M. Lacuesta

The oxidative stress caused by salinity in two barley cultivars is mitigated by elevated CO2.

Changes in antioxidant metabolism because of the effect of salinity stress (0, 80, 160 or 240 mM NaCl) on protective enzyme activities under ambient (350 micromol mol(-1)) and elevated (700 micromol mol(-1)) CO(2) concentrations were investigated in two barley cultivars (Hordeum vulgare L., cvs Alpha and Iranis). Electrolyte leakage, peroxidation, antioxidant enzyme activities [superoxide dismutase (SOD), EC 1.15.1.1; ascorbate peroxidase (APX), EC 1.11.1.11; catalase (CAT), EC 1.11.1.6; dehydroascorbate reductase (DHAR), EC 1.8.5.1; monodehydroascorbate reductase (MDHAR), EC 1.6.5.4; glutathione reductase (GR), EC 1.6.4.2] and their isoenzymatic profiles were determined. Under salinity and ambient CO(2), upregulation of antioxidant enzymes such as SOD, APX, CAT, DHAR and GR occurred. However, this upregulation was not enough to counteract all ROS formation as both ion leakage and lipid peroxidation came into play. The higher constitutive SOD and CAT activities together with a higher contribution of Cu,Zn-SOD 1 detected in Iranis might possibly contribute and make this cultivar more salt-tolerant than Alpha. Elevated CO(2) alone had no effect on the constitutive levels of antioxidant enzymes in Iranis, whereas in Alpha it induced an increase in SOD, CAT and MDHAR together with a decrease of DHAR and GR. Under combined conditions of elevated CO(2) and salinity the oxidative damage recorded was lower, above all in Alpha, together with a lower upregulation of the antioxidant system. So it can be concluded that elevated CO(2) mitigates the oxidative stress caused by salinity, involving lower ROS generation and a better maintenance of redox homeostasis as a consequence of higher assimilation rates and lower photorespiration, being the response dependent on the cultivar analysed.

Lipoic acid and redox status in barley plants subjected to salinity and elevated CO2

Future environmental conditions will include elevated concentrations of salt in the soil and an elevated concentration of CO(2) in the atmosphere. Because these environmental changes will likely affect reactive oxygen species (ROS) formation and cellular antioxidant metabolism in opposite ways, we analyzed changes in cellular H(2)O(2) and non-enzymatic antioxidant metabolite [lipoic acid (LA), ascorbate (ASA), glutathione (GSH)] content induced by salt stress (0, 80, 160 or 240 mM NaCl) under ambient (350 micromol mol(-1)) or elevated (700 micromol mol(-1)) CO(2) concentrations in two barley cultivars (Hordeum vulgare L.) that differ in sensitivity to salinity (cv. Alpha is more sensitive than cv. Iranis). Under non-salinized conditions, elevated CO(2) increased LA content, while ASA and GSH content decreased. Under salinized conditions and ambient CO(2), ASA increased, while GSH and LA decreased. At 240 mM NaCl, H(2)O(2) increased in Alpha and decreased in Iranis. When salt stress was imposed at elevated CO(2), less oxidative stress and lower increases in ASA were detected, while LA was constitutively higher. The decrease in oxidative stress could have been because of less ROS formation or to a higher constitutive LA level, which might have improved regulation of ASA and GSH reductions. Iranis had a greater capacity to synthesize ASA de novo and had higher constitutive LA content than did Alpha. Therefore, we conclude that elevated CO(2) protects barley cultivars against oxidative damage. However, the magnitude of the positive effect is cultivar specific.

Temporal study of the effect of phosphinothricin on the activity of glutamine synthetase, glutamate dehydrogenase and nitrate reductase in Medicago sativa L.

Summary In a preliminary work we showed that when plants were sprayed with several doses of glufosinate, GS activity of the leaf was reduced by 50% after 48 h of application of 250 µM herbicide. Enzyme inhibition was accompained by a dramatic accumulation of ammonia. In this paper we analyze the time-course effect of phosphinothricin on nitrogen metabolism. Our results show that GS activity is the first process affected; more than 50% activity reduction is observed after 2 h of a 1,000 µM treatment, whereby ammonia values 400% higher than control were reached. NRase did not change until 24 h of assay, whereas protein content was reduced after 48 h. GDH activity was enhanced, but only 24 h after starting the treatment. These results indicate that the main target of PPT action is on GS activity, and other processes are long-term modified. GDH could, perhaps, reassimilate some of the ammonia produced but its activity is not able to completely prevent, the injury caused by phosphinothricin.

Atmospheric CO2 concentration influences the contributions of osmolyte accumulation and cell wall elasticity to salt tolerance in barley cultivars.

Future environmental conditions will include elevated concentrations of salt in the soils and elevated concentrations of CO(2) in the atmosphere. Soil salinization inhibits crop growth due to osmotic and ionic stress. However, plants possess salt tolerance mechanisms, such as osmotic and elastic adjustment, to maintain water status. These mechanisms, which enhance the uptake and accumulation of ions and the synthesis of compatible solutes, require substantial energy expenditure. Under elevated CO(2), the carbon and energy supplies are usually higher, which could facilitate the energetically expensive salt tolerance mechanisms. To test this hypothesis, the factors involved in osmotic and elastic adjustments in two barley cultivars (Hordeum vulgare cv. Alpha and cv. Iranis) grown under several salt concentrations and at ambient or elevated [CO(2)] were evaluated. Under ambient [CO(2)] and salt stress, both cultivars (1) decreased the volumetric elasticity modulus (epsilon) of their cell walls, and (2) adjusted osmotically by accumulating ions (Na(+) and Cl(-)) from the soil, confirming barley as an includer species. The contributions of sugars and other unidentified osmolytes also increased, while the contribution of organic acids decreased. Under elevated [CO(2)] and salt stress, epsilon decreased less and osmotic adjustment (OA) was greater than at ambient [CO(2)]. In fact, the greater OA under elevated [CO(2)] was positively correlated with the contributions of sugars and other unidentified compounds. These results indicate that barley is likely to be successful in more salinized soils due to its capacity for OA under elevated [CO(2)].

Influence of water stress on photosynthetic characteristics in barley plants under ambient and elevated CO2 concentrations

We evaluated the combined effects of elevated CO2 and water availability on photosynthesis in barley. Soil and plant water content decreased with water stress, but less under elevated CO2 concentration (EC) compared with ambient CO2 concentration (AC). During water stress, stomatal conductance, carboxylation rate, RuBP regeneration, and the rate of triose phosphate utilisation (TPU) were decreased but less when plants grew under EC. Drought treatments caused only a slight effect on maximum photochemical efficiency (variable to maximum fluorescence ratio, Fv/Fm), whereas the actual quantum yield (ΦPS2), maximum electron transport rate (Jmax) and photochemical quenching (qP) were decreased and the non photochemical quenching (NPQ) was enhanced. Under water deficit, the allocation of electrons to CO2 assimilation was diminished by 49 % at AC and by 26 % at EC while the allocation to O2 reduction was increased by 15 % at AC and by 12 % at EC.

Effect of Phosphinothricin (Glufosinate) on Activities of Glutamine Synthetase and Glutamate Dehydrogenase in Medicago sativa L.

Summary The effect of the glutamate analogue phosphinothricin on enzymes of nitrogen assimilation was studied as well as the accumulation of ammonia and nitrate after treatment of lucerne plants with this herbicide. Glutamine synthetase (GS) was assayed with several phosphinothricin concentrations. When plants were sprayed with 250, 500 and 1,000 μM of the herbicide, GS activity of the leaf was reduced by 50% at 250 μM. Phosphinothricin caused a high ammonia accumulation in the tissues, reaching values of 70 μmol ammonia/g dry weight after treatment with 1,000 μM phosphinothricin. Glutamate dehydrogenase (GDH) activity increased ca. 40 % and nitrate reductase activity (NRase) was strongly decreased. Neither nitrate content nor proteolytic activity were altered.

Solute accumulation and elastic modulus changes in six radiata pine breeds exposed to drought

Drought is one of the main abiotic factors that determine forest species growth, survival and productivity. For this reason, knowledge of plant drought response and the identification of physiological traits involved in stress tolerance will be of interest to breeding programs. In this work, several Pinus radiata D. Don breeds from different geographical origins were evaluated along a water stress period (4 weeks) and subsequent rewatering (1 week), showing different responses among them. Leaf water potential (Ψ(leaf)) and osmotic potential decreases were accompanied by a variation in the total relative water content (RWC, %). The most tolerant breeds presented the lowest leaf water potential and RWC at turgor loss point, and showed the lowest elastic modulus (ε) values. A high ε value was a characteristic of a less-drought-tolerant plant and was related to membrane alterations (high electrolyte leakage percentages) that could favor cell water loss. Of the group of solutes that contributed to osmotic adjustment, soluble carbohydrates were the most abundant, although stressed plants also increased their content of free amino acids [mainly proline (Pro) and glutamic acid (Glu), and γ-aminobutyric acid (GABA)] and free polyamines. In addition, the most sensitive breeds had a higher GABA/Glu ratio. After rewatering, Pro and GABA were higher in rehydrated plants than in controls.

Immunolocalization of IAA and ABA in roots and needles of radiata pine (Pinus radiata) during drought and rewatering.

Anatomical, physiological and phytohormonal changes involved in drought tolerance were examined in different Pinus radiata D. Don breeds subjected to soil drying and rewatering. Breeds with the smallest stomatal chamber size had the lowest transpiration rate and the highest intrinsic water-use efficiency. Xylem cell size was positively correlated with leaf hydraulic conductance and needle indole-3-acetic acid (IAA) concentrations, whereas transpiration rate was negatively correlated with needle abscisic acid (ABA) levels. Since these two phytohormones seem important in regulating the P. radiata drought response, they were simultaneously immunolocalized in roots and needles of the most tolerant breed (P. radiata var. radiata × var. cedrosensis) during two sequential drought cycles and after rewatering. During drought, IAA was unequally distributed into the pointed area of the needle cross-section and mainly located in mesophyll and vascular tissue cells of needles, possibly inducing needle epinasty, whereas ABA was principally located in guard cells, presumably to elicit stomata closure. In the roots, at the end of the first drought cycle, while strong IAA accumulation was observed in the cortex, ABA levels decreased probably due to translocation to the leaves. Rewatering modified the distribution of both IAA and ABA in the needles, causing an accumulation principally in vascular tissue, with residual concentrations in mesophyll, likely favouring the acclimatization of the plants for further drought cycles. Contrarily, in the roots IAA and ABA were located in the exodermis, a natural barrier that regulates the phytohormone translocation to other plant tissues and hormone losses to the soil solution after rewatering. These results confirm that immunolocalization is an efficient tool to understand the translocation of IAA and ABA in plants subjected to different water stress situations, and clarify their role in regulating physiological responses such as stomata closure and epinasty in needles and root development.

Comparative study of the inhibition of photosynthesis caused by aminooxyacetic acid and phosphinothricin in Zea mays

Summary Maize plants fed with PPT (a glutamine synthetase inhibitor) accumulate ammonia. Approximately 50 % of the ammonia accumulated seems to come from the photorespiratory pathway, while the nonphotorespiratory ammonia is derived from nitrate reductase activity. The ammonia accumulated, however, is not sufficient to increase GDH activity. We also carried out a comparative study of the effect of PPT and AOA on photosynthesis. Both compounds inhibit the photorespiratory pathway, causing glycolate accumulation and diminishing photosynthesis.

Comparative effects of phosphinothricin on nitrate and ammonium assimilation and on anaplerotic CO2 fixation in N-deprived barley plants

Summary We have analyzed the comparative effect of nitrate and ammonium on the nitrogen-dependent induction of anaplerotic CO 2 fixation in detached leaves of N-limited barley plants. The supply of N, either in the form of NO 3 − or NH 4 + , provoked induction of phosphoenolpyruvate carboxylase activity (PEPCase), whereas the extractable pyruvate kinase, malate dehydrogenase and malic enzyme activities were not affected. The time course induction of PEPCase was dependent on the rate of ammonia assimilation and not on nitrate or ammonium uptake or accumulation. Treatment of N-deprived leaves with phosphinothricin (an inhibitor of glutamine synthetase activity) caused inhibition of ammonia assimilation, resulting in the reversion of ammonia-/nitrate-dependent enhancement of PEPCase. The results indicate that glutamine level controls phosphoenolpyruvate carboxylase activation in both nitrate and ammonium fed plants; consequently, if glutamine synthesis is inhibited, PEPCase activity is not enhanced. Determination of malate content showed that as PEPCase activity increased in response to increasing ammonia assimilation, there was a linear decline in the level of that metabolite. The highest rates of PEPCase enhancement in excised barley leaves when ammonia salt is the N source are in accordance with the highest rates of ammonia assimilation.