Algorithm and computer program Pro--Anal for analysis of relationship between structure and activity in a family of proteins or peptides

In this paper we introduce a computer algorithm and programPro—Anal for analysis of the structure-activity relationshipin a family of evolutionarily related (and/or artificially mutated)proteins/peptides. The program uses aligned amino acid sequenceswith data of their activity (pK, K_m, ED₅₀ or any other) andsearches for correlations between data on activity and variousphysico-chemical characteristics of different regions in primarystructures. In automatic mode, the program generates and verifieshypotheses on the disposition of a sequential modulating regionin a protein, and key characteristics of the region. In manualmode, users can generate and analyze their own hypotheses. Theprogram is implemented on IBM PC or compatible computers. Itis designed to be easily handled by the occasional computeruser and yet it is powerful enough for experienced professionals.Pro—Anal operation is demonstrated on the example of findingmodulating centers in a family of disintegrins—proteinsfrom snake venoms which inhibit fibrinogen interaction withplatelet receptors. In another example it is shown that theimmunogenicity of peptides is connected with their positivecharge.     

The Red-Eared Slider Turtle: An Animal Model for the Study of Low Doses and Mixtures

Current risk assessment techniques for almost all chemicalsinvolve establishing a threshold dose, or the dose below whichno adverse effects are seen. But threshold doses may not applyin situations where the chemical mimics the actions of an endogenouscompound—such as a steroid hormone—that is importantto development. In such cases, any exposure may exceed the threshold.Recent studies with the red-eared slider turtle have shown thatexogenous estradiol—even when applied to eggshell in dosesas low 0.4 ng—will affect sex development during embryogenesis.Considering that only 0.2% of the estradiol applied to the eggshellends up in the embryo, it becomes apparent that even very lowdosages of steroid hormones or their mimics can have profoundbiological effects. We tested this idea using eight compoundsidentified in the yolk of alligator eggs from Lake Apopka, Fla.Five of the compounds—the PCB mixture Aroclor 1242,trans-Nonachlor,cis-Nonachlor,p,p'-DDE, and chlordane—altered sex ratio outcomes whenapplied to eggshells during development. Aroclor 1242 producedthe most powerful effects, shifting the ratio of females almosttwofold, while chlordane had the greatest effect when combinedwith estradiol. Administration of all eight compounds togetheralso increased the ratio of females to males. However, comparingthe single-compound exposures at the same dosages indicate thatthese compounds behave differently in combination than theydo singly, emphasizing the need for further studies using chemicalmixtures reflecting proportions found in nature. The effectof chlordane and Aroclor 1242 on aromatase activity levels duringembryogenesis in the brain and adrenal-kidney-gonad (AKG) complexwas also examined. Chlordane, a suspected anti-androgen in thisspecies, did not affect aromatase activity in either the brainor the AKG. However, Aroclor 1242 significantly altered aromataseactivity levels in the red-eared slider turtle brain—butnot in the AKG—during a crucial developmental period.After this crucial period, Aroclor 1242 caused an increase inaromatase activity in the AKG of embryos just prior to hatch.Additionally, hatchling males treated during embryogenesis withAroclor 1242 and chlordane exhibited significantly lower testosteronelevels than controls in response to follicle-stimulating hormoneadministration, while chlordane-treated females had significantlylower progesterone, testosterone, and 5-dihydrotestosteronelevels relative to controls. These results are similar to thosefound in juvenile alligators from Lake Apopka. Males treatedwith Aroclor 1242 and trans-Nonachlor displayed an elevatedestradiol response to FSH administration vs. control males.Taken together, these results suggest that EDCs exert effectsduring embryonic development that extend beyond birth. Theyalso suggest that the alterations in sex steroid hormone levelsobserved in animals from contaminated areas may result fromEDC-induced alterations in the neuroendocrine axis controllinggonadal sex steroid hormone production.     

Cell Locomotion and Contact Guidance in Amphibian Gastrulation

Presumptive mesodermal cells in amphibian gastrulae migratefrom the blastopore toward the animal pole by using the innersurface of the ectodermal layer as their substratum. Duringmigration, the mesodermal cells form lamellipodia and filopodiapredominantly in a direction toward the animal pole. There isa network of the extracellular fibrils on the inner surfaceof the ectodermal layer. The fibrils seem to serve as an adequatesubstratum for attachment of the filopodia and locomotion ofthe mesodermal cells. A significant alignment of the fibrilnetwork along the blastopore—animal pole axis suggestsa hypothesis that it directs morphogenetic cell movements bycontact guidance in combination with contact inhibition of movement.New culture conditions allow the gastrula mesodermal cells tomove actively in vitro with a similar cell shape and at a similarrate as in vivo. Such culture conditions enabled an in vitroexperiment to test the hypothesis of contact guidance. Explantedectodermal layers deposit the fibril network on the surfaceof a cover slip. Dissociated gastrula mesodermal cells seededon such a conditioned surface attach to the surface and moveabout actively. A computer analysis of the time—lapsefilms shows that the cell trails are significantly aligned alongthe blastopore—animal pole axis of the ectodermal layerthat conditioned the surface. The deposited fibril network showsthe alignment along the same axis. There is also a tendencyof the mesodermal cells to move in a polarized fashion preferentiallytoward the animal pole. These results support the hypothesisof contact guidance of mesodermal cell migration in vivo byoriented extracellular fibrils     

Nuclear Chromatin Structure in Relation to Cell Differentiation and Cell Activation in the Cap and Quiescent Centre of Zea mays L.

Barlow, P. W. 1985. Nuclear chromatin structure in relationto cell differentiation and cell activation in the cap and quiescentcentre of Zea mays L —J. exp. Bot. 36: 1492–1503.Nuclear chromatin structure has been analysed by electron microscopyof thin sections of cells in four zones of the root cap—meristem,central, slime-secreting and outermost cells—and alsoin the quiescent centre of the root before and after decapping.The chromatin pattern has been related to the DNA and RNA syntheticactivity of the nuclei. During cap cell maturation there wasa progressive condensation of the chromatin and this was accompaniedby some reduction of RNA synthesis. The degree of condensationwas estimated from the area and number of pieces of electrondense chromatin which increased and decreased, respectively,during cap maturation. The volume fraction of condensed chromatinwas also estimated but, in the cap, was not found to be a goodindicator of nuclear activity. The outermost cells of the capshowed the greatest degree of chromatin condensation but werestill active in RNA synthesis. Microdensitometry of their nuclearDNA contents gave an indication of loss of DNA in some of thenuclei. Decapping activated DNA and RNA synthesis in the quiescentcentre and also stimulated a decondensation of chromatin: thenumber of condensed pieces of chromatin increased, and theirsize and volume fraction both decreased 4 h after decapping.The number of pores per unit length of nuclear envelope profilewas also estimated. In the cap this number increased duringcap maturation; in the activated quiescent centre the numberremained constant except for a small rise 4 h after decapping Key words: Zea mays, chromatin, root cap, quiescent centre     

Endosperm Development in Solanum nigrum L. Formation of the Zone of Separation and Secretion

Endosperm development in Solanum nigrum was ab initio cellular.During early seed development, a Zone of Separation and Secretion(ZSS) differentiated within the endosperm. There were threephases in the formation of the ZSS. Phase I—stainabilityof the cell walls and middle lamella increased followed by numeroussmall plasmalemma invaginations (blebs) which became filledwith a fibrillar material. Phase II—the plasmalemma withdrewfrom the cell wall as a fibrillar lipo-carbohydrate matrix accumulatedoutside the plasmalemma. The middle lamella was gradually removedfrom between the cells forming the central axis of the cone.Phase III—the lipo-carbohydrate matrix continued to accumulateoutside the plasmalemma and also within the developing intercellularspaces. Some axial cells were completely separated from theremaining ZSS cells and became embedded in the matrix. The formationof the ZSS did not entail the destruction of the endosperm cellsand cell divisions were frequent. The ZSS was initially cone-shaped,capping the globular embryo. As the embryo sac enlarged, theZSS continued to differentiate. This resulted in a narrow curvedcorridor through the peripheral region of the endosperm whichterminated above the vascular trace. The embryo grew throughthe centre of the ZSS and pushed aside the separated axial cells.The ZSS facilitated the growth of the embryo through the restof the endosperm.Copyright 1993, 1999 Academic Press Lipo-carbohydrate matrix, extracellular matrix, endosperm development, Solanum nigrum, Zone of Separation and Secretion     

cytokinin-induced amylolytic activity in bean cotyledons: Identification of the regulated enzyme

Separation of an extract of cotyledons of Phaseolus vulgarison a column of Sephadex G-200 revealed three amylolytic fractions.The slower migrating fraction hydrolyzed/ß-Limit dextrinazure and was inhibited by EDTA. The activity of this fractionwas enhanced by the embryo axis and this effect could be fullyreplaced by kinetin or benzyladenine. These results suggestthat the bean embryo axis exerts a promotive influence on theactivity of a-amylase in the cotyledons and that this effectis mediated by cytokinins. The other two amylolytic fractions did not show a-amylase activity.No effect of the embryo axis or of cytokinins on their activitycould be noted. ¹Present address: The Thimann Laboratories, University of California,Santa Cruz, California, U.S.A. (Received June 27, 1979; )     

Purification and Properties of Poly(A) Polymerase from Vigna unguiculata

Poly(A) polymerase was purified from germinating Vigna unguiculataseeds by successive column chromatography on phosphocellulose,Toyopearl HW-55S, heparin-Sepharose and TSKgel phenyl-5PW, whichyielded two activity fractions. The first fraction was purifiedas a single polypeptide with a mol wt of 63,000 as estimatedby SDS-PAGE. The enzyme activity was highly specific for ATPand required Mn²⁺ ion; an ATP-Mn complex may be the actual substrate.The polymerization reaction required a primer, with varioustypes of RNAs, poly(A) as well as dinucleoside phosphates having3'—OH, serving as efficient primers. The two forms ofthe enzyme had very similar properties with respect to divalentcation requirement and dependency on ion strength, but theyshowed some difference in primer preference. (Received March 4, 1988; Accepted May 2, 1988)     

Activation of Nitrite Reductase in a Cell-Free System from the Denitrifier Alcaligenes sp. by Freeze-Thawing

Nitrite reductase (NiR) activity of the cell-free extract orthe soluble fraction prepared from cells of Alcaligenes sp.NC1B 11015 grown anaerobically in the presence of nitrate wasexamined by measuring the rate of nitrite disappearance withdithionitemethyl viologen (MV) as an electron donor. Freezingat — 20?C and subsequent thawing of the fraction resultedin 5-40 times increase of the specific activity of NiR. Fromthe experiments on the effect of freezing conditions on theactivation, the phase change of solvent water due to freezingis considered to play an important role in the activation. Thisactivation occurred with the preparation in the exponentialgrowth phase, but not that in the stationary growth phase. Clearly,the low-molecular-weight (< 12,000) component which was obtainedfrom the soluble fraction through a collodion bag participatedin the activation. The activated enzyme proved to be the dissimilatory NiR, becauseNO production from nitrite, one of the typical characteristicsof the dissimilatory NiR, was also activated when assayed withascorbate-tetramethyl-p-phenylene diamine (TMPD) as an electrondonor. Nevertheless, the reaction products of nitrite reductionwere identified as hydroxylamine and ammonia with dithionite-MV.The possible pathway of nitrite reduction with this electrondonor is discussed. (Received May 26, 1983; Accepted February 2, 1984)     

The Effect of Temperature and Moisture Content on the Longevity of Seed of Ulmus carpinifolia and Terminalia brassii

Seeds of the Smooth-leafed Elm (Uimus carpinifolia) and of thetropical forest tree Terb (Terminalia brassii) were stored hermeticallyand sampled at intervals for periods of up to two years. Bothspecies possess ‘orthodox’ seed (increasing longevityis observed as either moisture content or temperature are reduced)within the temperature ranges from — 13 to 52°C (Elm)and from —4 to 42°C (Terb) and within the moisturecontent ranges from 3 to 19 per cent (Elm) and from 5 to 14per cent (Terb) on a fresh weight basis. Elm seed stored at—75°C showed the expected relationship between longevityand moisture content, but did not differ significantly in longevityfrom seed kept at — 13°C when moisture contents wereheld constant. Probit analysis of the relationship between germinationpercentage and time was performed for each storage environment,yielding a slope from which the standard deviation of the distributionof seed deaths over time () was calculated. Standard deviationvalues were used in turn to determine the values of constantsin a viability equation which had previously been applied toseed of barley, chickpea, cowpea and soybean. The equation,which gave a good fit to the results obtained, can be used topredict viability for seed in storage over a wide range of environmentalconditions. Some limitations to the applicability of the viability equationwere defined. At 22 per cent and higher moisture contents Elmseed survived longer than predicted. Furthermore, all Elm andTerb seed was killed quickly on placing in —75°C at22 and 20 per cent moisture content respectively, but high viabilitywas retained for several days at 19 and 17 per cent respectively.Practical implications of the results are discussed. Uimus carpinifolia Gleditsch, Smooth-leafed Elm, Terminalia brassii Exell, Terb, seed longevity, seed storage, moisture content, temperature     

The response of tomato roots (Lycopersicon esculentum Mill.) to iron deficiency stress: alterations in the pattern of protein synthesis

Dicotyledon plants adapt to iron (Fe) deficiency through a seriesof reactions that increase the ability of the plant to assimilateFe and to increase the efficiency of Fe utilization. In an attemptto gain an insight into these adaptive processes, the specificchanges in protein synthesis associated with the onset of theFe deficiency response in tomato roots (Lycopersicon esculentumMill cv. Rutgers) have been investigated. Roots were grown underFe—sufficient and —deficient conditions, and thepattern of protein synthesis was analysed by in vitro translationof root mRNA and by in vivo labelling of root proteins. Polypeptideswere resolved by two—dimensional polyacrylamide gel elec—trophoresis.Seven polypeptides were identified by in vitro translation,whose synthesis was significantly increased during Fe deficiency.The increase was probably specific to Fe deficiency in thatthe polypep—tide synthesis was not increased during phosphatedeficiency stress, was less prominent following prolonged Fedeficiency and was decreased following re—supply of Feto the hydroponic medium. The pattern of in vitro translation of mRNA isolated from Fe—deficientroots was compared to the results obtainedin vivo followingradiolabelling of proteins. In these analyses, eight polypeptideswere identified, tentatively including the seven polypeptidespreviously identified by in vitro translation. All polypeptideswere characterized with regard to molecular mass and pl andtheir localization in the cell, whether being membrane boundor soluble. It is suggested that members of this group of polypeptidesare involved in the response of the root to Fe deficiency: althoughtheir functions remain to be identified. Key words: In vitro protein synthesis, iron, iron deficiency, root, 2-dimensional PAGE     

Isolation and Identification of Nicotinic Acid as a Flower-Inducing Factor in Lemna

Extracts of flowering plants of the long-day plant Lemna gibbaG3 and the short-day plants Lemna paucicostata 151 and 381 weretested on L. paucicostata 151 for flower-inducing activity.Crude extracts failed to show any activity but after severalpurification steps three fractions with flower-inducing activitywere obtained. One fraction obtained from all three plants wasshown to contain nicotinic acid by mass spectroscopic and NMRspectroscopic analyses. These results raise the possibilitythat nicotinic acid may act to influence the flowering processin Lemna. (Received August 28, 1985; Accepted October 29, 1985)     

Cambial Activity in Zygophyllum dumosum Boiss

Cambial activity in Zygophyllum dumosum Boiss.—a dominantdesert shrub in Israel—was investigated. Cambial activityin young plants seems to be greatly affected by water supply,while cambial activity in old specimens was found to be moreendogenously controlled. A late-summer cambial dormancy wasobserved under field conditions even in plants that were irrigated.     

Glycosylation sites identified by solid-phase Edman degradation: O-linked glycosylation motifs on human glycophorin A

The human red blood cell sialoglycoprotein, glycophorin A (GpA),contains a ‘mucin-like’ extensively O-glycosylatedextracellular domain which carries the MN blood group antigens.We have revised the sites of O-glyccsylation in the extracellulardomain of GpA by automated solid-phase Edman degradation, whichallowed positive identification and quantitation of O-glycosylatedSer and Thr residues, as well as the single N-glycosylationsite. One N-linked and 16 O-linked sites were identified. Carbohydratewas absent on Ser 1, Ser14, Ser15, Ser23, Thr28 and Thr58 inGpA. We propose that the glycosyltransferases present in erythrocytesrecognize specific flanking sequences around potential O-glycosylationsites. All 16 O-glycosylation sites are explained on the basisof four motifs. Three motifs are associated with Thr-glycosylation:Xaa—Pro—Xaa—Xaa where at least one Xaa = Thr;Thr—Xaa—Xaa—Xaa where at least one Xaa = Thr;Xaa—Xaa—Thr—Xaa where at least one X = Argor Lys. The fourth motif is associated with Ser-glycosylation:Ser—Xaa—Xaa—Xaa where at least one Xaa = Ser.These simple rules explain the glycosylation (or lack of it)on 21 of 22 Ser/Thr in the extracellular domain of GpA. glycophorin A O-glycosylation motif solid-phase Edman degradation     

Changes in Abscisic Acid Content in Axis and Cotyledons of Developing Phaseolus vulgaris Embryos and their Physiological Consequences

Prevost, I. and Le Page–Degivry, M. Th. 1985. Changesin absicisic acid content in axis and cotyledons of developingPhaseolus vulgaris embryos and their physiological consequences.—J.exp. Bot. 36: 1900–1905.Changes in abscisic acid (ABA)content with time were measured in embryonic axes and in cotyledonsof Phaseolus vulgaris embryos using a radio–immunoassay.During embryogenesis, a similar pattern was observed in bothtissues: ABA increased to a maximum 29 d after an thesis, followedby a decrease as the seed matured. The level of ABA in the cotyledonswas always much higher than that in the axes. In in vitro cultures,the duration of the lag phase before germination of isolatedembryonic axes increased with ABA content. The presence of cotyledonsalways lengthened the lag phase; longer lag phases were associatedwith greater concentrations of ABA in the cotyledons. Moreoverthe presence of cotyledons stimulated the growth of seedlings. Key words: ABA distribution, embryo maturation, axis and embryo germinability     

The Messenger RNA Population in the Embryonic Axes of Phaseolus vulgaris During Development and Following Germination

Misra, S. and Bewley, J. D. 1985. The messenger RNA populationin the embryonic axes of Phaseolus vulgaris during developmentand following germination.—J. exp. Bot. 36: 1644–1652. Messenger RNAs were extracted from young, mid-maturation, late-maturation,mature-dry, and 20-h-germinated embryonic axes of Phaseolusvulgaris cv. Taylor's Horticultural. They were translated invitro in a rabbit reticulocyte lysate protein synthesizing system.Analysis of the translation products using two-dimensional polyacrylamidegel electrophoresis indicated that there were substantial changesin the messenger RNA populations of developing and germinatingaxes. The number of polypeptides synthesized increased sharplyat 20–22 d after pollination and then declined. Therewas a parallel increase and decrease in the Poly(A)⁺ contentof the seed axis. The analysis showed that certain messageswere present throughout development and were stored in maturedry seed. These messages were degraded upon subsequent rehydration.Some messages appeared during mid-maturation but declined duringlater stages of development and were absent from the matureseed. In the germinating seed a set of messages unique to germinationappeared. Key words: —Seed development, germination, mRNA, in vitro translation, Phaseolus vulgaris     

Response to Temperature in a Stand of Pearl Millet: 10. PARTITION OF ASSIMILATE

Effects of temperature on partition of assimilate between leaves,stems and panicles of pearl millet are analysed in terms ofa duration (t_w) over which a structure increased in weight,and a partition factor (p)—the fraction of new dry matterallocated to the structure during t_w. The value of tw was, forall structures, inversely proportional to temperature abovea base of 10 °C and below an optimum of 28 to 30 °C.For stems and panicles, the value of p was, with one exception,little affected by temperature. The dry weight of these structureswas, therefore, proportional to t_w, and decreased with risein temperature. (The exception was panicles at the lowest temperature,19 °C, for which p was reduced by 40% because few grainswere set.) For leaves, however, p increased with rise in temperature,counteracting the effect on t_w, such that dry weight changedlittle with temperature. The optimum temperature for reproductiveyield was 22 °C, but the proportion of the total dry matterallocated to reproductive structures changed little between22 °C and 31 °C. Key words: Pearl millet, temperature, thermal time, partitioning     

Endogenous Cytokinins in Developing Fruits of Seeded and Seedless Citrus Cultivars

This report describes the isolation and identification of endogenouscytokinins from Citrus ovaries. Cytokinin active fractions wereobtained by extraction with 80% (v/v) ethanol, followed by purificationwith hexane, n-butanol and a polyvinylpyrrolidone and SephadexLH-20 column chromatography. Five fractions with cytokinin activity were found in the organicphase, using the tobacco callus assay. The main active compoundsin these fractions were separated by HPLC, bioassayed and identifiedby GC—MS as ribosyl zeatin, zeatin and isopentenyl adenosine.Hydrolysis of the first fraction with B-glucosidase gave cytokininactive compounds that in paper chromatography had R_F's similarto those of zeatin and ribosyl zeatin. Treating the aqueousphase with alkaline phosphatase produced a cytokinin activecompound that in paper chromatography had the same R_F as isopentenyladenosine indicating that their ribotide was probably the majorphosphorylated cytokinin present in Citrus ovaries. Key words: Citrus, cytokinin fruit set and development     

Ethylene-bis-Nitrourethane: Investigation of the Plant Growth Stimulant Action of Analogous Compounds

The results of a systematic attempt to relate the growth-stimulatingfunction of compounds analogous to ethylene-bis-nitrourethaneto their chemical structure and properties showed that, withthe exception of a series of nitraminocarboxylic acids, NO₂NH(CH₂)COOH,activity was confined to compounds of the form R—N(NO₂)—X—N(NO₂)—R,where X is an alkylene or substituted alkylene chain. It issuggested that active compounds are converted to dinitramines(R = H) in the plant, and that the acidic nature of the —NHNO₂groups is of importance. The oil/water partition coefficientsand the geometrical structure of the molecules are also shownto influence their growth-stimulating activity.     

Water uptake resumption following soil drought: a comparison among four barley genotypes

Soil water resulting from episodic growing season rainfall evaporatesrapidly in semi-arid regions. Plants may mnot benefit from suchwater additions if near-surface roots are unable to resume wateruptake rapidly following periods of soil water deficit. Ourobjectives were to develop a means of quantifying root uptakeresponses in the upper soil layer following rewetting aftersoil water deficit, and to evaluate the existence of genotypicdifferences among four diverse barley (Hordeum vulgare L.) genotypesin this regard. Plants were grown in replicate soil columnshaving hydraulically isolated surface and subsoil layers, andinstrumented with time-domain reflectometry (TDR) waveguides.The upper 0.05 m soil layer was allowed to dry to —1.8to —3.0 MPa for 10-14 d, during which time subsoil wetnesswas maintained at about —0.6 to —0.7 MPa. The time-courseof soil water uptake was monitored at 0.5 h intervals followingrewetting of the surface layer. Substantial water uptake began1 d after rewetting following 10 d, and 2-3 d after rewettingfollowing 14 d of water deficit. Rate of water uptake was morerapid in response to a second rewetting 5-7 d later. Consistentgenotypic responses in terms of cumulative water uptake on awhole plant and leaf area-specific basis were observed duringeach trial. These results have application to evaluating droughthardiness and interspecific competitive ability under semi-aridconditions, and to investigations of root physiological andmorphological changes that contribute to recovery from waterdeficit Key words: Hordeum vulgare, root water uptake, soil water deficit, time-domain reflectometry     

Plasma membrane isolation from freshwater and salt-tolerant species of Chara: antibody cross-reactions and phosphohydrolase activities

Plasma membranes were isolated using the aqueous polymer two-phasepartition method from the algae Chara corallina and Chara longifolia,algae which differ in their ability to grow in saline environments.Enrichment of plasma membrane and depletion of tonoplast relativeto the microsomal fraction was monitored using phosphohydrolaseassays and crossreactions to antibodies raised against higherplant transporters. Antibodies to the vacuolar ATPase and pyrophosphatasecross-reacted with epitopes in the microsomal fraction, butshowed little affinity for the plasma membrane fraction. Pyrophosphataseactivity also declined in the plasma membrane fraction relativeto the microsomal fraction. The V-type H⁺ -ATPase activity,sensitive to nitrate or bafilomycin, was low in both fractions,though the cross-reaction to the antibody was reduced in theplasma membrane fraction. By contrast, the antibody recognitionof a P-type H⁺-ATPase amino acid sequence from Arabidopsis didnot occur strongly in the anticipated 90–100 kDa range.While there was enhanced recognition of a polypeptide at around140 kDa in the plasma membrane fraction, salt treatment of Charalongifolia resulted in plasma membrane fractions with reducedamounts of this epitope, but no change in vanadate-sensitiveATPase activity, suggesting that it does not represent the onlyP-type ATPase. Microsomal membranes from saltadapted C. longifoliahave higher reactivity with the antibody to the tonoplast ATPase. Key words: Chara, plasma membrane, salt tolerance, ATPase