Efflux of photosynthate and acid from developing seed coats of Phaseolus vulgaris L.: a chemiosmotic analysis of pump-driven efflux

Seed coats of Phaseolus vulgaris L. unload photosynthetic products,mineral ions and acid into the apoplastic space surroundingthe embryo. We report measurements, on detached seed coats,of the rates of unloading of photosynthates, ions and acid atdifferent external pH and in the presence of treatments intendedto alter the rate of proton pumping. We also report measurementsof membrane potential difference (PD) and of cytoplasmic pHunder the same conditions, measurements which have allowed usto validate the treatments we used and to investigate functionalrelationships between membrane processes. A chemiosmotic model of the seed-coat cell membrane is proposed,in which sucrose efflux and acid efflux are both driven by theproton pump. Sucrose efflux is proposed to occur by sucrose/protonantiport driven by the proton-motive force (PMF), and acid effluxto occur by pumped protons accompanied by a passive efflux ofanions. We use our measurements to estimate the net efflux ofsucrose on the antiporter and the total efflux of protons onthe pump. We have tested the model by using experimental treatments designedto manipulate the pump rate as the independent variable. Underthese conditions, and assuming the model is correct, the pumprate determines the cytoplasmic pH. Over the range covered byour experiments the net sucrose efflux is dependent on externaland cytoplasmic pH, the latter having the major role. The effluxof acid, under the same treatments, depends primarily on theproton pump rate, and was found to be well fitted by a quadraticfunction of pump rate. This means that, as pump rate increases,an increasing proportion of the pump output is used by acidefflux and a decreasing proportion by sucrose antiport. The membrane PD, although an important component of the PMF,does not appear to function in rate control of net sucrose orof acid efflux, since neither efflux is correlated with membranePD under our treatments which vary the pump rate. The PD correlateswell with external potassium concentration, and seems largelydetermined by the diffusion of potassium ions and anions. Key words: Phaseolus vulgaris L, photosynthate efflux, proton pump, sucrose/proton antiport, seed coat, membrane transport model     

Apical Senescence in Pisum: A Direct or Indirect Role for the Flowering Genes ?

Apical senescence was examined in a range of intact and defloweredflowering genotypes under both long and short photoperiods.The flower inhibitor produced by the gene Sn, appears to havea direct effect on apical senescence since it can delay apicalsenescence under short day conditions in the absence of flowerand fruit development or where the rate of such developmentis the same in different treatments. Gene Hr can magnify thiseffect. Gene E, on the other hand, appears to influence apicalsenescence only indirectly through the effect it has on flowerand fruit development. The flowering genes at the If, sn andhr loci are also thought to have indirect effects on apicalsenescence. Even in deflowered plants apical senescence appearsto occur eventually in continuous light in all genotypes testedindicating that the presence of developing fruits, althoughpromotory, is not essential for apical senescence. Pisum sativum L., garden pea, flowering, senescence     

Possible Roles of Methyl Glucoside andMyo-inositol in the Opening of Cut Rose Flowers

Methyl glucoside andmyo-inositol are present in all organs ofrose (Rosa hybridaL.). To investigate the possible role of thesecarbohydrates in the opening of cut roses, flowers with a 10,20 or 40-cm-long stem and a single flower bud (about 1.5 cmin diameter) were placed in water and flower opening and changesin sugar content in flowers and stems examined for 7 d. Thelonger the stem of the cut flower, the larger was the flowerdiameter. In stems, the concentration of carbohydrates, includingmethyl glucoside andmyo-inositol markedly decreased before floweropening. In petals, contents of glucose, methyl glucoside andmyo-inositolalso decreased before flower opening, but those of fructose,sucrose and xylose did not. When glucose and methyl glucosidewere added to the vase water (4%) flower opening was clearlypromoted; this was accompanied by an increase in methyl glucosideand fructose concentrations in petals. On the contrary,myo-inositolinhibited flower opening, and this was accompanied by an increaseinmyo-inositol and xylose concentrations in petals. These resultssuggest that methyl glucoside and/or its metabolites are transportedinto the petal cells, thereby lowering the osmotic water potentialand promoting flower opening.Myo-inositol is not readily metabolized,and exogenousmyo-inositol given at a high concentration mayact as an extracellular osmolyte, which inhibits water uptakeand flower opening.Copyright 1999 Annals of Botany Company Cut flowers, methyl glucoside,myo-inositol,Rosa hybrida,soluble carbohydrate.     

Stomatal Opening in Light of Different Wavelengths: Effects of Blue Light Independent of Carbon Dioxide Concentration

Stomatal opening in Xanthium pennsylvanicum was found to besignificantly greater in blue light than in red. Experimentsin which leaves were placed in a closed system and allowed toestablish their own steady-state carbon dioxide concentrationshowed that when the CO₂ concentration was about the same asthat in red, opening was much greater in blue light. Blue lightof low intensity could cause as great an opening as red of higherintensity, even though the CO₂ concentration was much higherin blue. Stomatal opening in light is considered as involvingat least two reactions: (1) a response to the removal of CO₂by photosynthesis; (2) a response to blue light not dependenton the removal of CO₂. Blue light became increasingly effective, relative to red, asthe length of night was increased over the range 2 to 14 hours.This might, in part, explain previously observed effects ofnight length on rate of opening in light. The initial very rapid phase of closure in darkness appearedto be independent of CO₂ accumulation, for it was not preventedby flushing the intercellular spaces with air free of CO₂. Itis suggested that closure in darkness, like opening in light,should be considered as involving components both dependentupon, and independent of, CO₂ concentration.     

Effect of the Rht3 dwarfing gene on dynamics of cell extension in wheat leaves, and its modification by gibberellic acid and paclobutrazol

The second leaf of wheat was used as a model system to examinethe effects of the Rht3 dwarfing gene on leaf growth. Comparedto the rht3 wild type, the Rht3allele decreased final leaf length,surface area and dry mass by reducing the maximum growth rates,but without affecting growth duration. Gibberellic acid (GA₃)increased final leaf length and maximum growth rate in the rht3wild type, but was without effect on the Rht3 mutant, whichis generally regarded as being non-responsive to gibberellin(GA). Paclobutrazol, an inhibitor of GA biosynthesis, decreasedfinal leaf length and maximum growth rate in the rht3 wild typeto values similar to those in the untreated Rht3 mutant. NeitherGA₃ nor paclobutrazol affected the duration of leaf growth.The decrease in leaf length was produced by reduction of celllength rather than cell number. The maximum relative elementalgrowth rate (REGR) for cell extension was essentially the samein all treatments, as was the time between the cells leavingthe meristem and achieving maximum extension rate. The differencesbetween the genotypes and treatments were all almost entirelydue to differences in the time taken from the attainment ofmaximum REGR of cell extension to the cessation of extension.This was reflected in the length of the extension zone, whichwas approximately 6–8 per cent of final leaf length. Theeffects of the Rht3 allele, GA₃ and paclobutrazol all appearto be on the processes which promote the cessation of cell elongation. Key words: Cell extension, gibberellin, leaf growth, Rht3 gene, Triticum, wheat     

Effects of Jasmonic Acid on Motor Cell Physiology in Mimosa pudica L. and Cassia fasciculata Michx

When applied to pulvini of Mimosa pudica, jasmonic acid (JA)affected neither proton fluxes nor the membrane potential ofthe motor cells. When added to leaflets of Cassia fasciculata,JA increased the rate of dark-induced pulvinar movements ina concentration-dependent manner. This effect was observed withinas little as 15 min after a 1-h treatment that preceded theinducing signal. Treatments in buffered media at acidic pH resultedin the greatest physiological responses. Light-induced pulvinarmovements were considerably reduced under the same conditions.With continuous illumination, JA induced a closing movementof the leaflets in a concentrationdependent manner. These resultsare discussed in relation to the ionic changes in the pulvinarmotor cells and in relation to results obtained previously upontreatment of Cassia plants with ABA. Although ABA and JA havesimilar physiological effects on the dark-induced closure, theydiffer in the type of response elicited by brief treatment andwith respect to light-induced opening. (Received September 27, 1993; Accepted January 15, 1994)     

An Analysis of Growth Rates in the Early Life of Apple Trees

A long-term continuing experiment to study the effect of rootstockand pruning on composite apple trees is described. Cox's OrangePippin was worked on to either M·26 or MM·104rootstocks. Two pruning regimes were applied to give a totalof four treatments. The course of whole plant growth expressed by the cumulativedry weight during the first 9 years is described by means ofgrowth curves. Special reference is made to the growth and functionof the perennial structure, and its contribution to whole plantgrowth is quantified by means of the parameter Unit ProductionRate. This expresses the rate of growth per unit weight of theperennating structure. Unit Production Rate declined with increasingtree age; in the first 5 years spur pruning and invigoratingrootstock treatments gave the highest values whilst later thissituation was reversed. The efficiency of the process of perennation is quantified interms of the production rate of the deciduous fraction per unitweight of the perennating structure. In this respect dwarfingrootstock treatments were appreciably superior to invigoratingrootstock treatments only after the fifth year. Spur pruninggave lower values than minimal pruning at 7 years with spur-prunedCox/MM·ro4 alone showing a decline with increasing ageafter 5 years.     

Stomatal responses to changing irradiance in Phaseolus vulgaris L.

The effects of air temperature (T_o), leaf-air vapour pressuredifferences [VPD) and water deficit on stomatal responses tochanging irradiance were studied in Phaseolus vulgaris L. Responseswere approximately sigmoidal, with rates of closure being fasterthan the rates of opening. The mean half-time for closure was5.4 min and for the opening 9.2 min. Under water deficit, bothstomatal opening and closing were faster than in well-wateredconditions. Stomata were more sensitive to VPD and water stressthan to T_o. The higher the VPD or T_o the more rapid was thestomatal response, except in stressed plants where there wasno significant effect of T_o. Under water stress, stomata weremore sensitive to water potential (     

Regrowth by Swards of Subterranean Clover after Defoliation. 1. Growth, Non-structural Carbohydrate and Nitrogen Content

Swards of subterranean clover (Trifolium subterraneum L.) atLAl 6 grown in N-free nutrient solution were subjected to threedefoliation treatments which removed 30, 70 and 80% of shootdry weight. Subsequent regrowth and changes in the concentrationsof carbohydrate and nitrogen in plant components were measuredat 0, 1, 5, 9 and 13 d after defoliation and compared with thosein uncut swards. The rate of shoot regrowth declined with increasing severilyof defoliation. In all defoliation treatments, growth was confinedto leaves for up to 5 d. Root growth ceased in all treatmentsfor a longer period. Reestablishment of the leaf area in severely-defoliatedswards was facilitated by the rapid opening of developing leavesand by changes in the allocation of carbon which favoured leafover branch and root, and lamina over petiole growth. Loss of carbohydrate and nitrogen from roots and branches lasting5–9 d was observed in the more severe defoliation treatments.Loss of protein (N x 6.25) exceeded that of total non-structuralcarbohydrate, and could have accounted for the nitrogen contentof new leaf during this period. Branches lost 62% of their initialcarbohydrate content compared with 25% from roots in the 80%cut swards. In contrast, roots, by virtue of their greater mass,were the principle source of mobilized nitrogen. Nitrogen accumulationceased in 80% cut swards for 9 d. However, carbohydrate levelsin the crown nodules were not severely depleted. It was concluded that partitioning of growth to lamina and themobilization of carbohydrates and nitrogen were important forrecovery from defoliation. Carbohydrates, carbon partitioning, defoliation, nitrogen, mobilization, regrowth, subterranean clover, Trifolium subterraneum L     

Tentoxin Suppresses Stomatal Opening by Inhibiting Photophosphorylation

Tentoxin and, to a lesser extent, dihydrotentoxin (both at 10mmol m^–3) reduce stomatal opening in epidermal stripsof Commelina communis in the light but not in darkness. Thiseffect was significantly greater in normal air than in CO₂-freeair. Fusicoccin overcame the tentoxin effect. However, tentoxindid not inhibit stomatal opening in the light in epidermal stripsof Paphiopedilum harrisianum, a species which lacks guard cellchloroplasts. It is concluded that tentoxin exerts its actionon stomata not by an ionophorous effect in the plasmalemma ofguard cells but by the inhibition of photophosphorylation intheir chloroplasts. The effects of DCMU and tentoxin on guardcells are discussed in terms of their effects on chloroplastsand the extent to which energy is supplied from this organelleduring stomatal opening in the light. The results indicate thatneither photophosphorylation nor non-cyclic electron transportin guard cell chloroplasts are essential for stomatal opening. Key words: Commelina, epidermal strips, Paphiopedilum, photophosphorylation, stomata, tentoxin     

Defoliation and its Effects on Pod and Seed Development in Oil Seed Rape (Brassica napus L.)

Both in vivo and in vitro techniques have been used to followthe development of individual pods on the terminal inflorescenceof undefoliated and defoliated plants of oil-seed rape (Brassicanapus cv. Maris Haplona). For any pod, a rapid increase in podlength occurred between 2 d and 8 d after flower opening andthis preceded by approximately 2 d the increase in pod width,the rate of which was less than that for length. An increasein the diameter of individual seeds coincided with the increasein pod width. Regional increases in the length and width ofpods were associated with the presence of developing seeds inthese regions. Key words: Brassica napus L., Development, In vivo, Pod and seed, Stress     

Studies in Stomatal Behaviour: X. AN INVESTIGATION OF RESPONSES TO LOW-INTENSITY ILLUMINATION AND TEMPERATURE IN XANTHIUM PENNSYLVANICUM

Two experiments are described in which stomatal sensitivityto low-intensity white light was studied for Xanthium pennsylvanicumWall. In the first experiment a daylength extension for 7, 9, or 15hrs. was given using 10, 40, or 160 lux to shorten a basic 16-hr.night, which was also given at its full length as a tenth treatment.Measurements were made of stomatal opening ability on the morningfollowing the different treatments. With a 15-hr. extensionthere was at all intensities a significant response, shown bya reduced rate of opening in the morning. With a 9-hr. extensionusing 40 or 160 lux, opening ability was reduced, but 9 hrs.of 10 lux was insufficinet to produce a detectable effect. The7-hr. extension was ineffective at all three intensities. In the second experiment stomatal behaviour was observed during20 hrs. of either darkness or 10 lux at four temperatures (15,22, 29, and 36°C.). During 20 hrs. of darkness there wasnight opening at all temperatures, but at lower temperaturesit began sooner and lasted longer. These responses to temperaturedid not fit a simple linear relationship, there being a significantcubic term revealed by non-linear regression analysis. Thiscould be explained if the response was considered in terms ofthe magnitude of the change in temperature (from 25°C.)at the beginning of the experiment; there appeared to be sometemperature compensation over a limited range. in 10 lux, nightopening was suppressed at 29° and 36°, but at 15°it was apparently unaffected by the light; at 22° it wasnot completely suppressed by 10 lux but the time of its occurrencewas delayed. Effects of light and temperature are discussed in relation toan endogenous rhythm in darkness which was previoulsy shownto operate in Xanthium pennsylvanicum (Part IX). It is considered that to explain effects of very low intensitylight it will be necessary to recognize a ‘low intensityresponse’ by stomata, which does not operate via changesin guard-cell carbon dioxide.     

Responses of Stomata to IAA and Fusicoccin at the Opposite Phases of an Entrained Rhythm

The stomata of Commelma communis showed reduced opening responsesto light and low CO₂ concentrations during the night phase oftheir entrained circadian rhythm. Increased supplies of potassiumions, and treatments with indol-3-ylacetic acid and fusicoccin,failed to promote opening during the night phase to a levelequivalent to that in the day phase. The inability of fusiccocinto overcome the suppression of opening during the night phasecontrasts with its ability to counteract the closure inducedby agents such as CO₂, darkness and abscisic acid. It is concludedthat there are at least two basic mechanisms by which the turgorof guard cells can be regulated, one which is susceptible tooverriding control by fusicoccin and another which is unaffectedby fusicoccin. Several previous studies had shown a positive correlation betweenmalate in the epidermis (mainly located in guard cells) andstomatal opening. In the present experiments the aperture/malatecorrelation was broken in epidermis treated with fusicoccinduring the night phase of the rhythm. The amount of malate presentexceeded that associated with the same stomatal aperture inthe day phase. Possible explanations are (1) that fusicoccinstimulates similar proton fluxes out of the guard cells duringboth phases of the rhythm, but an unknown factor imposes a restrictionon stomatal opening during the night phase; (2) that there arelower proton fluxes in the night phase (limited, for example,by a reduced supply of ATP) but chloride availability or transportis reduced to an even greater extent so that a larger productionof malate in the guard cells is required. Key words: Stomata, IAA, Fusicoccin, Rhythms     

Studies in Stomatal Behaviour: IX. PHOTOPERIODIC EFFECTS ON RHYTHMIC PHENOMENA IN XANTHIUM PENNSYLVANICUM

An investigation of the effects of different day-length treatmentson stomatal behaviour in Xanthium pennsylvanicum Wall. has shownthat there are differences in long-and short-day treatmentssimilar to those first reported by Schwabe (1952) for Chrysanthemumand Kalanchoe, viz. stomatal opening towards the end of thenight in short days, but not in long days (short night precededby a period of low intensity illumination). In Xanthium therewere in addition very marked differences in rates of stomatalopening in the morning after different lengths of night. Theseeffects were not persistent, there being an immediate reversalupon a change from long- to shrot-day treatment, or vice versa. Further investigation showed that there was an endogenous rhythmaffecting the stomata in continuous darkness; rate of openingwas slow after very short nights, but became greater with extensionof the night to 14–16 hrs., this being the first ‘peak’of an ‘opening ability’ rhythm. The rhythm diedout rapidly and the second cycle was much reduced in amplitude.The period of the rhythm appeared to be approximately 24 hrs.The phase was set mainly by the time of onset of darkness, butthe duration of the pretreatment with low intensity illuminationwas also important—prolonging this was found to reudcethe time in darkness before the first peak. Each hour of lightof 1,500 lux given before darkness was found to be equivalentot approximately 0.3 hr. to darkness. The predominant effect of the length of the preceding nightwas on the slope of the opening curves rather than on the timeof onset of opening following illumination. The first peak of the opening ability rhythm was often manifestedeven in continuous darkness by a period of ‘night opening’of the stomata. The results are discussed and compared with those of other authorsand attempts are made to relate them to theories of the stomatalmechanism.     

Microanalytical Studies of Potassium and Chloride Fluxes and Stomatal Movements of Two Species: Vicia faba and Pelargonium x hortorum

Potassium and chloride exchange kinetics in guard cells werefound to be related to stomatal opening and closing kineticsin Pelargonium x hortorum and Vicia faba by simultaneously measuringion contents and degrees of opening with an electron probe anda hydrogen diffusion porometer. Large shifts between both kinetics occur during opening andclosing. These results show that maximal potassium content isdelayed, which seems to question the primary role of potassiumin stomatal opening     

Inhibition of Stomatal Opening by Analogues of Abscisic Acid

Twenty analogues of abscisic acid have been tested for theiractivity as inhibitors of stomatal opening in isolated epidermisof Commelina communis. A number of derivatives showed slightactivity but only two treatments resulted in significant stomatalclosure and this was accompanied by destruction of the guardcell membranes. Such damage is characteristic of the stomatalresponse to farnesol, another sesquiterpenoid also thought tobe involved in control of water loss. The implication of theseresults in the study of antitranspirants is considered.     

The Spontaneous Formation of Antheridia in Onoclea is not Blocked by Light

Onoclea sensibilis gametophytes were grown from spores on ashedsoil and agar to determine if the spontaneous formation of antheridiacan be blocked by light. Under most conditions, dark-grown gametophytesformed antheridia later than or at the same time as gametophytesgrown in the light. Under no circumstances was there a rapidonset of maleness in the dark. These results contradict thehypothesis that, in Onoclea, antheridiogen is required to inducemaleness because light inhibits the formation of antheridia.In the light, antheridia formed on heart-shaped thalli. In darkness,antheridia formed on filamentous gametophytes. The timing ofonset of maleness was affected by temperature and the presenceof sucrose. The effect of sucrose on the comparison betweenlight and dark treatments depended on both substrate and temperature Onoclea sensibilis, L., sensitive fern, fern gametophytes, sexuality, light-induced block     

Induction of Near-vessellessness in Ephedra campylopoda C. A. Mey.

Near-vessellessness was induced in the secondary xylem of Ephedracampylopoda C. A. Mey. by mechanical bark blocking or by wounding.Both treatments resulted in regions of near-vesselless xylem.Xylem formed after the mechanical bark blocking also had regionsin which the orientation of the axial components was changedfrom axial to lateral. Since either mechanical arrest of phloemand cambial transport or wounding of the cambium almost stoppeddifferentiation into vessels, and instead induced differentiationinto tracheids, it seems that the developmental signal for tracheiddifferentiation is not the same as that for vessels. The possibleregulation of near-vessellessness in Ephedra is discussed.Copyright1994, 1999 Academic Press Differentiation, Ephedra campylopoda, near-vessellessness, wood formation, xylem     

An Empirical Model for Leaf Expansion in Cacao in Relation to Plant Water Deficit

Cacao (Theobroma cacao L.) seedlings at the onset of a flushcycle were exposed to five different irrigation treatments.The expansion of all leaves in an emerging flush was followed,and estimates of leaf water potential(_w) were made on each daythat leaf areas were measured. The growth in area of the leaveswas fitted with a modified logistic curve of the form y = a/[1+ be ^–(ct+dt2)], and parameters of leaf growth were derivedfrom the fitted constants. A coefficient of stress exposure,S, was derived as the slope of the relation between cumulative_w and time. Three parameters of the logisitc function (a, cand d) were strongly associated with S. By fitting regressionsfor the relations between S and estimates of each parameterfor the five treatments, values of a, b, c and d may be estimatedat any level of S encompassed by these data. The effects ofwater stress on leaf expansion rate and on the final leaf areaattained by the emerging flush can be adequately predicted bythis technique. Theobroma cacao L.cacao, leaf expansion, logistic model, water stress     

Changes in Anthocyanin, Carotenoid, Chlorophyll, and Protein in Developing Florets of the Chrysanthemum

Florets of a purple cultivar (Fandango) of the horticulturalchrysanthemum (Chrysanthemum morifolium Ramat) were removedfrom flower heads at seven stages of opening (from unopenedbud to dying flower) and segregated into different lengths,each of which was analysed. Wet weight per floret increased from 0.25–1 mg in thebud to about 3 mg (tubular florets) or to 20–40 mg (rayflorets) in the fully open flower. Protein decreased from 6per cent of the wet weight in the bud to about 2 per cent inthe open flower. In the ray florets anthocyanin concentrationreached a maximum in the half-open flower and then decreasedsharply, whereas carotenoid and chlorophyll declined continuouslyfrom the bud stage. Almost no anthocyanin was formed by thetubular florets and chlorophyll declined as in the ray florets,but carotenoid concentration increased to a maximum in the half-openflower and then decreased. In another cultivar (Light Bronze Fandango) the content of anthocyaninwas lower and that of carotenoid higher but similar changesin pigment levels were observed except that carotenoid roseto a maximum in the ray florets. In two other cultivars, Redand Cerise Fandango, the anthocyanin content was the same asin Fandango but the carotenoid concentration was the same orabout half that in Light Bronze Fandango respectively.