Inhibitory effect of methyl jasmonate and its related compounds on kinetin-induced retardation of oat leaf senescence

The senescence-promoting activities of methyl jasmonate and its related compounds were compared with respect to structure-activity relationships. The activities were assayed by using oat ( A vena saliva L. cv. Victor) leaf segments in the presence of 2 μg/ml kinetin. Dextrorotatory methyl jasmonate prepared from an authentic sample of the racemate mixture was less active than the naturally occurring levorotatory form especially at its low concentrations (0.1 to 2.5 μg/ml). The activity of jasmonic acid, the free acid form of methyl jasmonate, was much less than the methyl ester, and this relationship was true for the other compounds tested. The reduction of the unsaturated bond in the substituent at the C-2 position and the keto group at the C-3 position greatly reduced the activity. The length of the n -alkyl substituents at the C-2 position had also a significant effect on the activity. From these results, it is concluded that the important functional groups for the high senescence-promoting activity of the methyl jasmonate related compounds are the methyl acetate substituent at the C-l position, the 2' cis -pentenyl or n -pentyl group at the C-2, position and the keto group at the C-3 position in methyl jasmonate.     

The exudation of solutes during senescence of oat leaves

Exudation of cations and of amino acids from detached oat seedling leaves ( Avena sativa L. cv. Victory) floated on water or solutions was measured under varied conditions. A small amount of exudation in the first 4 h, greater in the dark than in white light, is followed, one to 8 days later, by a relatively great increase which approximately accompanies senescence. This second increase is delayed by kinetin and is accelerated by abscisic acid or methyl jasmonate. both of which accelerate senescence. A nitrogen atmosphere (in darkness) causes immediate large exudation, mainly of potassium ions, and this is effectively delayed by light. There are two exceptions to the parallel between exudation and senescence; n-pentanol, which strongly delays senescence in darkness, nevertheless increases exudation both of potassium and of amino acids, probably due to a direct effect on the plasmalemma. Cycloheximide, which also delays senescence, increases the exudation somewhat. Kinetin prevents or delays exudation under all conditions. Thus, the permeability of the plasmalemma increases greatly along with other criteria of senescence, but this change is probably not the principal cause of the senescence syndrome. Some of the effects are considered to result from reduction in available energy and others from a direct influence on plasmalemma permeability.     

Light-induced h secretion and the relation to senescence of oat leaves

When abraded oat (Avena sativa L. cv Victory) leaf segments are floated on KCl solution, white light causes acidification of the solution external to leaf tissue. The presence of mannitol amplifies the light-induced proton secretion. Mature leaves as well as young ones acidify the medium in light, while senescing leaves (after 3 to 4 days incubated in water in the dark) lose the ability to produce this response to light. The decrease in H⁺ secretion is already measureable after as little as 30 minutes in darkness, while the increase in proteolysis rate was detected only after 6 hours in dark. The decrease in capacity to secrete protons is one of the symptoms of leaf senescence. Moreover, fusicoccin mimics light in stimulating H⁺ pumping and delaying the senescence in the dark. On the other hand vanadate, an apparent inhibitor of plasma membrane H⁺ ATPase, blocks the acidification and promotes the chlorophyll and protein degradation in leaf segments during the 2-day period of incubation. These results, which show a parallel between cessation of H⁺ secretion and acceleration of senescence, may suggest a regulatory role for H⁺ secretion in leaf senescence.     

Effects of methyl jasmonate (MeJA) on the dark-induced senescence in oat (Avena sativa L.) leaf segments

To investigate the relationship between methyl jasmonate (MeJA) and ethylene in leaf senescence, we studied the effects of MeJA on ethylene production and ethylene biosynthetic enzyme activities in oat(Avena sativa L.) leaf segments incubated in darkness. MeJA promoted dark-induced senescence judged from the contents of chlorophyll and protein, and increased ethylene production 6 times of the control. MeJA also increased the activities of ethylene biosynthetic enzymes, 1-aminocyclopropane carboxylic acid (ACC) synthase and ACC oxidase as compared to control. In MeJA-treated leaf segments, ACC synthase activity reached its maximum level at 24 h of incubation and ACC oxidase activity peaked at 6 h of incubation. Aminoethoxyvinylglycine (AVG) and Co²⁺, inhibitors of ACC synthase and ACC oxidase respectively, reduced MeJA-induced ethylene production. They also delayed leaf senescence that was promoted by the treatment of MeJA. From these results, we can suggest that MeJA increased the activities of ACC synthase and ACC oxidase, these increased activities lead to increase in ethylene production and this increased ethylene production might promote dark-induced leaf senescence.     

The role of ethylene in the senescence of oat leaves

The evolution of ethylene, both from the endogenous source and from added 1-aminocyclopropane-1-carboxylic acid (ACC), has been followed in close relationship with the senescent loss of chlorophyll from seedling oat leaves. In white light, where chlorophyll loss is slow, the ethylene evolution increases slowly at first, but when the loss of chlorophyll becomes more rapid, ethylene evolution accelerates. CoCl₂ inhibits this increase and correspondingly maintains the chlorophyll content, with an optimum concentration of 10 micromolar. The rapid rate of chlorophyll loss in the dark is slightly decreased by 3-aminoethoxyvinyl glycine (AVG), by cobalt, and slightly stimulated by ACC. The slower chlorophyll loss in white light, however, is almost completely inhibited by silver ions, greatly decreased by cobalt and by AVG, and strongly increased by ACC. Since the chlorophyll loss is accompanied by proteolysis, it represents true senescence. Chlorophyll loss in light is also strongly antagonized by CO₂, 1% CO₂ giving almost 50% chlorophyll maintenance in controls, while in the presence of added ACC or ethylene gas, the chlorophyll loss is 50% reversed by about 3% CO₂. The ethylene system in leaves is thus more sensitive to CO₂ than that in fruits. Indoleacetic acid also clearly decreases the effect of ACC. It is shown that kinetin, CO₂, Ag⁺, and indoleacetic acid, all of which oppose the effect of ethylene, nevertheless increase the evolution of ethylene by the leaves, and it is suggested that ethylene evolution may, in many instances, mean that its hormonal metabolism is being prevented.     

Raman imaging of changes in the polysaccharides distribution in the cell wall during apple fruit development and senescence

Planta - Du ring on-tree ripening, the pectin distribution changed from polydispersed in cell wall to cumulated in cell wall corners. During apple storage, the pectin distribution returned to...     

Changes in cell wall polysaccharides associated with growth

Changes in the polysaccharide composition of Phaseolus vulgaris, P. aureus, and Zea mays cell walls were studied during the first 28 days of seedling development using a gas chromatographic method for the analysis of neutral sugars. Acid hydrolysis of cell wall material from young tissues liberates rhamnose, fucose, arabinose, xylose, mannose, galactose, and glucose which collectively can account for as much as 70% of the dry weight of the wall. Mature walls in fully expanded tissues of these same plants contain less of these constituents (10%-20% of dry wt). Gross differences are observed between developmental patterns of the cell wall in the various parts of a seedling, such as root, stem, and leaf. The general patterns of wall polysaccharide composition change, however, are similar for analogous organs among the varieties of a species. Small but significant differences in the rates of change in sugar composition were detected between varieties of the same species which exhibited different growth patterns. The cell walls of species which are further removed phylogenetically exhibit even more dissimilar developmental patterns. The results demonstrate the dynamic nature of the cell wall during growth as well as the quantitative and qualitative exactness with which the biosynthesis of plant cell walls is regulated.     

Changes in the rate of synthesis of wall polysaccharides during the cell cycle of yeast

Reevaluation and comparison of seemingly contradictory literature data on the mode of synthesis of wall polysaccharides during the cell cycle ofSaccharomyces cerevisiae explained the source of discrepancies and demonstrated their general consonance in the following points: 1. The rate of synthesis of glucan and mannan is not constant and does not increase continuously throughout the entire cell cycle. 2. The rate of synthesis of both polysaccharides is considerably reduced at the time of cell division and in the prebudding phase.     

Changes in the activities of enzymes involved in amino acid metabolism during the senescence of detached wheat leaves

The activities of several enzymes related to amino acid metabolism were investigated in senescing detached wheat leaves ( Triticum aestivum L. cv. Diplomat) in light and darkness and after kinetin treatment. Glutamine synthetase and glutamate synthase activities rapidly declined in darkness. In light, the decline of glutamate synthase activity was retarded, while the activity of glutamine synthetase remained high and even increased transitorily. Kinetin treatment counteracted the decline of the activities of both enzymes. The activity of glutamate dehydrogenase markedly increased during senescence, particularly in light, and kinetin treatment lowered its activity. The activities of glutamate-oxaloacetate and glutamate-pyruvate amino-transferases and of NADP-dependent isocitrate dehydrogenase also increased in detached wheat leaves in light. Kinetin treatment prevented the rise of these enzyme activities. In darkness, the activities of glutamate-oxaloacetate aminotransferase and NADP-dependent isocitrate dehydrogenase decreased slowly while the decline of glutamate-pyruvate aminotransferase activity was more rapid. The activity of NAD-dependent malate dehydrogenase decreased both in light and, more rapidly, in darkness. The pattern of changes of the enzyme activities provides an explanation for the amino acid transformations and the flow of amino nitrogen into transport metabolites in senescing leaves.     

Changes in cell wall polysaccharides of germinating barley grains

Decorticated barley grains were germinated at 25° for 6 days, until the endosperm reserves were nearly exhausted. The neutral monosaccharide components of the hydrolysates of the cell walls and gums from the embryo, aleurone layer and starchy endosperm and the endospermic starch were determined at daily intervals. The amount of embryo cell wall polysaccharide increased 40 times and glucose became the major component, followed in abundance by xylose and arabinose. The cell wall and gum polysaccharides of the aleurone layer (plus testa) and the starchy endosperm declined during germination and their compositions altered. The endospermic starch also decreased. In the early stages of germination the apparent composition of the cell walls of the aleurone layer and starchy endosperm depended upon how they had been prepared. After 6 days the cell walls and gums had provided a significant carbohydrate supply to the living tissues, equivalent to 18.5% of the endospermic polysaccharide degraded during growth, starch having provided the remaining 81.5%.     

Changes in endogenous growth regulators in nasturtium leaves during senescence

Summary By use of lettuce-hypocotyl and wheat-coleoptile bioassay, the presence of both gibberellin (GA)-like and abscisic-acid(ABA)-like components in acidic ethyl-acetate extracts of fully expanded nasturtium (Tropaeolum majus) leaves has been shown. During senescence of detached leaves there was a progressive decline in GA-like components and an increase in ABA-like components. Pretreatment of detached leaves with GA₃ or kinetin prevented changes in the levels of endogenous growth regulators and delayed senescence. The observations provide experimental verification for the concept that senescence is associated with changes in endogenous growth regulators.     

Changes in photosystem II function during senescence of wheat leaves

Analyses of chlorophyll fluorescence were undertaken to investigate the alterations in photosystem II (PSII) function during senescence of wheat ( Triticum aestivum L. cv. Shannong 229) leaves. Senescence resulted in a decrease in the apparent quantum yield of photosynthesis and the maximal CO₂ assimilation capacity. Analyses of fluorescence quenching under steady‐state photosynthesis showed that senescence also resulted in a significant decrease in the efficiency of excitation energy capture by open PSII reaction centers (F'_v/F'_m) but only a slight decrease in the maximum efficiency of PSII photochemistry (F'_v/F'_m). At the same time, a significant increase in non‐photochemical quenching (q_N) and a considerable decrease in photochemical quenching (q_P) were observed in senescing leaves. Rapid fluorescence induction kinetics indicated a decrease in the rate of Q_A reduction and an increase in the proportion of Q_B‐non‐reducing PSII reaction during senescence. The decrease in both F'_v/F'_m and q_P explained the decrease in the actual quantum yield of PSII electron transport ((φ_PSII). We suggest that the modifications in PSII function, which led to the down‐regulation of photosynthetic electron transport, would be in concert with the lower demand for ATP and NADPH in the Calvin cycle which is often inhibited in senescing leaves.     

Multiple actions of abscisic acid in senescence of oat leaves

The modifications induced by abscisic acid (ABA) on the senescence of oat leaves in darkness have been studied and are compared with its well-known effects in light. Contrary to the action in light, ABA preserves chlorophyll (Chl) in the dark almost as well as kinetin. Chlorophylla is decolorized more extensively thanb, and the content ofb is maintained by ABA almost at its initial level for 4 days. ABA also prevents proteolysis in darkness just as completely as chlorophyll loss, the relationship of both breakdown processes to ABA concentration being strictly log-linear over the range from 1 to 100 M. In line with this action, ABA inhibits formation of the neutral protease in the dark but not in the light. The data suggest that ABA and kinetin operate to preserve chlorophyll and protein by different mechanisms, since their actions are neither independent nor synergistic but actually interfere with one another. In this connection, protein values given by the Lowry and Bradford methods have been compared. In parallel with the effect on senescence, ABA slowly opens the stomata in the dark. This effect increases with time, and by day 3 the stomata in ABA are as open as in leaves on water in light. Thus all these effects of ABA in darkness are strikingly opposite to those commonly observed on leaves in natural lighting. In addition, ABA powerfully inhibits the formation of ethylene in the dark by the detached oat leaves, and this inhibition also tends to increase with time. Finally, a slight antagonism to ABA's action on senescence is exerted byp-coumaric acid in the light but not in the dark.     

Senescence in oat leaves: Changes in translatable mRNAs

Changes in translatable mRNA populations during the senescence of oat (Avena sativa L. cv. Victory) leaves were examined by analyzing the in vitro translation products of isolated RNA. Total RNA was isolated from oat leaves of 7-day-old seedlings, and also after these leaves were aged for different lengths of time under various conditions. Polypeptides from in vitro translations were separated by two-dimensional gel electrophoresis to estimate any changes in translatable mRNA populations associated with senescence. Corresponding leaf samples were monitored for loss of chlorophyll as a measure of the extent of senescence. The aging of excised leaves in the light for 4 days resulted in the disappearance or substantial quantitative decrease of a number of mRNA species, while only five new translatable mRNA species were produced. Three of these mRNAs were unique to aging of leaves under light. Two of these mRNA species were also produced during the early stages of senescence in attached leaves of seedlings grown under light. The translatable mRNA populations of leaves aged for 4 days either on intact seedlings or detached and kept in the light in the presence of kinetin were very similar. Aging of excised leaves in the dark on water for 24 h resulted in very extensive changes in translatable mRNA populations. Over thirty polypeptides disappeared or were substantially reduced in quantity, while about an equal number appeared de novo or were substantially increased in quantity. Aging of these leaves for an additional 24 or 48 h resulted in only a few additional changes in translatable mRNAs. The presence of kinetin during aging of excised leaves in the dark inhibited few of the numerous changes in mRNAs that occured during the first 24 h, but did inhibit most of the changes that occured after 48 or 72 h of aging in the dark. When leaves were first aged in the dark and then returned to light, most of the initial changes in translatable mRNAs expression were reversed. Such changes in mRNAs thus appear to be light-regulated and not necessarily associated with senescence.     

Relation of polyamine synthesis and titer to aging and senescence in oat leaves

Polyamine biosynthesis in senescing leaves of Avena sativa L. was measured by determining the activities of arginine decarboxylase (EC 4.1.1.19), ornithine decarboxylase (EC 4.1.1.17) and S-adenosyl-l-methionine decarboxylase (EC 4.1.1.50). Polyamine content was also estimated by thin layer chromatography and high performance liquid chromatography. Arginine decarboxylase activity decreases progressively in aging attached first leaves and in senescing excised leaves in the dark. Conversely, it increases during light exposure of excised leaves, which retards senescence. Ornithine decarboxylase activity is high and constant in the attached leaf, irrespective of age; it decreases in excised leaves kept in the dark and in the light, irrespective of senescence. S-Adenosyl-l-methionine decarboxylase shows no correlation with age or senescence. Levels of putrescine, diaminopropane, agmatine, and spermidine are high in young leaves and decline with age. The best single indicator of senescence is usually spermidine, which decreases in excised leaves incubated in the dark, but increases in such leaves with time of light exposure. Spermidine generally has a reciprocal relationship with putrescine, indicating that spermidine synthase, which converts putrescine to spermidine, may exert important physiological control. These data support the view that polyamines play an important role in the regulation of plant development.     

Accumulation of chloroplast-targeted lipoxygenase in passion fruit leaves in response to methyl jasmonate

Wounding caused local and systemic induction of lipoxygenase (LOX) activity in passion fruit (Passiflora edulis f. flavicarpa) leaves, while exposing intact plants to methyl jasmonate (MJ) vapor provoked a much stronger response. Western blot analysis of these leaf protein extracts using polyclonal antibodies against cucumber LOX, revealed an accumulation of a 90 kDa protein, consistent with LOX enzymatic assays. The inducible LOX was purified to apparent homogeneity, and in vitro analysis of LOXactivity using linoleic acid as substrate showed that it possesses C-13 specificity. Immunocytochemical localization studies using leaf tissue from MJ-treated plants demonstrated that the inducible LOX was compartmented in large quantities in the chloroplasts of mesophyll cells, associated with the stroma. The results suggest that the wound response in passion fruit plants may be mediated by a chloroplast 13-LOX, a key enzyme of the octadecanoid defense-signaling pathway.     

Inhibition of the synthesis of cell wall polysaccharides in oat coleoptile segments by jasmonic acid: Relevance to its growth inhibition

The inhibitory mode of action of jasmonic acid (JA) on the growth of etiolated oat (Avena sativa L. cv. Victory) coleoptile segments was studied in relation to the synthesis of cell wall polysaccharides using [¹⁴C]glucose. Exogenously applied JA significantly inhibited indoleacetic acid (IAA)-induced elongation of oat coleoptile segments and prevented the increase of the total amounts of cell wall polysaccharides in both the noncellulosic and cellulosic fractions during coleoptile growth. JA had no effect on neutral sugar compositions of hemicellulosic polysaccharides but substantially inhibited the IAA-stimulated incorporation of [¹⁴C]glucose into noncellulosic and cellulosic polysaccharides. JA-induced inhibition of growth was completely prevented by pretreating segments with 30 mm sucrose for 4 h before the addition of IAA. The endogenous levels of UDP-sugars, which are key intermediates for the synthesis of cell wall polysaccharides, were not reduced significantly by JA. Although these observations suggest that the inhibitory mode of action of JA associated with the growth of oat coleoptile segments is relevant to sugar metabolism during cell wall polysaccharide synthesis, the precise site of inhibition remains to be investigated.Abbreviations JA jasmonic acid - ABA abscisic acid - IAA indoleacetic acid - T ₀ minimum stress relaxation time - TFA trifluoroacetic acid - TCA trichloroacetic acid - HPLC high-performance liquid chromatography - EtOAc ethyl acetate - TLC thin-layer chromatography - JA-Me methyl jasmonate - GLC-SIM gas-liquid chromatography-selected ion monitoring     

Changes in cell wall polysaccharides in the internodes of submerged floating rice

In excised stem segments of floating rice (Oryza sativa L.), as well as in intact plants, submergence greatly stimulates the elongation of internodes. The differences in the composition of cell wall polysaccharides along the highest internodes of submerged and air-grown stem segments were examined. The newly elongated parts of internodes that had been submerged for two days contained considerably less cellulosic and noncellulosic polysaccharides than air-grown internodes, an indication that the cell walls of the newly elongated parts of submerged internodes are extremely thin. In the young parts of both air-grown and submerged internodes, the relative amounts of noncellulosic polysaccharides were equal to those of -cellulose, whereas the relative amounts of -cellulose were higher than those of noncellulosic polysaccharides in the upper, old parts. In the cell-elongation zones of both air-grown and submerged internodes, glucose was predominant among the noncellulosic neutral sugars of cell wall. The relative amount of glucose in noncellulosic neutral sugars decreased toward the upper, old parts of internodes, whereas that of xylose increased.     

Growth and cell wall changes in azuki bean epicotyls I. Changes in wall polysaccharides during intact growth

Measurement of endogenous growth rates and the mechanical propertyof the cell wall in various regions of light-grown azuki beanepicotyls revealed diat the minimum stress-relaxation time (T_o)was the shortest in the upper region (0–30 mm below theapex) of the epicotyl, where vigorous endogenous growth tookplace, and became longer toward the basal region, which wasmature and not growing. In the upper region of the epicotyl, a lower percentage of a-celluloseand a higher percentage of pectic substances than in the lowerregion were found. The percentage of hemicellulose content wasalmost constant over the whole epicotyl. Major components ofnoncellulosic neutral sugars in the cell wall were galactoseand xylose. The percentage of the galactose content to the noncellulosicpolysaccharide was highest in the upper region and lowest inthe basal region of the epicotyl, and a clearly negative correlationbetween the galactose composition and the T_o value was obtained.On the contrary, the percentage of die xylose content was highestin the basal region and lowest in die upper region, and a clearlypositive correlation between die xylose composition and theT_o value was obtained. During die endogenous growth of die intactepicotyl, all die neutral sugars, particularly galactose, increasedin die upper region, whereas in die middle and basal regions,only xylose increased. Similar changes in sugar compositionswere observed during IAA-induced elongation of die segment excisedfrom various regions of die epicotyl. (Received July 27, 1978; )     

Changes in cell wall polysaccharides and monosaccharides during development and ripening of Japanese pear fruit

1. Changes in polysaccharide and monosaccharide components in thecell wall were studied during cell division, cell enlargmementand softening in Japanese pear fruit. Wall polysaccharides werefractionated into water soluble carbohydrate, NaClO₂ solublecarbohydrate, EDTA soluble carbohydrate, acid soluble hemicellulose,alkali soluble hemicellulose and cellulose. These polysaccharideswere composed of glucose, uronic acid, xylose, arabinose, galactose,rhamnose, mannose and fucose.
2. The total polysaccharide contentof the cell wall per cell (DNAcontent basis) remained constantduring the cell division period(S1). But during the pre-enlargementperiod (S2) it began toincrease rapidly in spite of the slightnessof cell enlargement.Thereafter, during the enlargement period(S3) the polysaccharidesremained almost constant although thefruits enlarged dramatically,and the polysaccharides increasedsomewhat with ripening. Thequality of the polysaccharides,however, seemed to change activelyat each stage. This suggestedthat the extensive fruit enlargementdid not require an increasein polysaccharide content, and wasrather accompanied by thepartial breakdown or partial interconversionof polysaccharidecomponents already present.
3. The loss of arabinose and galactosein acid soluble hemicellulosewas prominant in fruit softeningoccurring in the ripening stage.The cellulose component decreasedwith overripening. Water solublepectin increased parallel tothe increase in total pectin withripening. On the other hand,xylose and non-cellulosic glucoseresidues did not alter withripening or overripening. Non-cellulosicglucose continued toaccumulate during cell enlargement.

¹ This paper is Contribution A-88, Fruit Tree Research Station. (Received August 4, 1978; )