Energy Metabolism of Rumex Leaf Tissue in the Presence of Senescence-regulating Hormones and Sucrose

209

In spinach (Spinacia oleracea Hybrid 102 INew World seedsl) and wheat (Triticum aestivum L. cv Gabo) leaves, 02 uptake rates in the dark were faster after the plants had been allowed to photosynthesize for a period of several hours. Alternative path activity also increased following a period of photosynthesis in these leaves. No such effects were observed with isolated mitochondria. In spinach and wheat leaves, the level of fructose plus glucose decreased during a period of darkness. In pea (Pisum sativum cv Alaska) leaves, the level of these sugars did not vary significantly during the day, and respiratory rates were also constant. In slices cut from wheat leaves harvested at the end of the night, addition of sugars increased the rate of respiration and engaged the previously latent alternative oxidase. Several lines of evidence suggest that there is a direct relationship between the rate of respiration and the carbohydrate status of plant tissues. We have shown previously that the rate of respiratory CO2 efflux of mature wheat leaves is correlated with the rate of prior CO2 assimilation, via carbohydrate accumulation (3), and similar correlations have been found in leaves of other species (5, 14). Exogenous sugars stimulate CO2 efflux and 02 consumption in the dark in the leaves of Rumex acetosa (9) and Avena sativa (17), and similar effects have been observed with maize root tips (16).Recent work in our laboratory has shown that the cyanideinsensitive, alternative pathway of respiration is present in the leaves of many plant species, and that it may contribute significantly to measured respiratory rates in the dark (12). However, little is known concerning the regulation of this pathway in leaves. In this report, we present results from an investigation of the relationship between the rate of respiratory 02 uptake (in the dark), the involvement of the alternative path, and the level of (12). Quantum flux (400-700 nm) was 600 to 700 ,uE m-2 S-1, supplied by Sylvania Gro-Lux and cool-white fluorescence tubes.The day/night temperature regime was 25/21°C with a daylength of 13 h for wheat and 10 h for peas and spinach. Mature leaves were selected from these plants at the end of the night or after several hours in the light in the cabinet.Measurement of Oxygen Uptake in Leaves. Intact pea and spinach leaves or wheat leaf segments (30-80 cm2 area) were placed in an 02 electrode cuvette containing 105 ml of reaction medium (50 mm Hepes, 10 mm Mes buffer [pH 6.61,0.2 mM CaCl2, for wheat; and the same solution but with 10 mm Hepes for peas and spinach) in equilibrium with air. The wheat leaf segments (5-8 cm long) were obtained by sectioning leaves perpendicularly to the veins under buffer (see above) with a sharp razor blade. The linear depletion of the 02 concentration in a closed system was then measured polarographically using a Clark type 02 electrode. The solution was strongly stirred and 02 became limiting for respiration only at low concentrations (less than 100 tM). Measurements were made in the dark...

supporting

confidence: 49%

Effect of Photosynthesis and Carbohydrate Status on Respiratory Rates and the Involvement of the Alternative Pathway in Leaf Respiration

Azcón-Bieto¹,

Lambers²,

Day³

1983

209

“…Both proteolysis and Chl loss can be effectively prevented by cytokinins and, in a few cases, also by gibberellins (e.g. 4,13). The mechanism of this interference with hydrolytic reactions is under active study.…”

mentioning

confidence: 99%

The Metabolism of Oat Leaves during Senescence

Choe¹,

Thimann²

1975

The changes in chlorophyll and protein in senescing chloroplasts isolated from the first leaves of 7-day-old oat (Avena sativa) seedlings have been investigated. In darkness the chlorophyll in these plastids is highly stable, losing only 5 to 10% of its content after 7 days at 26 C. This result contrasts with the behavior of chlorophyll in intact leaves, in which about 80% of the pigment would have disappeared in that time. The protein is less stable than the chlorophyll, though more stable than in the leaf; probably a small amount of protease is present in the plastids. Some protein is also being synthesized in the chloroplasts along with its breakdown; gains of up to 38% in protein and 13% in chlorophyll were observed under different conditions. L-Serine, which actively promotes senescence in the leaf, has only a very slight effect on the chloroplasts, and kinetin antagonizes it. Kinetin also has a small but significant effect in preserving the protein from breakdown. Acid pH somewhat promotes the breakdown, both of chlorophyll and protein. A loss of chlorophyll and protein comparable to that occurring in the senescence of the leaf could not be induced in the chloroplasts by suspending them in malate, in cytoplasmic extract, or in any of a number of enzymes tested alone. Incubation with a mixture of four enzymes was the only treatment which approximated the senescent process in the leaf, causing 34% loss of chlorophyll at pH 5 and 40% loss of protein at pH 7.4, both in 72 hours.In white light, the chlorophyll and the carotenoids, but not the protein, disappear rapidly. This disappearance was shown to be prevented in an atmosphere of nitrogen or in air by a number of reducing agents, of which ascorbic acid was the most effective. It is, therefore, ascribed to photooxidation rather than to normal senescence.The senescence of leaves, which has been studied in a number of laboratories over the years (3, 7, 10, 11, 18, 19, 21-23, 25, 26, 29, 31, 33-39), involves loss of Chl, decrease in carotenoids, enhanced proteolysis, hydrolysis of starch and other polysaccharides, and, in some cases, increase in respiration. In detached leaves, the hydrolyzed products accumulate and may be partly converted to organic acids (21, 23, 37), whereas if the leaves remain on the plant the amino acids, and perhaps 'This work was supported in part by National Science Foundation Grant GB35238 to K. V. T. other products, are re-exported to the stem or base (34). Both proteolysis and Chl loss can be effectively prevented by cytokinins and, in a few cases, also by gibberellins (e.g. 4, 13). The mechanism of this interference with hydrolytic reactions is under active study.In our own laboratory, detailed studies of the senescent process in the detached first leaves of 7-day-old oat seedlings have shown that senescence is largely prevented not only by cytokinins but also by anaerobiosis and by certain inhibitors of protein synthesis, i.e. it is dependent on some synthetic processes, probably formation of enzymes, at the start. There ...

“…Furthermore, isocitrate oxidation activity both in the presence and absence of ATP was enhanced by ABA. (6), and Rumex leaf discs (5). In these experiments, whole plants and tissues treated with ABA have been used.…”

Section: Resultsmentioning

confidence: 99%

“…Goldthwaite (5) observed that ABA stimulates oxygen consumption during senescence of Rumex leaf discs. Hourmant and Penot (6) reported that the rate of respiration of aerated potato tuber discs increased after a 24-h aging period with ABA in the dark.…”

mentioning

confidence: 99%

Activation of O₂ Uptake and NAD-Specific Isocitrate Dehydrogenase in Mitochondria Isolated from Cotyledons of Castor Bean by cis, trans-Abscisic Acid

Tezuka

Yamamoto²,

Kondo³

1990

The rate of 02 uptake and the activity of NAD-specific isocitrate dehydrogenase (NAD-ICDH) of mitochondria isolated from castor bean cotyledons were increased by added cis,trans-abscisic acid (ABA) in an in vitro system, while the NADP-specific isocitrate dehydrogenase (NADP-ICDH) was not affected by cis,trans-ABA. Trans,trans-ABA showed only a slightly inhibitory effect on 02 uptake. The Vmax value for the isotherm of isocitrate by the enzyme was also increased by cis,trans-ABA. The isocitrate Km value for the enzyme with cis,trans-ABA was calculated to be approximately 249.8 micromolar, while the S05 for the enzyme without the ABA was 151.6 micromolar. The n value calculated from the slopes of Hill plots of the reaction velocity of NAD-ICDH against isocitrate concentration was 1.5 in the mitochondrial fraction in the absence of ABA, and cis,trans-ABA treatment decreased the value to 1.0. Cis,trans-ABA also partly overcame the inhibition of NAD-ICDH activity by ATP. It has been reported that the respiratory rate (10) and ABA content (9) in grape, expressed on a per berry basis, increased during the ripening process after veraison (which means the onset of maturation [13]). These phenomena suggest a close relationship between ABA content and respiratory metabolism in mature tissues and organs of plants.Goldthwaite (5) observed that ABA stimulates oxygen consumption during senescence of Rumex leaf discs. Hourmant and Penot (6) reported that the rate of respiration of aerated potato tuber discs increased after a 24-h aging period with ABA in the dark. Tillberg et al. (16) found that respiration in Lemna gibba L. was significantly stimulated 2 to 7 d after ABA treatment. In the unicellular green algae, Chlorellafusca and Ankistrodesmus brauni, dark respiration was increased by ABA (17).In the present work, we studied the stimulating action of ABA on respiration of mitochondria isolated from cotyledons of castor beans and found evidence for the regulation of enzyme activity by ABA in an in vitro system. MATERIALS AND METHODS Plant MaterialsCastor beans (Ricinus communis L.) were grown under the conditions described previously by Kondo and Sugahara (7).The plants were grown for 2 to 3 weeks at 25 ± 0.5°C during the day and 20 ± 0.5°C at night with 70 ± 5% RH in an environmentally controlled glasshouse under natural light conditions. The growth substratum was composed of vermiculite: peat moss:perlite:fine gravel (2:2:1:1, v/v/v/v). A nutrient solution containing 8 g of magnesia lime and 4 g of hyponex per liter was supplied twice a week to each 1.9 L pot. Isolation of MitochondriaFive pairs of castor bean cotyledons were cut into small pieces (about 15 g) and homogenized with 45 mL of 25 mM Hepes buffer (pH 7.8) containing 0.35 M mannitol, 0.25 M sucrose, and 0.1 mg Na-2-mercaptobenzothiazole and 1 mg BSA per mL (14) using a Homo blender (18,000 rpm, 100 W, Sakuma Seisakusho, Ltd., Tokyo) at a quarter of maximum speed for 20 s. The homogenate was squeezed through four layers of gauze and centrifuged for 10...