can cause cell plasmolysis and leakage of osmolytes in plant. Also, the gene expression pattern is influenced by the type of methods used in increasing the salinity. Therefore, this chapter discusses the adaptive mechanism in plant responding to both types of salinity increment, which include the morphological changes of plant roots and aerial parts, involvement of signalling molecules in stress perception and regulatory networks and production of osmolyte and osmoprotective proteins

Segmented Polyethylene Oxides: A New Class of Polyethers Prepared via Melt Transetherification

Several segmented polyethylene oxides (SPEOs) were prepared by a melt-transetherification process using 1,4-bis(methoxymethyl)-2,3,5,6-tetramethylbenzene and polyethylene glycols (PEGs) of different molecular weights (di-, tri-, and tetraethylene glycols and PEGs of molecular weights 300, 600, 1000, 1500, and 3400) as the monomers. The effect of polymerization temperature (185 and 150 °C) on the molecular weight of SPEOs was studied, and it was shown that the molecular weight is larger at a higher polymerization temperature. The reversal of the polycondensation (transetherification) equilibrium by treatment of the polyethers with excess methanol transformed them completely into the starting monomers. The analysis of the degraded products by mass and NMR spectroscopies revealed that side reactions, such as the self-condensation of diols, are insignificant. The polymers containing shorter PEG spacers are amorphous, whereas the ones with longer PEG spacers are semicrystalline. The glass-transition temperature $(T_g)$ of the SPEOs decreased with increases in the spacer length and attained the value of PEO at PEG-600, whereas the melting transition $(T _m)$, crystallization temperature $(T_c)$, and their enthalpies of transition, $(\Delta H_m)$ and $(\Delta H_c)$, increased with increases in the spacer length. The introduction of "molecular kinks" into SPEOs by the use of another monomer, 1,3-bis(methoxymethyl)-2,4,5,6-tetramethylbenzene, surprisingly, showed little effect on their thermal properties. A "branched-PEO" analogue, containing pendant oligoethyleneoxy units, was also prepared, and its thermal properties were compared with its linear analogue. Preliminary ionic conductance measurements showed that some of these SPEOs could serve as potential candidates for solid polymer-electrolyte applications

Novel Melt Transurethane Process for Cycloaliphatic Polyurethanes and Their Self-Organization for Nano-Materials

In this introduction part, importance has been given to the elastomeric properties of polyurethanes. Emphasis has been laid to this property based on microphase separation and how this could be modified by modifying the segment lengths, as well as the structure of the segments. Implication was also made on the mechanical and thermal properties of these copolymers based on various analytical methods usually used for characterization of polymers. A brief overview of the challenges faced by the polyurethane chemistry was also done, pointing to the fact that though polyurethane industry is more than 75 years old, still a lot of questions remain unanswered, that too mostly in the synthesis of polyurethanes. A major challenge in this industry is the utilization of more environmental friendly “Green Chemistry Routes” for the synthesis of polyurethanes which are devoid of any isocyanates or harsh solvents.The research work in this thesis was focused to develop non-isocyanate green chemical process for polyurethanes and also self-organize the resultant novel polymers into nano-materials. The thesis was focused on the following three major aspects:(i) Design and development of novel melt transurethane process for polyurethanes under non-isocyanate and solvent free melt condition. (ii) Solvent induced self-organization of the novel cycloaliphatic polyurethanes prepared by the melt transurethane process into microporous templates and nano-sized polymeric hexagons and spheres. (iii) Novel polyurethane-oligophenylenevinylene random block copolymer nano-materials and their photoluminescence properties. The second chapter of the thesis gives an elaborate discussion on the “Novel Melt Transurethane Process ” for the synthesis of polyurethanes under non-isocyanate and solvent free melt condition. The polycondensation reaction was carried out between equimolar amounts of a di-urethane monomer and a diol in the presence of a catalyst under melt condition to produce polyurethanes followed by the removal of low boiling alcohol from equilibrium. The polymers synthesized through this green chemical route were found to be soluble (devoid of any cross links), thermally stable and free from any isocyanate entities. The polymerization reaction was confirmed by various analytical techniques with specific references to the extent of reaction which is the main watchful point for any successful polymerization reaction. The mechanistic aspects of the reaction were another point of consideration for the novel polymerization route which was successfully dealt with by performing various model reactions. Since this route was successful enough in synthesizing polyurethanes with novel structures, they were employed for the solvent induced self-organization which is an important area of research in the polymer world in the present scenario. Chapter three mesmerizes the reader with multitudes of morphologies depending upon the chemical backbone structure of the polyurethane as well as on the nature and amount of various solvents employed for the self-organization tactics. The rationale towards these morphologies-“Hydrogen Bonding ” have been systematically probed by various techniques. These polyurethanes were then tagged with luminescent 0ligo(phenylene vinylene) units and the effects of these OPV blocks on the morphology of the polyurethanes were analyzed in chapter four. These blocks have resulted in the formation of novel “Blue Luminescent Balls” which could find various applications in optoelectronic devices as well as delivery vehicles

Synthesis and Thermal Analysis of Branched and "Kinked" Poly (ethylene terephthalate)

Poly(ethylene terephthalate) (PET) was synthesized by self-condensation of bis-(2-hydroxyethyl) terephthalate (BHET). Copolymerization of BHET with ethyl, bis-3,5-(2- hydroxyethoxy) benzoate (EBHEB) and ethyl, 3-(2-hydroxyethoxy) benzoate (E3HEB) yielded copolymers that ontain varying amounts of branching and kinks, respectively. Copolymers of BHET with ethyl, 4-(2-hydroxyethoxy) benzoate (E4HEB), in which only the backbone symmetry is broken but without disruption of the linearity, were also prepared for comparison. The composition of the copolymers were established from their $^1H-NMR$ spectra. The intrinsic viscosity of all the copolymers indicated that they were of reasonably high molecular weights. The thermal analysis of the copolymers using DSC showed that both the melting temperatures $(T_m)$ and the percent crystallinity (as seen from the enthalpies of melting $(\Delta H_m)$ decreased with increasing comonomer (defect concentration) content, although their glass transition temperatures $(T_g)$ were less affected. This effect was found to be most pronounced in the case of branching, while the effects of kinks and linear disruptions, on both $T_m$ and $\Delta H_m$, were found to be similar

Synthesis and thermal analysis of branched and "kinked" poly(ethylene terephthalate)

Poly(ethylene terephthalate) (PET) was synthesized by self-condensation of bis-(2-hydroxyethyl) terephthalate (BHET). Copolymerization of BHET with ethyl, bis-3,5-(2-hydroxyethoxy) benzoate (EBHEB) and ethyl, 3-(2-hydroxyethoxy) benzoate (E3HEB) yielded copolymers that contain varying amounts of branching and kinks, respectively. Copolymers of BHET with ethyl, 4-(2-hydroxyethoxy) benzoate (E4HEB), in which only the backbone symmetry is broken but without disruption of the linearity, were also prepared for comparison. The composition of the copolymers were established from their <SUP>1</SUP>H-NMR spectra. The intrinsic viscosity of all the copolymers indicated that they were of reasonably high molecular weights. The thermal analysis of the copolymers using DSC showed that both the melting temperatures (T<SUB>m</SUB>) and the percent crystallinity (as seen from the enthalpies of melting) (ΔH<SUB>m</SUB>) decreased with increasing comonomer (defect concentration) content, although their glass transition temperatures (T<SUB>g</SUB>) were less affected. This effect was found to be most pronounced in the case of branching, while the effects of kinks and linear disruptions, on both T<SUB>m</SUB> and ΔH<SUB>m</SUB>, were found to be similar

Effect of Branching on the Crystallization Kinetics of Poly(ethylene terephthalate)

The effect of branching on the crystallization behavior of poly(ethylene terephthalate) has been examined by nonisothermal crystallization studies, using DSC. It was found that branching causes a significant change in the crystallization behavior, in that the Avrami exponent n lies between 1 and 2, suggesting a rodlike growth process compared to a spherulitic one observed in the case of PET. In addition, the effect of molecular kinks and linear disruptions were also examined; in both these cases, however, the same spherulitic growth, as seen in the case of PET, is observed. Further, the presence of branching, kinks and linear disruptions, in small concentrations, appears to enhance the crystallization process, possibly, by acting to facilitate nucleation. At higher concentrations of such defects, however, the crystallization process is slowed down and the overall crystallinity of the PET copolymers is reduced

Effect of branching on the crystallization kinetics of poly(ethylene terephthalate)

The effect of branching on the crystallization behavior of poly(ethylene terephthalate) has been examined by nonisothermal crystallization studies, using DSC. It was found that branching causes a significant change in the crystallization behavior, in that the Avrami exponent n lies between 1 and 2, suggesting a rodlike growth process compared to a spherulitic one observed in the case of PET. In addition, the effect of molecular kinks and linear disruptions were also examined; in both these cases, however, the same spherulitic growth, as seen in the case of PET, is observed. Further, the presence of branching, kinks and linear disruptions, in small concentrations, appears to enhance the crystallization process, possibly, by acting to facilitate nucleation. At higher concentrations of such defects, however, the crystallization process is slowed down and the overall crystallinity of the PET copolymers is reduced