Light-dependent fluorescence variations in intact leaves

Transient variations in the fluorescence from intact Phytolaccaamericana leaves after the onset of illumination were measuredunder various light and dark conditions. Dark-adapted leaveswhen illuminated with strong light underwent an intensity variationwith a peak; the fluorescence intensity reaching its peak severalseconds after the onset of illumination then decreasing to asteady level. The peak height relative to the steady level increasedwith the increasing intensity of actinic light. Pre-illuminationof the dark-adapted leaves with strong light caused a markedlowering of the peak. About 20 min of dark incubation was requiredfor the light-adapted leaves to return to the dark-adapted state.All of the action spectra, for the peak, the steady level andthe effect of light in post-illumination to inhibit recoveryto the dark state, showed high bands due to chlorophyll b andcarotenoid absorption and low bands due to chlorophyll a absorption.We concluded that the light absorbed by photosystem 2 is responsiblefor these phenomena. (Received April 21, 1975; )     

Effects of 5-Aminolevulinic Acid on the Accumulation of Chlorophyll b and Apoproteins of the Light-Harvesting Chlorophyll a/b-Protein Complex of Photosystem II

The effects were examined of 5-aminolevulinic acid (ALA) onthe accumulation of Chl and apoproteins of light-harvestingChl a/b-protein complex of photosystem II (LHCII) in cucumbercotyledons under intermittent light. A supply of ALA preferentiallyincreased the accumulation of Chl a during intermittent illumination.However, when cotyledons were pretreated with a brief exposureto light or benzyladenine (BA), the stimulatory effect of ALAon the increase in the level of Chl b was greater than thatin the level of Chl a, resulting in decreased ratios of Chla/b. Time-course experiments with preilluminated cotyledonsrevealed that LHCII apoproteins accumulated rapidly within thefirst 30 min of intermittent illumination with a decline duringsubsequent incubation in darkness. A supply of ALA did not affectthe accumulation of LHCII apoproteins during the intermittentlight period, but it efficiently inhibited the decline in theirlevels during the subsequent darkness. After exposure to a singlepulse of light of BA-treated cotyledons, the prompt increasein levels of LHCII apoproteins was not accompanied by the formationof Ch b, which began to accumulate later. The pattern of changesin levels of LHCII apoproteins was quite similar to that inlevels of Chl a. These results suggest that LHCII apoproteinsare first stabilized by binding with Chl a and that an increasedsupply of Chl a and the accumulation of LHCII apoproteins areprerequisites for the formation of Chl b. ¹Present address: Department of Chemistry, Faculty of Scienceand Technology, Meijo University, Aichi, 468 Japan.     

Active oxygen participation in chlorophyll destruction and lipid peroxidation in SO2-fumigated leaves of spinach

Chlorophyll a and carotenoids of spinach began to be destroyed2 to 3 hr after fumigation with 2 ppm SO₂ under light, whereaschlorophyll b was undamaged during 8 hr of exposure to SO₂.Pheophytin a was not affected by the fumigation. When disks excised from leaves fumigated with SO₂ at 2 ppm for2 hr were illuminated, chlorophyll a and carotenoids were brokendown, while they were not destroyed in darkness. The destructionof these pigments was suppressed under nitrogen. Chlorophylla destruction was inhibited by l,2-dihydroxybenzene-3,5-disulfonate(tiron), hydro-quinone and ascorbate, but not by l,4-diazabicyclo-[2,2,2]-octane(DABCO), methio-nine, histidine, benzoate and formate. Chlorophylla destruction was inhibited by phenazine methosulfate but stimulatedby methyl viologen. Addition of superoxide dismutase (SOD) tothe homogenate of SO₂-fumigated leaves inhibited the chlorophylla destruction. The activity of endogenous SOD was reduced to40% by 2-hr fumigation before the loss of chlorophyll was observed.These results suggest that chlorophyll a destruction by SO₂was due to superoxide radicals (O₂^–). Moreover, malondialdehyde (MDA), a product of lipid peroxidation,was formed in SO₂-fumigated leaves. MDA formation was inhibitedby tiron, hydroquinone and DABCO but not by benzoate and formate.MDA formation was increased by D₂O. These results suggest thatlipid peroxidation in SO₂-fumigated leaves was due to singletoxygen ¹O₂ produced from O_2^–. (Received May 15, 1980; )     

Active oxygen participation in chlorophyll destruction and lipid peroxidation in SO2-fumigated leaves of spinach

Chlorophyll a and carotenoids of spinach began to be destroyed2 to 3 hr after fumigation with 2 ppm SO₂ under light, whereaschlorophyll b was undamaged during 8 hr of exposure to SO₂.Pheophytin a was not affected by the fumigation. When disks excised from leaves fumigated with SO₂ at 2 ppm for2 hr were illuminated, chlorophyll a and carotenoids were brokendown, while they were not destroyed in darkness. The destructionof these pigments was suppressed under nitrogen. Chlorophylla destruction was inhibited by l,2-dihydroxybenzene-3,5-disulfonate(tiron), hydro-quinone and ascorbate, but not by l,4-diazabicyclo-[2,2,2]-octane(DABCO), methio-nine, histidine, benzoate and formate. Chlorophylla destruction was inhibited by phenazine methosulfate but stimulatedby methyl viologen. Addition of superoxide dismutase (SOD) tothe homogenate of SO₂-fumigated leaves inhibited the chlorophylla destruction. The activity of endogenous SOD was reduced to40% by 2-hr fumigation before the loss of chlorophyll was observed.These results suggest that chlorophyll a destruction by SO₂was due to superoxide radicals (O₂^–). Moreover, malondialdehyde (MDA), a product of lipid peroxidation,was formed in SO₂-fumigated leaves. MDA formation was inhibitedby tiron, hydroquinone and DABCO but not by benzoate and formate.MDA formation was increased by D₂O. These results suggest thatlipid peroxidation in SO₂-fumigated leaves was due to singletoxygen ¹O₂ produced from O_2^–. (Received May 15, 1980; )     

Enzymatic Conversion of Pheophorbide a to the Precursor of Pyropheophorbide a in Leaves of Chenopodium album

Soluble proteins extracted from leaves of Chenopodium albumcatalyzed the conversion of pheophorbide a to a precursor ofpyropheophorbide a, putatively identified as C-13²-carboxyl-pyropheophorbidea. The precursor was then decarboxylated non-enzymatically toyield pyropheophorbide a. Soluble proteins and pheophorbidea, as the substrate, were required for the formation of theprecursor, and boiled proteins were enzymatically inactive.The maximum rate of conversion of pheophorbide a to the precursoroccurred at pH 7.5. The K_m for pheophorbide a was 12.5 µMat pH 7.0. Both pheophorbide b and bacteriopheophorbide a couldserve as substrates, but protopheophorbide a could not. Formationof methanol was detected during the enzymatic reaction, an indicationthat the enzyme is an esterase. Among seven alcohol analogstested, only methanol inhibited the enzymatic activity uncompetitively,with a K₁ of 71.6 mM. Mass-spectrometric (MS) analysis of theprecursor yield a peak at m/z 579 that indicated the releaseof a methyl group from pheophorbide a. It appears thereforethat the enzyme catalyzes the demethylation of the carbomethoxygroup at C-13² of pheophorbide a by hydrolysis to yield methanoland the precursor, C-13²-carboxyl-pyropheophorbide a, whichis converted to pyropheophorbide a by spontaneous decarboxylation.We have tentatively designated the enzyme "pheophorbidase".The presence of the enzyme was dependent on plant species andit was expressed constitutively. ¹Present address: Faculty of Science, Shizuoka University, Ohya,Shizuoka, 422 Japan     

Growth of grana from "primary" thylakoids in Phaseolus vulgaris

The plastids of young dark-grown bean leaves, exposed to periodiclight are agranal, devoid of chlorophyll b and contain primarythylakoids and chlorophyll a. Transfer of these plants to continuousillumination results in synthesis of new chlorophyll a, chlorophyllb and grana. This study was done in order to study whether andhow the grana are formed from preexisting primary thylakoids.¹⁴C--aminolevulinic acid was used to label the chlorophyll aof the primary thylakoids, and its fate was studied after transferof the plants to continuous light. It was found that chlorophyll b and grana become ¹⁴C-labelled.The total radioactivity of chlorophyll b per bean increasedwith the parallel decrease of that of chlorophyll a. All subchloroplastfractions, obtained after digitonin disruption of chloroplasts,contained chlorophyll a of equal specific radioactivity. Thespecific radioactivity of chlorophyll b was lower than thatof chlorophyll a, and, in addition, it was lower in the granathan in the stroma lamellae fraction. The data suggest that chlorophyll b is formed from chlorophylla; the grana are formed by stacking of preexisting primary thylakoids;chlorophyll b is synthesized faster in the grana than the stromalamellae; the newly formed chlorophyll a molecules are distributedat random throughout the developing photosynthetic membraneand not on specific growing sites. (Received April 24, 1976; )     

Appearance of Chlorophyll-Protein Complexes in Greening Barley Seedlings

The sequential appearance of chlorophyll-protein complexes (CP)in greening barley leaves was studied by an improved methodof SDS-polyacrylamide gel electrophoresis (PAGE). Solubilizedthylakoid membranes were purified using a sucrose step gradientand CPs were separated by PAGE with low concentrations of SDSin solubilizing and reservoir buffers. At 10 min after the onsetof illumination, a chlorophyll-protein complex (CPX) was detected.It was a labile CP, its chlorophyll (Chl) being easily releasedfrom the apoprotein during electrophoresis. The P700-chlorophylla/b-protein complex (CPl) appeared after 45–60 min ofillumination together with P700 activity. Light-harvesting chlorophylla/b-protein complex (LHCP) began to accumulate at 2.5 h withthe beginning of Chl b synthesis. In some cases a small amountof CPa could be detected after 6 h of greening. The time-differencespectrum between homogenates of leaves illuminated for 30 and60 min had an absorbance maximum at 677 nm, showing that a redshift indicative of CPl formation began soon after completionof the Shibata shift. The time-difference spectrum between 3.5-hand 4.0-h illuminated leaves resembled the absolute spectrumof fully greened leaves, indicating that at this stage, spectralcomponents were being synthesized at the same ratio at whichthey exist in fully greened tissues. Both absolute and time-differencespectral data supported the SDS-PAGE results. (Received February 27, 1985; Accepted May 8, 1985)     

Ultrastructural Changes in Chloroplasts of Quercus rotundifolia Lam. in Response to Evernic Acid

The amount of total chlorophyll, chlorophylls a and b as wellas the ratio of a to b decreased in chloroplasts isolated fromQuercus rotundifolia leaves, kept for 17 d in a solution of35.5 µM evernic acid in 1 mM Na HCO₃, when compared withthe chloroplasts of control leaves (kept in NaHCO₃). The chloroplastsin the spongy parenchyma were smaller and the amount of starchand plastoglobuli lower. The number of grana per chloroplastsection, the number of thylakoids per grana and the height ofgrana stacks were also less in the chloroplasts of leaves treatedwith evernic acid. Quantitative ultrastructural differenceswere determined by means of electron microscopy and image analysistechniques. Quercus rotundifolia Lam., chloroplasts, ultrastructure, lichens, evernic acid     

Formation of Chlorophyll-Protein Complexes during Greening 1. Distribution of Newly Synthesized Chlorophyll among Apoproteins

The relationship between the accumulation of Chl and the apoproteinsof the light-harvesting Chl a/b-protein complex of PS II (LHCII)during the greening of cucumber cotyledons was studied. LHCIIapoproteins were not detected in etiolated cotyledons. Uponillumination, Chl a was formed as a result of photoconversionof protochlorophyllide (Pchlide) which had accumulated in thedark. During the lag period that preceded the accumulation ofChl, a small amount of LHCII apoproteins appeared. The amountof LHCII apoproteins increased with increases in levels of Chlb, though somewhat more rapidly during the first 10 h of greening.Treatment with benzyladenine (BA) or levulinic acid (LA) wasused to vary the supply of Chl a for apoproteins by promotingor inhibiting the synthesis of Chl a, respectively. LA decreasedbut BA increased the rate of accumulation of Chl b and LHCIIapoproteins. Only small amounts of Chl b and LHCII apoproteinswere formed under intermittent illumination. However, in thepresence of chloramphenicol (CAP), which inhibits the synthesisof plastome-coded proteins including apoproteins of the P700-Chla-protein complex (CP1) and a Chl a-protein complex of PS II(CPa), we observed the accumulation of Chl b and LHCII apoproteins,both of which are of nuclear origin. During incubation in thedark after intermittent exposure to light, CAP alone allowedneither destruction nor accumulation of Chl b and LHCII apoproteins,but it did enhance the effect of CaCl₂ in inducing both Chlb and these apoproteins. These results can be explained by assumingthat apoproteins of CP1 and CPa have a higher affinity for Chla than do LHCII apoproteins. When the availability of Chl ais limited, these apoproteins compete with one another for Chla, with the resultant preferential formation of CP1 and CPa.However, when the supply of Chl a becomes large enough for saturationof apoproteins of CP1 and CPa, some of the Chl a is incorporatedinto LHCII apoproteins either directly or after conversion toChl b. Thus, the formation of different Chl-protein complexes(CPs) is regulated by the relative rates of synthesis of Chla and apoproteins and by differential affinities of the apoproteinsfor Chl a. ⁴Present address: Kyowa Hakko Co., Ltd., 4041, Ami-machi, Inashiki,Ibaraki, 300-03 Japan (Received September 14, 1989; Accepted April 26, 1990)     

Determination and Differentiation of Leaf and Petal Primordia in Impatiens balsamina

The development of primordia as leaves, petals, or as organsintermediate between leaves and petals can be regulated by photoperiodin Impatiens. In intermediate organs only some parts of theorgan differentiated as petal, and then only in some cell layers.Allometric measurements of primordium shape suggested that intermediateorgans may begin development as petals, and that their intermediatecharacter at maturity resulted from a switch of some parts ofthe organs from petal to leaf development when the primordiawere between 0.5 and 1 mm long. In reverted apices made to re-flower,primordia were not completely determined as leaves until theywere about 750 µm long. Determination typically occurredfirst at the tips and last at the bases of these primordia.The determination of primordia as leaves or petals in Impatiensis discussed in relation to primordium determination in otherspecies. It is suggested that the lack of commitment to flowermay result in relatively late primordium determination in Impatiens. Impatiens balsamina, determination, differentiation, leaf and petal development, flowering, reversion     

Estimation of the sensitivity to photoinhibition in Striga hermonthica-infected sorghum

Sensitivity to photoinhibition was assessed in sorghum infectedwith the angiosperm root parasite Striga her-monthica and inuninfected sorghum plants, at four times during the developmentof the host-parasite association. Photoinhibition was inducedby exposing either leaf discs or intact leaves to a photosyntheticphoton flux density of 2000 µmol m^–2 s^–1 for4 h. The inhibition of apparent quantum yield (_a) and photosynthesisin high light (A₁₅₀₀) were assessed in leaf discs using an oxygenelectrode and the recovery of these from photoinhibition wasfollowed in intact leaves using an infra-red gas analyser. Fromsoon after attachment of the parasite, infected sorghum plantshad a lower A₁₅₀₀. During the period when Striga induced a loweringof A₁₅₀₀, _a was more sensitive to photoinhibition in Striga-infectedplants. However, at the same time, the high-light-induced inhibitionof A₁₅₀₀ was similar in Striga-infected and uninfected plants.Recovery of both _a and A₁₅₀₀ was incomplete after 6 h and thetime-course of recovery was similar in Striga-infected and uninfectedplants. The results indicate that Striga-infected plants weremore sensitive to photoinhibition and that photoinhibition wasprimarily due to damage to electron transport/photo-phosphorylationand not disablement of the recovery processes. Key words: Photoinhibition, quantum yield, recovery from photoinhibition, parasitic plants     

Regulation of the Activity of Ribulose-1,5-Bisphosphate Carboxylase in Response to Changes in the Photosynthetic Source-Sink Balance in Intact Soybean Plants

Sink-limited conditions cause a reduction in the rate of photosyntheticfixation of CO₂ in single-rooted soybean leaves (Glycine max.Merr.). We suggested previously that this reduction is due tothe deactivation of ribulose-1,5-bisphosphate carboxylase (RuBPcase;EC 4.1.1.39 [EC] ) that is caused by a decrease in the level of freeP_i via a decrease in the rate of conversion of phosphorylatedintermediates to the end-product (sucrose) in sink-limited leaves[Sawada et al. (1989) Plant Cell Physiol. 30: 691, Sawada etal. (1990) Plant Cell Physiol. 31: 697, Sawada et al. (1992)Plant Cell Physiol. 33: 943]. In the present study, we investigatedwhether, in intact soybean plants, sink-limited conditions wouldalso cause a reduction in the rate of photosynthesis and whethersuch a reduction might be due to the deactivation of RuBPcasevia the same regulatory mechanism as that suggested from previousstudies with single-rooted leaves. Continuous removal of flowerbuds from intact plants brought a large decrease in ratio ofthe dry weight of sink organs (stem, roots, pods) to sourceorgan (leaves) as a result of the absence of pod formation.Pods are likely to function as the major sink at the reproductivestage. Upon continuous removal of flower buds, the treated (sink-limited)plants showed a large decrease in the rate of photosyntheticfixation of CO₂ as compared to control plants. RuBPcase in theleaves of treated plants was continuously inactivated with thedecrease in photosynthetic activity. However, the inactivatedenzyme was totally reactivated upon incubation in the presenceof 10 mM NaHCO₃ and 5 mM MgCl₂. The levels of sucrose and ribulose-1,5-bisphosphatein leaves of the treated plants increased significantly. Allthese results coincide exactly with those obtained in previousstudies of single-rooted leaves under the sink-limited conditions. (Received July 14, 1994; Accepted February 21, 1995)     

Growth and Dry-weight Distribution in Callistephus chinensis as Influenced by Lighting Treatment

Plants of two cultivars of Callistephus chinensis (Queen ofthe Market and Johannistag) were grown in 8 h of daylight perday with one of the following treatments given during the 16h dark period: (a) darkness—‘uninterrupted night’,(b) I h of light in the middle of the dark period—a ‘nightbreak’, (c) I min of light in every hour of the dark period—‘cycliclighting’, (d) light throughout—‘continuouslight’. The plants receiving uninterrupted dark periods remained compactand rosetted in habit with small leaves, while leaf expansion,stem extension, and flower initiation were promoted in all threeillumination treatments (b, c, d). Although these three treatmentsproduced similar increases in leaf area, continuous light wasthe most effective for the promotion of both stem growth andflower initiation while cyclic lighting was generally more effectivethan a I-h night break. Continuous light also caused more dry matter to be divertedto stems at any given vegetative dry weight and it was shownthat the stem weight ratio of both varieties was correlatedwith stem length.     

Transpiration Stream of Desert Species: Resistances and Capacitances for a C3, a C4, and a CAM Plant

Transpiration rates and water potentials of three sympatricdesert perennials, a C3 subshrub (Encelia farinosa), a C₄ bunchgrass(Hilaria rigida), and a CAM succulent (Agave deserti), wereanalysed using an electrical circuit analogue that includedresistances and capacitances for the leaves, stems, and roots.The water storage capability of the organs differed considerably,capacitance ranging over 1000-fold from the thin leaves of H,rigida to the massive leaves of A. deserti, although the capacitanceper unit volume varied only 1.9-fold. The diurnal changes inwater storage could support maximum transpiration rates of H.rigida for 4 min, E. farinosa for 7 min, and A. deserti for16 h. The time constant for equilibration of water from storageto the xylem ranged from 29 s for roots of H. rigida to 52 minfor leaves of A. deserti. Resistances for such movement wererelatively low for the succulent leaves of A. deserti and wereup to about 50-fold higher for the three organs of E. farinosa.Xylem resistances calculated using the Hagen-Poiseuille lawand measured xylem dimensions were 2.1- to 2.1-fold lower thanresistances estimated from observed water potential drops, adiscrepancy which is in agreement with other published data.Contrary to data on other plants, the xylem resistances in theroots and leaves of E. farinosa and H. rigida averaged only15% of the stem xylem resistance. Key words: Capacitance, Xylem resistance, Transpiration stream, Desert     

Energy-dependent redox state of heme a+a3 and copper of cytochrome oxidase in perfused rat brain in situ

Using theblood-free perfused rat brain, we examined the redox behavior ofcytochrome oxidase of two chromophores, hemea + a₃ and copper.When perfusate inflow was stopped to induce global ischemia,the reduction of heme a + a₃ was triphasic,with a rapid phase, a slow phase, and a second rapid phase. Incontrast, the reduction of copper was monophasic after the rapid phaseof heme a + a₃. The triphasicreduction of heme a + a₃ was diminished by energy-depleting treatments, such as addition of an uncoupler. Thetime course of the reduction of copper was not affected by the energydepletion. During global ischemia the decrease in creatine phosphate nearly paralleled the reduction of hemea + a₃, whereas ATPremained at the control level until ~60% of hemea + a₃ was reduced inthe rapid phase. In the slow phase, ATP started to decrease with thereduction of copper. The redox behavior of copper was similar to theslow phase of the reduction of heme a + a₃ because ofthe higher oxygen affinity of copper than of hemea + a₃. Therefore,the rapid phase of the reduction of hemea + a₃ can be used asan alarm before a decrease in ATP, whereas the reduction of copperindicates a decrease in ATP under severe hypoxia. Thus the coppersignal in noninvasive near-infrared spectroscopy is a useful parameterfor the clinical setting.

     

Participation of Inorganic Orthophosphate in Regulation of the Ribulose-1,5-bisphosphate Carboxylase Activity in Response to Changes in the Photosynthetic Source-Sink Balance

Sink-limited conditions, defined as treatment with continuousillumination, cause a reduction in the rate of photosyntheticfixation of CO₂ in single-rooted leaves of soybean (Glycinemax. Merr. cv. Turunoko). We suggested previously that thisreduction is due to a deactivation of ribulose-1,5-bisphosphatecarboxylase (RuBPcase, E.C. 4.1.1.39 [EC] ) that is caused by a decreasein the level of P_i in the leaves [Sawada et al. (1989) PlantCell Physiol. 30: 691, Sawada et al. (1990) Plant Cell Physiol.31: 697]. In the present study, the mechanism of regulationof RuBPcase activity by P_i was examined. The activity of RuBPcasein the sink-limited leaves, exposed for 6 or 7 d to continuousillumination to alter the source/sink balance, was enhancedwith increasing concentrations of P_i, in a CO₂-free preincubationmedium in the presence of 5 mM MgCl₂ The maximum value [6.3µmole CO₂ (mg Chl)^–1 min^–1] was obtained atapproximately 5 mM P_i after a 5 min incubation, being 3 timesof the activity without the preincubation. The activity of acrude preparation of RuBPcase that had been deactivated by removalof CO₂ and Mg²⁺ ions by the gel filtration was 5.2–9.3nmole CO₂ (mg protein)^–1 min^–1 and was also enhancedby P_i plus Mg²⁺ ions. The maximum value [147–151 nmoleCO₂ (mg protein)^–1 min^–1] was attained at 5 mM P_iafter a 5 min incubation. The cycle of activation and inactivationof deactivated crude RuBPcase was perfectly reversible by additionof P_i to the enzyme and removal of P_i from the enzyme. Levelsof free P_i and of esterified phosphate in the sink-limited leaveswere 69% and 31% of the total phosphate, respectively. By contrast,in the control leaves, these values were 87% and 13%, respectively.These results support our previously stated hypothesis and indicatean important role for free P_i in the regulation of RuBPcaseactivity, in particular in sink-limited plants. (Received February 21, 1992; Accepted July 23, 1992)     

Studies on chlorophyllase of Chlorella protothecoides III. Purification and catalytic properties

Chlorophyllase was extracted from green cells of Chlorella protothecoidesby n-butanol treatment and purified 600-fold, as measured byenzyme activity in chlorophyll a hydrolysis, by ammonium sulfateprecipitation, chromatography on TEAE-cellulose column and gelfiltration with Sephadex G-200. At each purification step the following activities were compared:hydrolyses of chlorophyll a and methyl chlorophyllide a, methanolysisof chlorophyll a and transphythylation of methyl chlorophyllidea to chlorophyll a. The ratio of activities of chlorophyll a hydrolysis to chlorophylla methanolysis changed on purification and partial inactivationby heat, PCMB and phytol, as well as by varying the reactiontemperature, thus suggesting that the two reactions are notcatalyzed by a single enzyme. In contrast, the activity ratio of chlorophyll a methanolysisto transphytylation of methyl chlorophyllide a remained unaltered,indicating that these reactions can be forward and backwardreactions catalyzed by one enzyme. Results of kinetic studies also indicated that the chlorophyllaseof Chlorella protothecoides consists of at least two enzymes.One enzyme catalyzes chlorophyll a hydrolysis and the other,chlorophyll a methanolysis and the reverse reaction, transphytylationof methyl chlorophyllide a. (Received May 24, 1973; )     

Positive and Negative Messages from Roots Induce Foliar Desiccation and Stomatal Closure in Flooded Pea Plants

Flooding the soil for 5–7 d caused partial desiccationin leaves of pea plants (Pisum sativum. L. cv. ‘Sprite’).The injury was associated with anaerobiosis in the soil, a largeincrease in the permeability of leaf tissue to electrolytesand other substances, a low leaf water content and an increasedwater saturation deficit (WSD). Desiccating leaves also lackedthe capacity to rehydrate in humid atmospheres, a disabilityexpressed as a water resaturation deficit (WRSD). This irreversibleinjury was preceded during the first 4–5 d of floodingby closure of stomata within 24 h, decreased transpiration,an unusually large leaf water content and small WSD. Leaf waterpotentials were higher than those in well-drained controls.Also, there was no appreciable WRSD. Leaflets detached fromflooded plants during this early phase retained their watermore effectively than those from controls when left exposedto the atmosphere for 5 min. Stomatal closure and the associated increase in leaf hydrationcould be simulated by excising leaves and incubating them withtheir petioles in open vials of water. Thus, such changes inflooded plants possibly represented a response to a deficiencyin the supply of substances that would usually be transportedfrom roots to leaves in healthy plants (negative message). Ionleakage and the associated loss of leaf hydration that occurswhen flooding is extended for more than 5 d could not be simulatedby isolating the leaves from the roots. Appearance of this symptomdepended on leaves remaining attached to flooded root systems,implying that the damage is caused by injurious substances passingupwards (positive message). Both ethylene and ethanol have beeneliminated as likely causes, but flooding increased phosphorusin the leaves to concentrations that may be toxic. Key words: Pisum sativum, Flooding, Foliar desiccation, Stomata, Ethylene     

Flag Leaf Anatomy of Triticum and Aegilops Species in Relation to Photosynthetic Rate

Quantitative anatomical and other measurements were made onfully expanded flag leaves of a series of diploid, tetraploidand hexaploid Triticum and Aegilops species, and photosyntheticrates per unit leaf area were measured at light saturation (P_max). Diploids had the highest P_max, hexaploids the lowest with tetraploidsbeing intermediate. The anatomical features of tetraploids andhexaploids were generally similar, but different from the diploids.The diploids had thinner leaves with less dry matter and chlorophyllper unit area. The surface area of the mesophyll cells per unitvolume of mesophyll tissue was similar for all ploidy levels,as was the ratio mesophyil cell surface area per unit leaf area.It is argued that while these anatomical features are unlikelyto account for the observed variation in Pmax, it is possiblethat other structural factors with which they are correlatedmay causally influence P_max. One such feature is the averagediffusion path length from the plasmalemma at the cell surfaceto the sites of carboxylation. Anatomy, photosynthesis, mesophyll, cell size, Triticum, Aegilops, polyploidy     

Luminal L-alanine stimulates exocytosis at the K+-conductive apical membrane of Aplysia enterocytes

In Aplysia intestine,stimulation of Na⁺ absorption withluminal alanine increases apical membraneK⁺ conductance(G_K,a), whichpresumably regulates enterocyte volume during stimulatedNa⁺ absorption. However, themechanism responsible for the sustained increase in plasma membraneK⁺ conductance is not known forany nutrient-absorbing epithelium. In the present study, we have begunto test the hypothesis that the alanine-induced increase inG_K,a inAplysia enterocytes results fromexocytic insertion of K⁺ channelsinto the apical membrane. We used the fluid-phase marker horseradishperoxidase to assess the effect of alanine on apical membraneexocytosis and conventional microelectrode techniques to assess theeffect of alanine on fractional capacitance of the apical membrane(fC_a). Luminalalanine significantly increased apical membrane exocytosis from 1.04 ± 0.30 to 1.39 ± 0.38 ng · min¹ · cm².To measure fC_a,we modeled the Aplysia enterocyte as adouble resistance-capacitance (RC) electric circuit arranged in series. Several criteria were tested to confirm application of the model to theenterocytes, and all satisfied the model. When added to the luminalsurface, alanine significantly increasedfC_a from 0.27 ± 0.02 to 0.33 ± 0.04 (n = 10)after 4 min. There are two possible explanations for our findings:1) the increase in exocytosis, whichadds membrane to the apical plasma membrane, prevents plasma membranefracture, and 2) the increase inexocytosis delivers K⁺ channels tothe apical membrane by exocytic insertion. After the alanine-induceddepolarization of apical membrane potential (V_a), there isa strong correlation (r = 0.96)between repolarization ofV_a, whichreflects the increase inG_K,a, andincrease in fC_a. This correlation supports the exocytic insertion hypothesis for activation ofG_K,a.