Malate Accumulates in the Vacuole of Chara australis During Uptake of Ammonium From Chloride-free Solution

Internodal cells of Chara australis can accumulate ammoniumto high concentrations (10 to 70 mol m^–3) in their vacuoles.When Cl^– is included in the bathing solution, changesin the cellular concentrations of ammonium, K⁺, Cl^– andNa⁺ have been shown to meet the requirements for electroneutralityand to account for the changes in vacuolar osmotic pressureassociated with ammonium uptake. If accumulation occurs in theabsence of external Cl^–, however; changes in the inorganicions do not meet these criteria. Malate is found in the vacuolesof cells accumulating amine in the absence of external Cl^–and its presence (at 0·5 to 8·5 mol m^–3)allows us to account for electroneutrality and for changes inthe osmotic potential. Key words: Malate, Chara, electroneutrality, ammonium     

Habitat scale, female cannibalism, and male reproductive success in three-spined sticklebacks (Gasterosteus aculeatus)

We studied the effects of habitat scale (pool size) on the riskof egg cannibalism and the breeding behavior of three-spinedsticklebacks (Gasterosteus aculeatus). Laboratory experimentsshowed that in pools with similar population density, a largerfraction of males nested in pools of 4.8-m² surface area thanin 15-m² pools. Breeding males in the 4.8-m² pools also hadbigger territories, obtained more eggs, and were more aggressivetoward intruders than those in the 15-m² pools. The numbersof raids attempted on nests were similar in the two pool sizes.However, raiding-shoal sizes were larger, and hence individualsin these larger shoals were more succesful at overwhelming maleterritorial defenses in the 15-m² pools than in the 4.8-m² pools.At the field site, a salt marsh containing hundreds of tidepools of different sizes, sticklebacks settled at similar densitiesin small (<30 m² surface area) and large (>355 m²) pools.This resulted in significantly more fish being present in largepools than in small pools, a concomitant increase in the amountof nest raiding, and a decrease in individual male reproductivesuccess in large compared to small pools. A photogrammetry analysisof the salt-marsh pools suggests that the majority of fish breedingat the site will be subjected to a high risk of cannibalismbecause most fish nest in large pools. We conclude that spatialscale has a major influence on this species' breeding behaviorand reproductive success.     

Glutamine's protection against cellular injury is dependent on heat shock factor-1

Glutamine (GLN) has been shown to protect cells, tissues, and whole organisms from stress and injury. Enhanced expression of heat shock protein (HSP) has been hypothesized to be responsible for this protection. To date, there are no clear mechanistic data confirming this relationship. This study tested the hypothesis that GLN-mediated activation of the HSP pathway via heat shock factor-1 (HSF-1) is responsible for cellular protection. Wild-type HSF-1 (HSF-1^+/+) and knockout (HSF-1^–/–) mouse fibroblasts were used in all experiments. Cells were treated with GLN concentrations ranging from 0 to 16 mM and exposed to heat stress injury in a concurrent treatment model. Cell viability was assayed with phenazine methosulfate plus tetrazolium salt, HSP-70, HSP-25, and nuclear HSF-1 expression via Western blot analysis, and HSF-1/heat shock element (HSE) binding via EMSA. GLN significantly attenuated heat-stress induced cell death in HSF-1^+/+ cells in a dose-dependent manner; however, the survival benefit of GLN was lost in HSF-1^–/– cells. GLN led to a dose-dependent increase in HSP-70 and HSP-25 expression after heat stress. No inducible HSP expression was observed in HSF-1^–/– cells. GLN increased unphosphorylated HSF-1 in the nucleus before heat stress. This was accompanied by a GLN-mediated increase in HSF-1/HSE binding and nuclear content of phosphorylated HSF-1 after heat stress. This is the first demonstration that GLN-mediated cellular protection after heat-stress injury is related to HSF-1 expression and cellular capacity to activate an HSP response. Furthermore, the mechanism of GLN-mediated protection against injury appears to involve an increase in nuclear HSF-1 content before stress and increased HSF-1 promoter binding and phosphorylation. knockout cells; amino acid; heat stress mechanism     

Heat shock-induced attenuation of hydroxyl radical generation and mitochondrial aconitase activity in cardiac H9c2 cells

A mild heat shock (hyperthermia) protects cells from apoptotic and necrotic deaths by inducing overexpression of various heat shock proteins (Hsps). These proteins, in combination with the activation of the nitric oxide synthase (NOS) enzyme, play important roles in the protection of the myocardium against a variety of diseases. In the present work we report that the generation of potent reactive oxygen species (ROS), namely ·OH in cardiac H9c2 cells, is attenuated by heat shock treatment (2 h at 42°C). Western blot analyses showed that heat shock treatment induced overexpression of Hsp70, Hsp60, and Hsp25. The observed ·OH was found to be derived from the superoxide (O₂^–·) generated by the mitochondria. Whereas the manganese superoxide dismutase (MnSOD) activity was increased in the heat-shocked cells, the mitochondrial aconitase activity was reduced. The mechanism of O₂^–· conversion into ·OH in mitochondria is proposed as follows. The O₂^–· leaked from the electron transport chain, oxidatively damages the mitochondrial aconitase, releasing a free Fe²⁺. The aconitase-released Fe²⁺ combines with H₂O₂ to generate ·OH via a Fenton reaction and the oxidized Fe³⁺ recombines with the inactivated enzyme after being reduced to Fe²⁺ by other cellular reductants, turning it over to be active. However, in heat-shocked cells, because of higher MnSOD activity, the excess H₂O₂ causes irreversible damage to the mitochondrial aconitase enzyme, thus inhibiting its activity. In conclusion, we propose that attenuation of ·OH generation after heat shock treatment might play an important role in reducing the myocardial ischemic injury, observed in heat shock-treated animals. proteins; free radicals; spin trapping; reactive oxygen species     

Heat Stress Activates Fission Yeast Spc1/StyI MAPK by a MEKK-Independent Mechanism

Fission yeast Spc1/StyI MAPK is activated by many environmental insults including high osmolarity, oxidative stress, and heat shock. Spc1/StyI is activated by Wis1, a MAPK kinase (MEK), which is itself activated by Wik1/Wak1/Wis4, a MEK kinase (MEKK). Spc1/StyI is inactivated by the tyrosine phosphatases Pyp1 and Pyp2. Inhibition of Pyp1 was recently reported to play a crucial role in the oxidative stress and heat shock responses. These conclusions were based on three findings: 1) osmotic, oxidative, and heat stresses activate Spc1/StyI in wis4 cells; 2) oxidative stress and heat shock activate Spc1/StyI in cells that express Wis1AA, in which MEKK consensus phosphorylation sites were replaced with alanine; and 3) Spc1/StyI is maximally activated in Δpyp1 cells. Contrary to these findings, we report: 1) Spc1/StyI activation by osmotic stress is greatly reduced in wis4 cells; 2) wis1-AA and Δwis1 cells have identical phenotypes; and 3) all forms of stress activate Spc1/StyI in Δpyp1 cells. We also report that heat shock, but not osmotic or oxidative stress, activate Spc1 in wis1-DD cells, which express Wis1 protein that has the MEKK consensus phosphorylation sites replaced with aspartic acid. Thus osmotic and oxidative stress activate Spc1/StyI by a MEKK-dependent process, whereas heat shock activates Spc1/StyI by a novel mechanism that does not require MEKK activation or Pyp1 inhibition.     

Effect of osmotic shock on auxin-induced cell extension, cell wall changes and acidification in Avena coleoptile segments

The effect of plasma membrane alteration caused by osmotic shockof different strengths on the auxin-induced responses of Avenacoleoptile cells was observed. Osmotic shock brought about by0.5–0.7 M mannitol solution for 10 or 30 min, followedby phosphate-buffer (1 mM, pH 6.0) treatment for 10 min at 4?Ccaused no significant inhibition of auxin-induced cell extension.The osmotic shock did not affect auxin-induced cell wall looseningrepresented by stress-relaxation time and a decrease in thenoncellulosic glucose level of the cell wall. The shock causedonly a temporary inhibition of transmembrane potential and noinhibition of oxygen consumption. However, it inhibited auxin-stimulatedH⁺ secretion which was reversed by 0.1 mM CaCl₂. We concludedthat the Osmotic shock may partly modify the plasma membranerelated to the hydrogen ion pump which interacts with auxin,but this modification which is reflected little by the transmembranepotential and cellular metabolism, is not closely related toauxin-induced cell wall loosening and thus cell extension inAvena coleoptiles. ³ Present address: Department of Botany, Faculty of Science,University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113, Japan (Received February 17, 1978; )     

AtsHsp17.6-CⅠ和AtsHsp17.6-CⅡ基因对非生物胁迫的应答研究

小分子热激蛋白是植物受到热胁迫后的主要表达产物之一,与植物细胞耐热有密切关系。该研究发现,拟南芥小分子热激蛋白基因AtsHsp17.6-CⅠ和AtsHsp17.6-CⅡ 除热激之外,重金属离子Ni⁺、Pb²⁺、Cu²⁺、Zn²⁺和Al³⁺均能诱导这2个热激蛋白基因的表达;氧化胁迫和渗透胁迫同样也能诱导它们表达。该研究将由CaMV35S启动子驱动的这2个小分子热激蛋白基因导入拟南芥,RT-PCR分析表明,2个小分子热激蛋白基因在转基因植物中呈现组成型表达。实验结果表明,组成型表达小分子热激蛋白基因AtsHsp17.6-CⅠ的转基因植物表现出对6 μmol·L^-1 Cd²⁺胁迫、0.4% NaCl胁迫的耐受性。研究表明,这2个小分子热激蛋白基因可能参与着多种抗逆途径,推测其能够减轻或抵抗逆境胁迫引起的伤害并对其进行修复。     

Electrophysiological characterization of the node in Chara corallina: functional differentiation for wounding response

Electrical characteristics of the node were analyzed in comparisonwith those of the flank of the internodal cell in Chara corallina.The dependence of the membrane potential of the node on pH andK⁺ concentration was almost the same as that of the flank. Inthe flank, the increase in the Ca²⁺ concentration stopped thedepolarization in the presence of 100 mM KCl. In the node, however,Ca²⁺ could not stop the depolarization induced by 100 mM KCl.It has been reported that the node has a function to tranducethe signal of osmotic shock into a transient depolarization.In combination with osmotic shock, 10 mM K⁺ could induce a long-lastingdepolarization of the node. These electrical characteristicsof the node were suggested to be responsible for the electricalresponse to wounding in Characeae.     

The Effect of Methionine Sulphoximine on the Absorption of Ammonium by Maize and Barley Roots over Short Periods

Pre-treating intact, ammonium-grown plants of maize and barleywith the glutamine synthetase inhibitor, L-methionine sulphoximine(MSO), increased subsequent accumulation of ¹³N or ¹⁵N fromlabelled ammonium very markedly—up to 8-fold in extremecases. The phenonemon could not be satisfactorily explainedin terms of an effect of MSO on the cation-exchange propertiesof the root free space, because neither potassium (⁸⁶Rb⁺) norsodium absorption was enhanced by MSO pretreatment, and rapidabsorption of ammonium continued for too long in relation tothe likely free-space content of a monovalent cation. Instead,the response is seen as a consequence of the enlargement ofthe cytoplasmic and vacuolar ammonium pools that occurs whenammonium assimilation is inhibited by MSO. The results, whichincluded comparisons between short-term and longer-term ratesof ammonium absorption, provide tentative evidence of rapidfluxes of ammonium through the plasma membrane. Key words: Cytoplasm, vacuole, flux, transport     

Phosphate Absorption, Fluxes, and Symplasmic Transport in Osmotically-Shocked Zea mays Roots

Osmotic shock with sequential 30 min treatments in ice-coldsaline solutions and distilled water inhibited both the subsequentuptake of orthophosphate (Pi) and its transport into the xylemof excised corn (Zea mays L.) roots. Measurements of Pi fluxeswith ³²P indicated that the decrease in net Pi uptake over a24 h period caused by osmotic shock was due primarily to delayedrecovery of Pi influx rather than to increasing efflux. Despitecomplete recovery of Pi absorption within 2–6 h aftershocking with 150–200 mM NaCl, transport to the xylemduring the subsequent 24 h only partially recovered. Leucineuptake and incorporation into protein was also markedly inhibitedby osmotic shock but both almost completely resumed controlrates within 24 h after shocking with up to 150 mM NaCl. Tetracyclineinhibited recovery of Pi uptake after NaCl treatment whereaspuromycin did not. These results with corn roots are consistentwith the hypothesis that recovery of Pi uptake activity aftermoderate osmotic shock requires de novo synthesis of membraneproteins. Incomplete recovery of Pi transport to the xylem suggeststhat osmotic shock may damage plasmodesmata. Key words: Corn, Ion uptake, Leucine uptake, NaCl, Puromycin, Tetracycline     

Osmotic and ionic regulation in Nitella

When the osmotic value of an internodal cell of Nitella flexiliswas modified by the method of transcellular osmosis, the normalosmotic value was chiefly restored by the release or absorptionof K⁺. The release or uptake of Na⁺ was observed only when themodification of osmotic value was significant. Both the uptakeand release of K⁺ were linearly dependent on the degree of modificationof the osmotic value. The effectiveness of alkali metal cationsin restoring the osmotic value in cells of lower osmotic valueswas in the order K⁺>Rb⁺>Na⁺, Cs⁺>Li⁺. The absorptionof K⁺ by cells of lower osmotic values depended strongly ontemperature, while the release of K⁺ from cells of higher osmoticvalues did not. To clarify whether the Nitella cell regulates the osmotic valueor regulates the concentration of K⁺ in the vacuole, the cellsap was exchanged for artificial cell saps whose osmotic valuesand ionic concentrations were varied independent of each other.It was shown that in Nitella two regulating mechanisms are operating,one which regulates the osmotic value of the cell sap irrespectiveof the level of vacuolar K⁺ (0.1–140 mM) and another whichregulates the vacuolar K⁺-level when it is abnormaly high (>160mM). Both mechanisms are assumed to operate in order to keepthe concentration of K⁺ in the cytoplasm at a constant level.The presence of Na⁺ (0–100 mM) and Ca²⁺ (5–40 mM)did not affect the movement of K⁺ during osmoregulation. ¹Present address: Sanki Engineering Limited, Nagaokakyo, Kyoto,Japan. (Received December 19, 1973; )     

Decreased cation channel activity and blunted channel-dependent eryptosis in neonatal erythrocytes

Eryptosis or apoptosis-like death of erythrocytes is characterized by phosphatidylserine exposure and erythrocyte shrinkage, both typical features of nucleated apoptotic cells. Eryptosis is triggered by activation of nonselective Ca²⁺-permeable cation channels with subsequent entry of Ca²⁺ and stimulation of Ca²⁺-sensitive scrambling of the cell membrane. The channels are activated and thus eryptosis is triggered by Cl^– removal, osmotic shock, oxidative stress, or glucose deprivation. The present study has been performed to compare cation channel activity and susceptibility to eryptosis in neonatal and adult erythrocytes. Channel activity was determined by patch-clamp analysis, cytosolic Ca²⁺ activity by fluo-3 fluorescence, phosphatidylserine exposure by FITC-labeled annexin V binding, and cell shrinkage by decrease in forward scatter in fluorescence-activated cell sorting analysis. Prostaglandin E₂ (PGE₂) formation, cation channel activity, Ca²⁺ entry, annexin V binding, and decreased forward scatter were triggered by removal of Cl^– in both adult and neonatal erythrocytes. The effects were, however, significantly blunted in neonatal erythrocytes. Osmotic shock, PGE_2, and platelet-activating factor similarly increased annexin V binding and decreased forward scatter, effects again significantly reduced in neonatal erythrocytes. On the other hand, spontaneous and oxidative (addition of tert-butylperoxide) stress-induced eryptosis was significantly larger in neonatal erythrocytes. In conclusion, cation channel activity, Ca²⁺ leakage, and thus channel-dependent triggering of eryptosis are blunted, whereas spontaneous and oxidative stress-induced eryptosis is more pronounced in neonatal erythrocytes. annexin V; osmotic cell shrinkage; calcium; apoptosis     

Activation of ERK by Ca2+ store depletion in rat liver epithelial cells

In rat liver epithelial (WB) cells,Ca²⁺ pool depletion induced by twoindependent methods resulted in activation of extracellular signal-regulated protein kinase (ERK). In the first method,Ca²⁺ pool depletion bythapsigargin increased the activity of ERK, even when rise in cytosolicCa²⁺ was blocked with theCa²⁺ chelator BAPTA-AM. For thesecond method, addition of extracellular EGTA at a concentration shownto deplete intracellular Ca²⁺pools also increased ERK activity. In each instance, ERK activation, asmeasured by an immunocomplex kinase assay, was greatly reduced by thetyrosine kinase inhibitor genistein, suggesting thatCa²⁺ store depletion increased ERKactivity through a tyrosine kinase pathway. The intracellularCa²⁺-releasing agent thapsigarginincreased Fyn activity, which was unaffected by BAPTA-AM pretreatment,suggesting that Fyn activity was unaffected by increased cytosolic freeCa²⁺. Furthermore, depletion ofintracellular Ca²⁺ with EGTAcaused inactivation of protein phosphatase 2A and protein tyrosinephosphatases. ANG II-induced activations of Fyn, Raf-1, and ERK wereaugmented in cells pretreated with BAPTA-AM, but ANG II-inducedexpression of the dual-specificity phosphatase mitogen-activatedprotein kinase phosphatase-1 was blocked by BAPTA-AM pretreatment.Together these results indicate that ERK activity is regulated by thebalance of phosphorylation vs. dephosphorylation reactions in intactcells and that the amount of Ca²⁺stored in intracellular pools plays an important role in this regulation.

     

Differential Stimulation of PEP-carboxylation in Guard Cells and Mesophyll Cells by Ammonium or Fusicoccin

Dark CO₂-fixation in guard cells of Vicia faba was much moresensitive to ammonium than in mesophyll cells. Addition of ammonium(5.0 mol m^–3; pH₀ 7.6) caused up to a 7-fold increasein dark CO₂-fixation rates in guard cell protoplasts (GCP),whereas in leaf slices, mesophyll cells, and mesophyll protoplaststhe increase was only about 1.4-fold. In both cell or tissuetypes, total CO₂-fixation rates were higher in the light (2–12-foldhigher in GCP and 28-fold in mesophyll); these rates were onlyslightly changed by ammonium treatment. However, separationof ¹⁴C-labelled products after fixation of CO₂ in the lightby GCP revealed a large ammonium-induced shift in carbon flowfrom starch and sugars to typical products of C₄-metabolism(mainly malate and aspartate). In contrast, in mesophyll cellsamino acid and malate labelling was only moderately increasedby ammonium at the expense of sucrose. The data suggest thatin vivo ammonium might facilitate stomatal opening and/or delaystomatal closing through an increased production of organicacids. Key words: PEP-carboxylation, guard cell protoplasts, ammonium, fusicoccin     

OSMOTIC EFFECT ON THE PHOTOSHRINKAGE OF CHLOROPLASTS

Shrinkage of spinach chloroplasts by illumination and that byexposure to a high tonicity raised by addition of sucrose wereinvestigated by means of the rapid measurement of chloroplastvolume with a COULTER counter. The osmotic shock in the darkinduced two steps of volume change; almost instantaneous shrinkagefor the osmotic shock and follow-up gradual swelling. The finalvolume attained after equilibration was smaller than the originalvolume below 0.6 M, and greater above this concentration. Whenchloroplasts under osmotic shock were illuminated, the photoshrinkagecompeted with the swelling induced by the osmotic shock, andthese reverse effects were balanced at a certain volume. Photoshrinkageactivity measured after equilibration decreased with increasingconcentrations, and the activity curve plotted against sucroseconcentration showed a stationary level of 50% of the originalactivity between 0.2 and 0.6 M, indicating the resistance ofa structure in chloroplasts to the denaturation by the osmoticeffect of sucrose. The osmotic effect in the dark as well asin the light was completely reversible below 0.2 M and was partiallyreversible or irreversible above this concentration. Glucoseshowed qualitatively the same osmotic effect as sucrose. ¹ Present address: Laboratory of Chemistry of Natural Products,Tokyo Institute of Technology, Meguroku, Tokyo.     

Solute Accumulation In Plant Cells IV. Effects of Ammonium lons on Growth and Solute Content

Procedures previously described were used to study growth andsolute content of aseptically cultured carrot explants as affectedby supplementary salts in the medium. The salts chosen (KC1,KNO₃, NH₄,Cl, and NH⁴,NO₃) contrasted, with appropriate controls,the effects due to nitrate and ammonium. Growth was measuredin terms of fresh weight, the number and average size of cells:solute concentrations were recorded for total solutes, sugars,soluble nitrogen compounds, and the electrolytes K⁺, Na⁺, C1^–,NO^–₃, and organic acids. The time-response curves of thecultures were traced at a fixed concentration of the added saltsand the effects due to the concentration of the supplementarysalts were tested after a fixed time period, For the same nitrogensource the concentrations of metabolites and solutes in cellswere very similar despite some clonal differences in their growth.When cells in a nitrate medium were small and dividing, thecultures had a low osmotic value, contained K⁺ as the principalcation balanced by organic acid, had relatively low sugar content,and their enriched total nitrogen content emphasized proteinrather than soluble nitrogen compounds. Later, as the cellsbecame older and larger, salts (K⁺, organic anions, Cl^–)contributed substantially to their increased osmotic value butthey accumulated sugar as their main, osmotically active solute,and the ratio of soluble to protein nitrogen declined as proteinsynthesis progressed. The extra nitrogen supplied by the additionalpotassium nitrate contributed more to protein and caused potassium,organic acids, and sugars to accumulate to higher levela. Supplementaryammonium salts required that more sugar be metabolized to organicnitrogen compounds (e.g. glutamine), contributed more to solublethan to protein nitrogen, and sharply reduced. both the osmoticvalue of the cells and the potassium linked to organic anions.The selectivity of the growing cells for K⁺ over Na⁺ and theirdiscrimination. between alkali cations (Ka⁺+Na⁺) and halides(C1^–) were relaxed in the presence of ammonia. Attentionis drawn to the implications of these results for the accumulationof solutes, organic and inorganic, by dividing and enlargingcells.     

The Mechanism of Photosynthetic Product Secretion by Carrot-Tissue 2

Green carrot-callus cultures exposed for 3 h in liquid mediato light and ¹⁴CO₂ secreted a constant 6–8 per cent oftheir total ¹⁴C-photosynthate into the medium over a wide rangeof ¹⁴CO₂-fixation rates, obtained by varying ¹⁴CO₂ levels. Themajor secreted products (sucrose and glutamine) or analoguesof these compounds did not affect secretion when included inthe bathing medium. Dual-labelling studies with [6-T] glucoseand ¹⁴CO₂ demonstrated that secretion occurred from a metabolicrather than a storage compartment. The effects of long preincubationperiods in liquid media and of osmotic shock showed that secretiontook place from specific sites on the boundary of the metaboliccompartment.     

Ionic Currents across the Plasmalemma of Chara inflata Cells: I. OSMOTIC EFFECTS OF SORBITOL ON K+, CI- AND LEAK CURRENTS

This paper describes experiments designed to investigate theeffects of increases in external osmotic pressure on the electrophysiologicalbehaviour of the plasmalemma in cells of the brackish-wateralga, Chara inflata. The electrical conductance of the plasmalemmaof these cells of, due to the diffusion of ions, consists mainlyof K⁺, Cl^– and leak components. The addition of sorbitolat concentrations in the range 40–280 mol m^–3 tothe external solution bathing the cells, progressively and reversiblydepolarized the cell membrane and increased the total membraneconductance, chiefly due to increases in each of the separateionic conductances. At concentrations higher than about 280mol m^–3 when the cells became plasmolysed, the effectsof sorbitol on the electrical properties of the cell ceasedto be fully reversible. When the membrane electrical potentialdifference is stepped in a negative direction with a voltage-clamp,the resulting inward current has voltage-dependent componentsconsisting of an inactivating K⁺ current, an activating Cl^–current and a constant leak current. The addition of sorbitolto the external solution modified these currents by increasingtheir magnitude, by increasing the half-time of the inactivationof the K⁺ current, and by decreasing the half-time of activationof the Cl^– current. Key words: Chara inflata, osmotic effects, K⁺ and Cl^– currents     

Evidence of ammonium assimilation via the glutamine synthetase-glutamate synthase system in rice seedling roots

When rice seedling roots were fed ¹⁵N-ammonium for 1 hr, theamide nitrogen of glutamine showed the highest ¹⁵N abundance.Moreover, glutamine amino, glutamic acid, aspartic acid andalanine showed higher ¹⁵N abundance than ammonium did. In roots whose GS activity was inhibited with MS, both the amountof ammonium and its ¹⁵N abundance were increased. In contrast,both the amount of all examined amino acids containing glutamicacid and their ¹⁵N abundance decreased in roots whose GS activitywas inhibited. From these results, it could be concluded thatthe first step of ammonium assimilation in rice seedling rootswas mainly glutamine synthesis by GS and the second was glutamicacid formation by the GOGAT system. The results of an experiment using ¹⁵N glutamine also supportedthis conclusion. (Received February 23, 1977; )     

Osmotic stress at membrane level and photosystem II activity in two C4 plants after growth in elevated CO2 and temperature

In the last two decades, several studies have evaluated plant physiology, growth and survival under forecasted climate changes and the effects of these environmental factors in plants are started to be understood. However, there are few studies evaluating such effects at the tissue or cellular level, especially for plants with photosynthetic C4 metabolism that are believed to respond less to elevated CO₂ concentration. For this reason, we tested maize and pearl millet plants to consider cellular physiological responses to induce osmotic stress and acute heat shock. Plants were grown under elevated CO₂ concentration and temperature, simulating global climate changes and then were subjected to osmotic stress and acute heat shock in vitro. The results indicated that the growth under elevated CO₂ and temperature improved cellular tolerance to osmotic stress and acute heat shock for both species, but maize seemed to benefit more from increased CO₂ concentration whereas pearl millet seemed to benefit more from increased temperature. Taken together, the results indicated that the current and expected global climate changes, besides operating differentially in these two species, can similarly affect other C4 plant species in different ecosystems whether undisturbed or managed.