Intracellular pH Regulation in the Giant-Celled Marine Alga Chaetomorpha darwinii

The vacuolar pH (pH_v) and the cytoplasmic pH (pH_c) of the marinegiant-celled green alga Chaetomorpha darwinii were measuredby pH microelectrode techniques on extracted vacuolar sap, andby the [^I4C]DMO distribution method respectively. Equilibrationof DMO occurred with a half-time of about 2 h, with an apparentP_DMO of 3.6 x 10^–5 cm s^–1, but the vacuolar concentrationof free, undissociated DMO was always less than the externalconcentration. The explanation offered for freshwater giant-celledalgae of net DMO^– leakage across the plasmalemma cannotapply to Chaetomorpha darwinii, since electrically-driven DMO^–exit from the cytoplasm should be similar across the plasmalemmaand the tonoplast in these cells with large, vacuole-positivepotential differences across the tonoplast. pHc was accordinglycomputed assuming either tonoplast or plasmalemma equilibrationof DMO, with correction for DMO metabolism within the cell.pH_c was 8.0–8.3 in the light in artificial seawater (pH_oabout 8.0), was some 0.5 units lower in the dark, and was slightlylower with an external pH of 7. Vacuolar pH was 6.5–6.9,without consistent effects of illumination or of external pHof 7 rather than 8. While µ_H+ at the tonoplast was similarto that in giant-celled freshwater algae (although with a greatercontribution from relative to pH), µ_H+ at the plasmalemmawas less than 8 kJ mol^–1, i.e. less than one-third ofthe value in freshwater green algae. µ_Na+ was some 13kJ mol^–1 at the plasmalemma. The possibility that theprimary active transport process at the plasmalemma of Chaetomorphadarwinii (and certain other marine algae) is Na⁺ efflux ratherthan H⁺ efflux is discussed.     

Characteristics of Radial Solution Flow in Roots of Cucumber (Cucumis sativus L.)

Low salt roots of Cucumis sativus L. cv. Burpeeana Hybrid weresubjected to iso-ionic treatments in which the external solutionconcentration of K⁺ was maintained at 14 mM. Solution concentrationof varied from 0 to 14 mM, other anions compensating. When Cl^– was the compensating ion, its concentrationin the exudate increased during the first 4 h and thereafterwas nearly the same as that of the external solution in alltreatments containing I mM or more. After 8 h of equilibration the concentration in the exudate increased almost exactly as its concentrationin the external solution. Rates of exudation and K⁺ transportwere almost constant between I and 14 mM KNO₂. More Cl^–was transported from solutions of similar Cl– concentrationwhen was also present. When water transport was inhibited with mannitol in treatments containing both KNO₃and KCI, exudate concentrations of K⁺ and were increased, but exudate concentration of Cl^– was notsignificantly affected except at the highest Cl     

The Uptake and Distribution of Copper in Some Forage Grasses as Affected by Nitrate-nitrogen Supply in Flowing Solution Culture

The effect of changes in nitrate-nitrogen supply on the absorptionand distribution of copper was examined in grasses grown inflowing solution culture with a maintained concentration ofcopper. Absorption by roots (µg Cu g^–1 dry root)decreased markedly when nitrogen had been depleted or was maintainedat 0.1 mg l^–1 N, but there was an immediate increase whennitrogen was maintained at 1.0 or 10.0 mg l^–1. There werealso large increases in the concentration of copper in the shootsof plants grown with 1.0 and 10.0 mg 1^–1 N. The rootsof plants grown with 0.1 or 1.0 mg 1^–1 N retained similarproportions of uptake, but a lower proportion was retained whenthe plants were grown with 10.0 mg 1^–1. Although a lowerproportion of the copper was associated with cell walls in theplants grown at 10.0 mg 1^–1 N this was the result of alower content of cell walls rather than an effect on copperitself. In a longer-term experiment in conventional solutionculture with a range of nitrogen concentration, the concentrationof copper in shoots was largely determined by shoot growth. Dactylis glomerata, Festuca arundinacea, Lolium perenne, cell walls, copper absorption, copper distribution, flowing solution culture, nitrate-nitrogen     

Effects of External Potassium Supply on Compartmentation and Flux Characteristics of Radiocaesium in Intact Spring Wheat Roots

Originally published in Annals of Botany84: 639–644 1999.For technical reasons beyond our control the flux symbol wasomitted from this paper. The paper is reprinted here in itsentirety. Short term experiments investigated the effects of potassiumsupply on radiocaesium influx/efflux and the radiocaesium compartmentationin intact spring wheat roots. Short term (24–72 h) influxanalysis showed that net influxes of radiocaesium to both rootand xylem were reduced approximately ten-times by increasingexternal potassium concentration from 50 µM to 200 µM.Efflux analysis distinguished three components for radiocaesium(namely cell wall+free space, cytoplasm and vacuole) and showedthat the rates of Cs⁺efflux at an external potassium concentrationof 100 µM (19.16 and 1.70 Bq g^-1min^-1for _coand _vo, respectively)were about three-times faster than those at 50 µM (7.24and to 0.41 Bq g^-1min^-1for _coand _vo, respectively). The resultsalso showed that external potassium concentration did not havea significant effect on the distribution of¹³⁷Cs between cytoplasmand vacuole, as indicated by the ratio of¹³⁷Cs in the two compartments.Results obtained in this study suggested that the inhibitoryeffect of potassium on the net uptake of radiocaesium by theplant root may be partially ascribed to the fact that at higherexternal potassium concentrations Cs⁺efflux rates were muchhigher. The mechanisms involved are discussed. Copyright 2000Annals of Botany Company Compartmentation, efflux analysis, potassium, radiocaesium, Triticum aestivum, wheat.     

Effect of Light Intensity on Adaptation of Dunaliella to Very High Salt Concentrations

Light intensity was found to have a strong effect on the adaptationto high salt concentrations of a green microalga, Dunaliellaparva, normally grown at low and medium salt concentrations.At high light intensities (200 µmol m^–2 s^–1)the cell glycerol content increased in parallel with an increasein external salt concentration; protein synthesis and cell divisioncontinued with no period of arrest. At low light intensitiesno glycerol synthesis occurred as the external salt concentrationwas raised; protein synthesis and cell division were equallyarrested. The importance of high light intensity during theinitial phase of increase of salt concentration was demonstratedand was found to be a requirement for protein synthesis andcell division. In experiments designed to discover the effectof light intensity on cells growing in media in which the saltconcentration was kept constant, it was confirmed that the lightintensity needed for growth increases as the salt concentrationof the medium is increased. Key words: Dunaliella, salt concentration, light intensity, growth rates     

Effects of sodium, potassium and chloride ions on the membrane potential of Valonia aegagropila

Changes of the resting potential of Valonia cell in sea wateragainst a 10-fold increase of the external concentrations ofK^$, Na^$ and Cl^– were 1±1, 6.2±0.1 and 38.9±4mV, respectively. The potassium conductance was smaller than7 µ/cm², while the Na and Cl conductances were 45 and281 µ/cm², respectively, in normal sea water. The positivevacuolar potential could be explained by these ionic conductances.On the other hand, the membrane became more sensitive to K^$,if the cell was incubated for about 30 min in K-rich (100 mM)sea water. It is worth noting, however, that the membrane conductancewas lower in the K-rich sea water than in the normal sea water. (Received October 7, 1974; )     

A Novel Method for Extracting Protoplasts from Large Brown Algae

Protoplasts have been isolated without the application of walldegrading enzymes from three large brown algal species: Macrocystisangustifolia, Ecklonia radiata and Durvillaea potatorum. Thecentral feature of this new protocol is the removal of wall-boundcalcium by substitution with sodium from the isolation medium.The new protocol is specific for cortex and inner meristodermcell walls with highest yields obtained from meristematic oryoung tissue. Protoplasts, extracted with this method, are approximately5–10 µm in diameter with viability estimates rangingfrom 73–86%. Consistent yields of 10⁷ protoplasts g^–1fresh weight have been obtained within 2–3 for all threespecies and this compares favourably with yields achieved usinga conventional enzyme-based system. Key words: Brown algae, protoplasts, alginate, calcium, enzymes     

Corrigendum

Due to an apparent fault in the telex system, a number of mistakeswere not corrected in this paper. The corrected lines are givenbelow p. 539: line 17In vivo fluorescence action spectra of chl a p. 541: Figure 2 legend, line 92.5 µW cm^–2 at 550nm (0.12 µE m^–2 s^–1 p. 542: Figure 3 legend, line 5( 89 µE m^–2 s^–1).Monochromatic beam intensity was 6 µW cm^–2 at 550nm (–0.28 µE m^–2 s^–1), Figure 3 legend, lines 8 and 9with intensity of 3 mW cm^–2( 179 µE m^–2 s^–1) Monochromatic beam intensitywas 7 5 µW cm^–2 at 550 nm (0.35 µE m^–2s^–1). line 6tivity. The match between the spectra of chl a fluorescenceand PSII O₂ evolution is lines 23–25fluorescence increasingly deviate from thoseof PSII O2 evolution. We attribute this discrepancy to selectivelight scattering by the algae. This scattering increases substantiallywith decreasing wavelength at that region when using a standardspectrofluorometer p. 543: Figure 4 legend, lines 3 and 4as in Chroomonas, withintensity of 2 mW cm^–2 ( 120 µE m^–2 s^–1).Monochromatic beam intensity was 15 µW cm^–2 at 550nm ( 0.7 µE m^–2 s^–1). line 7:ment types in the oceans. While all photoautotrophicorganisms have chl a (the bulk lines 12 and 13:and Barrett (1983). Our spectral data reflectthe great variability in pigment composition and functionalassociation in the major groups of algae. lines 25 and 26:Hiller, 1983). However, blue-violet and redPSII activity is much lower in cryptomonads than might be expectedfrom their absorption spectra (Haxo and Fork, 1958; Haxo, 1960), p. 544: Table I, column 3, entry 6:R-phycocyanin Table 1 legend, lines 1 and 2:^aHaxo and Blinks (1950); ^bFork(1961); ^cHaxo et al. (1955); ^dO'Carra and O'hEocha (1976); ^cresemblesDelesseria decipiens, Haxo and Blinks (1950, Figure 20); ^fHaxoand Fork (1969), like lines 1 and 2:I) indicates that these pigments are affiliatedwith PSII, perhaps exclusively, as in the red algae. In Rhodomonas,the peak of activity at 465 nm may be due to chl c absorp- p. 545: line 2:xanthophyll (fucoxanthin in diatoms, peridininin dinoflagellates and chl c. The ac line 7:shown to be similar to those of the much-studied Chlorella(Vidaver, 1966; Ried, 1972). line 14:tivity of PSII in algae. Spectrofluorometers, with theirsuperior sensitivity and stability, line 34:of natural populations, making these spectra more similarto the PSII photosynthesis lines 36 and 37:Large differences between the values of FIIfor different components within the algal population can distortfluorescence spectra, if they do not correspond with dif lines 41 and 42:different components are not similar to eachother. The necessary handling procedures of natural samples,such as filtration (Yentsch and Yentsch, 1979; Neori et al.,1984), p. 546: lines 20 and 21:DOE contract DE-AT03-82ER60031, anda grant to A.N., O.H.H. and F.T.H. from the Foundation for OceanResearch. Travel to the IInd GAP workshop was facilitated by lines 25 and 26:the culture of Chroomonas, J.Lance for helpwith cultures, J. and E.Yguerabide for the use of their spectrofluorometer,C.R.Booth, Y.Blatt and L.Petrosian for technical line 28:This study was in partial fulfilment for a Ph.D. degreeby A.N. line 42:Dutton.H.J., Manning.W.M. and Duggar.B.M. (1943) Chlorophyllfluorescence and energy transfer in the     

The Titrimetric Assay of OH- and HCO3- Excreted by Ricinus Cultivated on NO3--containing Nutrient Media: The Influence of Ionic Strength, End Point pH and CO2 Supersaturation

Allen, S. and Allen, C. R. 1987. The titrimetric assay of OH^–and excreted by Ricinus cultivatedon -containing nutrient media: the influence of ionic strength, end point pH and CO² supersaturation.–J. exp. Bot. 38: 607–617. When spent -containing nutrient media were titrated to the starting pH of 6.5 the titre wasequivalent to 50%orless of the base (i.e. ) excreted. Calculation of the total amount of baseexcreted could only be made from data obtained by titrationto pH 4–5. An accurate calculation of the amount of baserequired the inclusion of activity coefficients, estimated fromionic strength, in the calculations. Spent nutrient media contained from four to ten times the concentrationof CO₂ predicted from equilibrium values. It is probable that is very slow. Key words: Ricinus, nitrate-N nutrient medium, base excretion, ionic activity coefficients, carbon dioxide supersaturation     

Effect of Abscisic Acid on the K+ Efflux and Membrane Potential of Nicotiana tabacum L. Leaf Cells

K⁺ efflux from tobacco (Nicotiana tabacum L, cv. Samsun NN)leaf discs into the external medium was increased and the membranepotential (Em) changed in the positive direction with a changein pH from 8.0 to 4.0. E_m was affected by the external concentrationof KCl, greatly decreasing with a change in concentration from1 mM to 100 mM. The equilibrium potential of the membrane forK⁺ (E_k) was decreased in a Nernst fashion with increasing externalconcentrations of KCl. E_k is more positive than E_m above ca.50 µM KCl. Most of the experiments were carried out underconditions in which the difference between the electrochemicalpotential for K⁺ on the inside to the outside of the cell (µ_kis positive. Thus, K⁺ may passively flow to the outside of thecells accompanied by the depolarization of the membrane. Abscisic acid (ABA) increased the K⁺ efflux under conditionsof passive transport. K⁺ efflux was accelerated with an increasingconcentration of ABA, being maximal at 10^–4 M–10^–3M. This acceleration was due to the enhancement of the potassiummotive force (µ_k/F) which is the force causing the netpassive transport of K⁺. The membrane potential was decreasedfrom –205 mV to –170 mV by 2 x 10^–4 M ABAwithin 10 min. The depolarization was not transient, being lostfor at least 3 hr. These results show that ABA accelerated passive K⁺ efflux, whichaccompanied depolarization of the membrane. (Received June 22, 1981; Accepted August 24, 1981)     

The Uptake of Nitrate by Lolium perenne from Flowing Nutrient Solution: I. EFFECT OF CONCENTRATION

Experiments with simulated swards of perennial ryegrass (Loliumperenne L.) show the relationship between concentration in flowing nutrient solution, nitrate uptake,plant growth, and the chemical composition of roots and shoots.Rates of uptake exceeding 1 g N m^–2 d^–1 were maintainedat concentrations in solution down to 0•02 mg N l^–1. Short-term studies confirmed that at such lowconcentrations the plants were able to maintain rates of uptakeof about 85% of maximum. Between 0•2 and 200 mg N l^–1the concentration of in solution had little effect on rate of uptake or plant growth. With at 1000 and 2000 mg N l^–1 there was a marked reductionin weight of the shoots and, more particularly, in the lengthand tensile strength of the roots. There were several significanttrends in mineral composition of the plants (notably in S, Ca,Mg) which were apparently correlated with increasing concentrationof in solution.     

Carbon Metabolism in Seagrasses: III. ACTIVITIES OF CARBON-FIXING ENZYMES IN RELATION TO INTERNAL SALT CONCENTRATIONS

The internal salt content and distribution in photosynthetictissues as well as the effect of NaCl on photosynthetic carbonfixation enzymes was investigated in two seagrass species fromthe Red Sea. Concentrations of both Na⁺ and Cl^– were lower in the chloroplast-richepidermis than in underlying cell layers in Halophila stipulacea.In Halodule uninervis, the concentration of Na⁺ was lower inthe epidermis than in the underlying cells, while K⁺ was evenlydistributed between cell layers. The epidermal concentrationsof Na+ were estimated to be 0.17 and 0.10 M for Halophila stipulaceaand Halodule uninervis, respectively, which were about to the average leaf concentrations. Epidermal Cl^– concentrationof Halophila stipulacea was estimated to be 0.08 M, a valueonly about of the overall leaf concentration. Phosphoenolpyruvate carboxylase (PEPcase) extracted from leavesof these seagrasses showed increased activity at 0.05–0.3M NaCl in vitro. Ribulose-l, 5-bisphosphate carboxylase (RuBPcase)activity, on the other hand, was inhibited by NaCl at all testedconcentrations. At epidermal NaCl concentrations, PEPcase activitywas thus stimulated while RuBPcase was inhibited. The reducedRuBPcase activity at such concentrations compared to salt-freeconditions was still sufficient to account for observed photosyntheticrates. We conclude that these seagrasses have adapted to a saline environmentboth by maintaining relatively low ion concentrations in theepidermis where photosynthesis occurs and by having carbon-fixingenzymes capable of functioning in the presence of salt.     

Apoplastic Sugar Concentration and pH in Barley Leaves Infected with Brown Rust

Soluble sugars were extracted by low speed centrifugation fromthe apoplast of leaves of barley (Hordeum distichum L.) infiltratedwith water. Infection of the leaf with the brown rust fungus(Puccinia hordeii) resulted in a reduction in the concentrationof sucrose, glucose and fructose in the apoplast. Sugars werepresent in an apoplastic space occupying 12 and 17 cm³ m^–2of leaf area in healthy and infected tissue, respectively. Uptakeof hexoses by intercellular hyphae is suggested as a cause ofthis reduction. The pH of apoplastic sap extracted from rust-infectedleaves was increased to pH 7·3 from pH 6·6 incontrols. The effect of a reduced apoplastic sugar pool andincreased pH on export from infected leaves is discussed. Key words: Apoplast, barley (Hordeum distichum L.), brown rust (Puccinia hordeii Otth.), pH, sucrose, hexose     

Changes in the Levels of Ribulose-l,5-bisphosphate Carboxylase/Oxygenase (Rubisco) in Tetraedron minimum (Chlorophyta) during Light and Shade Adaptation

Exponentially growing cultures of the chlorophyta Tetraedronminimum were allowed to photoadapt to low (50µmole quantam^–2s^–1) and high (500µmole quanta m^–2–1)irradiance levels. In these cultures, various aspects of theorganization of the photosynthetic apparatus and related differencesin its performance were studied. In this organism, the observed five-fold increase in pigmentationof low-light adapted cells was due to increases in the numbersof PSU's, while their sizes remained constant. Using radioimmunoassay technique, we found that high-light adaptedalgae had over five times more Rubisco per PSU than their low-lightadapted counterparts. The high-light adapted algae also exhibited far higher (x2.3)light saturated photosynthetic rates per chl a. This increasewas the result of a reduction of tau, , the turnover time ofPS II reaction centers. We propose that the increase in Rubisco per PSU in high-lightadapted algae explains the reduction in , which results in thehigher P_max rates per chl a in these algae. The relationship is non linear, since the increase in Rubiscoper PSU was x5.3 whereas that in P_mM per chl a was only x2.3. (Received July 30, 1988; Accepted December 2, 1988)     

The Investigation of Stomatal Ionic Relations using Guard Cell Protoplasts: II. OSMOTIC RELATIONS OF GUARD CELL PROTOPLASTS IN SHORT AND LONG-TERM INCUBATION

Clint, G. M. 1985. The investigation of stomatal ionic relationsusing guard cell protoplasts. II. Osmotic relations of guardcell protoplasts in short and long-term incubation.—J.exp. Bot 36: 1739–1748 Measurements were made of the volume changes exhibited by isolatedguard cell protoplasts (GCPs) of Commelina communis L, whenexposed to a range of concentrations of external osmotica Inshort-term incubation, GCPs behaved as osmometers and showedrapid volume changes in response to changing external osmoticpressure (₀). In long-term incubation, GCPs prepared and incubatedwith added external KCl showed further slow changes in volume,in a manner suggesting that regulation of volume occurred. Protoplastsprepared and incubated without added external KCl had smallervolumes for a given value of ₀, and their ability to regulatevolume in long-term incubation was reduced or absent. Treatment with fusicoccin caused an increase in both the volumeand the K⁺ content of GCPs. The increase in volume continuingafter the increase in K⁺ content had ceased, in a manner similarto that observed in walled guard cells in epidermal strips. Key words: Guard cell protoplasts, volume regulation, Commelina communis     

Steady-state rotifer growth in a two-stage, computer-controlled turbidostat

Steady-state populations of rotifers (Brachionus calyciflorus)were maintained in twostage, continuous-flow turbidostatic cultureon the green alga Chlorella pyrenoidosa. In this system, themaximum specific growth rate,µ^max of the rotifers wasmaintained by using a computer to control the concentrationof algae, as rotifer food, in the rotifer culture. As rotifersconsumed algae, the turbidity decreased until a set-point wasreached. Then fresh algal suspension (supplied from a steady-statealgal chemostat) was metered into the rotifer culture, whichwas held in the dark. Rotifer and algal populations, as wellas rotifer µ^max entered steady states. These steady-stateresults were consistent with previous data from chemostat studies,but growth transients indicated that the of the µ^maxrotifersmay be subject to selection. The system is unique in providinga means to explore population dynamics of a metazoan maintainednear its µ^max.     

Development of Eodiaptomus japonicus Burckhardt (Copepoda, Calanoida) reared on different sized fractions of natural plankton

The nutritional value of different sized fractions of naturalplankton was investigated for the growth of Eodiaptomus japonicusBurckhardt by comparing the development of its naupliar andcopepodid stages fed on differentially fractionated planktonicassemblages of a eutrophic pond, at 20°C. Water filteredthrough a 0.8 µm Nuclepore filter, containing mainly smallcoccoid bacteria (0.45–0.6 µm in cell diameter),at a concentration of 82.7 µg C 1^–1 could not supportthe development of E.japonicus. The 3 µm filtered water,containing bacteria and picoalgae. at a total concentrationof 259 µg C 1^–1, supported development but not eggproduction. The 20 µm filtered water, containing bacteria,picoalgae and large algae, at a total concentration of 2600µg C 1^–1, supported rapid development of the juvenilesand continuous egg production by the adults. The separated 3–20µm fraction, containing only large algae, could not supportthe development at concentrations of 131 and 196 µg C1^–1. However, the same rapid development of the juvenilesand continuous egg production by adults occurred at all of thetested concentrations between 261 and 3920 µg C1^–1of the large algae. The results suggest that E.japonicus favoursalgae larger than 3 µm during its complete lifespan, andthat the threshold food concentration for its development variesbetween 200 and 250 µg C 1^–1.     

Calcification in the Green Alga Halimeda: IV. THE ACTION OF METABOLIC INHIBITORS ON PHOTOSYNTHESIS AND CALCIFICATION

The effects of a number of metabolic inhibitors on calcificationand photosynthesis in Halimeda tuna, H. discoidea, and H. macrolobaare described. The inhibitors used are CCCP, DNP, DCMU, azide,cyanide, chloramphenicol, cycloheximide, and Diamox. The effectsof these inhibitors, although complex, are consistent with ourmodel of calcification in Halimeda. Inhibition of photosyntheticCO₂ uptake inhibits calcification as does stimulation of respiratoryCO₂ evolution (i.e. uncoupling). There is also indirect evidencefor the presence of a possible light stimulated H⁺ efflux whichinhibits calcification. The observed calcification rate is thereforethe result of a number of factors which affect the concentrationof CO^–and the pH in the intercellular space of the Halimedathallus. The results obtained with the carbonic anhydrase inhibitor Diamoxprovide further evidence for the effective separation of theintercellular space from the external medium by the appressedperipheral utricles.     

The Titrimetric Assay of H+ Excreted by Ricinus communis Cultivated on NH4+-N Nutrient Media: The Effect of Ionic Strength and Nutrient Phosphate Concentration

Allen, C. R. and Allen, S. 1987. The titrimetric assay of H⁺excreted by Ricinus communis cultivated on -N nutrient media; the effect of ionic strengthand nutrient phosphate concentration.— J. exp. Bot. 38:597–606. The simple titrimetric assay of H⁺ excreted by -N plants into spent nutrient solution was criticallyassessed on the basis of equations for charge balance, conservationof matter and chemical equilibrium (including activity coefficients).For Ricinus communis, a simple volumetric calculation from back-titrationof the spent medium to 6–5 underestimated proton excretionby 3%, compared with the corresponding error of up to 50% fortitrimetric assay of base excreted into spent -N media. Key words: Ricinus, ammonium-N nutrient medium, proton excretion, ionic activity coefficients     

Enhanced suicidal death of erythrocytes from gene-targeted mice lacking the Cl-/HCO(3)(-) exchanger AE1

Genetic defects of anion exchanger 1 (AE1) may lead to spherocytic erythrocyte morphology, severe hemolytic anemia, and/or cation leak. In normal erythrocytes, osmotic shock, Cl^– removal, and energy depletion activate Ca²⁺-permeable cation channels with Ca²⁺-induced suicidal erythrocyte death, i.e., surface exposure of phosphatidylserine, cell shrinkage, and membrane blebbing, all features typical for apoptosis of nucleated cells. The present experiments explored whether AE1 deficiency favors suicidal erythrocyte death. Peripheral blood erythrocyte numbers were significantly smaller in gene-targeted mice lacking AE1 (AE1^–/– mice) than in their wild-type littermates (AE1^+/+ mice) despite increased percentages of reticulocytes (AE1^–/–: 49%, AE1^+/+: 2%), an indicator of enhanced erythropoiesis. Annexin binding, reflecting phosphatidylserine exposure, was significantly larger in AE1^–/–erythrocytes/reticulocytes (10%) than in AE1^+/+ erythrocytes (1%). Osmotic shock (addition of 400 mM sucrose), Cl^– removal (replacement with gluconate), or energy depletion (removal of glucose) led to significantly stronger annexin binding in AE1^–/– erythrocytes/reticulocytes than in AE1^+/+ erythrocytes. The increase of annexin binding following exposure to the Ca²⁺ ionophore ionomycin (1 µM) was, however, similar in AE1^–/– and in AE1^+/+ erythrocytes. Fluo3 fluorescence revealed markedly increased cytosolic Ca²⁺ permeability in AE1^–/– erythrocytes/reticulocytes. Clearance of carboxyfluorescein diacetate succinimidyl ester-labeled erythrocytes/reticulocytes from circulating blood was more rapid in AE1^–/– mice than in AE1^+/+ mice and was accelerated by ionomycin treatment in both genotypes. In conclusion, lack of AE1 is associated with enhanced Ca²⁺ entry and subsequent scrambling of cell membrane phospholipids. annexin; cell volume; osmolarity; phosphatidylserine; energy depletion