An increase in ethylene sensitivity is associated with jasmonate-promoted senescence of detached rice leaves

The role of ethylene in jasmonate-promoted senescence of detached rice leaves was investigated. Ethylene production in methyl jasmonate-treated leaf segments of rice was lower than in the control leaves. Treatment of leaf segments with silver nitrate or/and silver thiosulfate, inhibitors of ethylene action, inhibited methyl jasmonate-, jasmonic acid-, linolenic acid-, and abscisic acid-promoted senescence of detached leaves. We suggest that an increase in ethylene sensitivity, but not ethylene level, is the initial event triggering the enhanced senescence by jasmonates of detached rice leaves.Abbreviations JA jasmonic acid - MJ methyl jasmonate - STS silver thiosulfate - ABA abscisic acid     

Ammonium, calcium, and leaf senescence in rice

The possible involvement of calcium in the regulation of ammonium-promoted senescence of detached rice leaves was investigated. Calcium effectively reduced ammonium-promoted senescence of detached rice leaves. The effect of ammonium on the senescence was also significantly reduced by the calcium ionophore A23187. Ammonium-promoted senescence of detached rice leaves may be mediated through blocking the entrance of calcium ions into the cytosol.     

Promotive effect of jasmonates on the senescence of detached maize leaves

Promotion of senescence of detached maize leaves by jasmonates was investigated. Senescence of detached maize leaves was promoted by linolenic acid, the precursor of biosynthesis of jasmonic acid, and retarded by inhibitors of lipoxygenase, the first enzyme in the biosynthetic pathway of jasmonic acid. Results support a role of endogenous jasmonates in the regulation of senescence of detached maize leaves. Silver thiosulfate, an inhibitor of ethylene action, was found to inhibit methyl jasmonate, linolenic acid- and abscisic acid-promoted senescence of detached maize leaves. It seems that jasmonate-promoted senescence is mediated through an increase in ethylene sensitivity in detached maize leaves.Abbreviations ABA abscisic acid - MJ methyl jasmonate - STS silver thiosulfate     

Endogenous abscisic acid levels are not linked to methyl jasmonate-promoted senescence of detached rice leaves

Both abscisic acid (ABA) and jasmonates are known to promote leaf senescence. Since ABA and jasmonates have both chemical and physiological similarities, we are interested to know whether senescence of detached rice leaves induced by methyl jasmonate (MJ) is mediated through an increase in endogenous ABA levels. In darkness, the endogenous level of ABA in detached rice leaves remained unchanged in the first day of incubation in water and increased about 5 times its initial value in the second day. However, the pattern of senescence, as judged by protein loss, was rapid during the first day. MJ significantly promoted senescence of detached rice leaves. Contrary to our expectation, endogenous ABA levels decreased in MJ-treated detached rice leaves. Similar to the effect of MJ, endogenous ABA levels decreased in detached rice leaves which were induced to senesce by treatment with NH₄Cl. These results suggest that endogenous ABA levels are not linked to MJ-induced senescence of detached rice leaves.     

Water stress, ammonium, and leaf senescence in detached rice leaves

Ammonium accumulation in relation to water stress-promoted senescence of detached rice leaves was investigated. The effect of water stress on the senescence of detached rice leaves is associated with the accumulation of ammonium. The accumulation of ammonium in detached rice leaves by water stress is attributed to a decrease in glutamine synthetase activity. Ammonium accumulation in detached rice leaves, induced by water stress, was accompanied by an increase in tissue sensitivity to ethylene which, in turn, accelerated leaf senescence.     

Catalase, Peroxidase, and Polyphenoloxidase Activities during Rice Leaf Senescence

The activities of catalase, peroxidase, and polyphenoloxidase were studied in attached and detached rice (Oryza sativa L. cv. Ratna) leaves. Catalase activity decreased while peroxidase and polyphenoloxidase activities increased during senescence of both attached and detached rice leaves. Kinetic (5 mum) and benzimidazole (1 mm), which are known to delay the senescence of detached rice leaves, retarded the decrease of catalase activity during detached leaf senescence. On the other hand, these chemicals accelerated the increase of peroxidase and polyphenoloxidase activities over the water control. Total phenolics accumulated in detached and darkened rice leaves, but in attached leaf senescence in light no accumulation of phenolics was observed.     

Ammonium accumulation is associated with senescence of rice leaves

The relationship between ammonium accumulation and senescence of detached rice leaves was investigated. Ammonium accumulation in detached rice leaves coincided closely with dark-induced senescence. Exogenous NH4Cl and methionine sulfoximine, which caused an accumulation of ammonium in detached rice leaves, promoted senescence. Treatments such as light and benzyladenine, which retarded senescence, decreased ammonium level in detached rice leaves. Abscisic acid, which promoted senescence, increased ammonium level in detached rice leaves. The current results suggest that ammonium accumulation may be involved in regulating senescence. Evidence was presented to show that ammonium accumulated in detached rice leaves increases tissue sensitivity to ethylene. The accumulation of ammonium in detached rice leaves during dark-induced senescence is attributed to a decrease in glutamine synthetase activity and an increase in reduction of nitrate.     

Senescence of Rice Leaves XXIV. Involvement of Calcium and Calmodulin in the Regulation of Senescence

Effects of compounds that influenced calcium uptake and calmodulininhibitors on the senescence of detached rice leaves were examined.Chelators, ethyleneglycol-bis-(ß-aminoethyl ether)-N,N,N',N'-tetraaceticacid (EGTA) and l,2-bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraaceticacid (BAPTA), significantly promoted senescence of detachedrice leaves in the dark and light. The effect of EGTA can bereversed by treating detached rice leaves with calcium. Verapamil,a calcium channel blocker, and lanthanum chloride, a calciumantagonist, promoted dark-induced, and suppressed BA- and light-retardedsenescence of detached rice leaves. Calcium ionophore A23187 [GenBank] and ruthenium red, believed to raise cytosolic level of Ca²⁺,were quite effective in retarding dark-induced and ABA-promotedsenescence of detached rice leaves. Calmodulin inhibitors, W-7,compound 48/80, chlorpromazine and trifluoperazine, significantlypromoted dark-induced, and suppressed BA- and light-retardedsenescence of detached rice leaves. It is concluded that cytosoliclevel of Ca²⁺ may regulate senescence of detached rice leavesthrough a calmodulin-dependent mechanism. (Received June 13, 1990; Accepted August 3, 1990)     

A comparative study of the effects of methyl jasmonate and abscisic acid on some rice physiological processes

The effects of methyl jasmonate (MJ) and abscisic acid (ABA) on some physiological processes of rice were compared. MJ exhibited ABA-like effects by promoting senescence of detached leaves, by inducing acid phosphatase activity of detached leaves, by inhibiting ethylene production and shoot growth of seedlings, as well as inhibiting callus formation from anthers. However, MJ and ABA had opposite effects on 1-aminocyclopropane-1-carboxylic acid-dependent ethylene production in detached leaves. The regeneration ability of anther-derived callus was inhibited by MJ but not by ABA. MJ but not ABA markedly induced peroxidase activity in senescing detached leaves. It is concluded that not all physiological processes of rice affected by MJ are similar to those by ABA.Abbreviations ABA abscisic acid - MJ methyl jasmonate - ACC 1-aminocyclopropane-l-carboxylic acid - Apase acid phosphatase     

Hydrogen peroxide is involved in methyl jasmonate-induced senescence of rice leaves

The role of H₂O₂ in the senescence of detached rice leaves induced by methyl jasmonate (MJ) was investigated. MJ treatment resulted in H₂O₂ production in detached rice leaves, which was prior to the occurrence of leaf senescence. Dimethylthiourea, a chemical trap of H₂O₂, was observed to be effective in inhibiting MJ‐induced senescence and MJ‐increased malondialdehyde (MDA) content in detached rice leaves. Diphenyleneiodonium chloride (DPI) and imidazole (IMD), inhibitors of NADPH oxidase, prevented MJ‐induced H₂O₂ production, suggesting that NADPH oxidase is a H₂O₂‐generating enzyme in MJ‐treated detached rice leaves. DPI and IMD also inhibited MJ‐promoted senescence and MJ‐increased MDA content in detached rice leaves. Phosphatidylinositol 3‐kinase inhibitors wortmannin (WM) or LY 294002 (LY) inhibited MJ‐induced H₂O₂ production and senescence of detached rice leaves. Exogenous H₂O₂ reversed the inhibitory effect of WM or LY. In terms of leaf senescence, it was observed that rice seedlings of cultivar Taichung Native 1 (TN1) are jasmonic acid (JA)‐sensitive and those of cultivar Tainung 67 (TNG67) are JA‐insensitive. On treatment with JA, H₂O₂ accumulated in the leaves of TN1 seedlings but not in the leaves of TNG67. Evidence was also provided to show that MJ‐induced H₂O₂ production in detached rice leaves is abscisic acid (ABA)‐independent. Ethylene action inhibitor, silver thiosulfate, was observed to inhibit MJ‐ and ABA‐induced H₂O₂ production and senescence of detached rice leaves, suggesting that the action of MJ and ABA is ethylene‐dependent.     

Inhibition of methyl jasmonate-promoted senescence in rice leaves by a metal chelator, 2,2′-bipyridine

The possible mediatory role of transition metals in methyl jasmonate- (MJ-)induced senescence of rice leaves was investigated. Metal chelators(2,2-bipyridine, 8-hydroxylquinoline and 1,10-phenanthroline) reducedMJ-promoted senescence of rice leaves. The reduction of MJ-promoted senescenceby 2,2-bipyridine(BP) is closely associated with the decrease in lipidperoxidation and increase in activity of superoxide dismutase (SOD). Our resultssuggest that iron or copper plays a major role in MJ-promoted senescence ofdetached rice leaves. BP-reduced senescence of detached rice leaves induced byMJ was reversed by adding Fe²⁺ or Cu²⁺, but notby Mn²⁺ or Mg²⁺. Reduction of MJ-promotedsenescence of detached rice leaves by BP is most likely mediated throughchelation of iron or copper and an increase in SOD activity.     

Ammonium ion, ethylene, and NaCl-induced senescence of detached rice leaves

The possibility that NH₄ ⁺ accumulation is linkedto the senescence of detached rice (Oryza sativa) leavesinduced by NaCl was investigated. NaCl was effective in promoting senescenceandin increasing NH₄ ⁺ content of detached rice leaves.NaCl-promoted senescence is mainly due to the effect of both Na⁺ andCl^- ions. NaCl had no or slight effect on relative water content,suggesting that an osmotic effect is unlikely to be a major factor contributingto senescence of these leaves. NaCl-induced NH₄ ⁺accumulation was due to enhanced nitrate reduction and decreased glutaminesynthetase activity. Exogenous NH₄Cl, which caused an accumulationofNH₄ ⁺ in detached rice leaves, also promoted senescence.Itwas found that an increase in NH₄ ⁺ content preceded theoccurrence of senescence caused by NaCl. Results also show that NaCl-promotedsenescence is unlikely to be due to the lack of glutamate, glutamine,aspartate,and asparagine. The current results suggest that NH₄ ⁺accumulation is linked to NaCl-induced rice leaf senescence. Since ethylene isknown to be a potent promoter of leaf senescence, we also investigated the roleof ethylene in the regulation of NH₄ ⁺-promoted senescenceof detached rice leaves. NaCl or NH₄Cl treatment resulted in adecrease of ethylene production. Evidence was presented to show thatNH₄ ⁺ accumulation in detached rice leaves does not changetissue sensitivity to ethylene. Clearly, the possible involvement of ethyleneinNH₄ ⁺-promoted senescence is excluded.     

Protease activity during rice leaf senescence

A protease activity was detected in rice (Oryza sativa L. cv. Ratna) leaves that hydrolysed hemoglobin more efficiently than bovine serum albumin. The activity was high when the enzyme was extracted and assayed with tris-maleate buffer [tris (hydroxymethyl) methyl amino-maleate] pH 7.0 rather than with water or with citrate-phosphate buffer pH 7.0. The enzyme had a strong dependence on sulfhydryl groups for its activity without which it was inaotive. The pH optimum was 7.0 and the temperature optimum was 40 °C. Protease activity expressed per unit leaf fresh weight (absolute activity) increased only little during senescence of detached rice leaves while the same activity expressed per unit soluble protein content (specific activity) increased by a greater factor (about 5 times) than absolute activity. Total and soluble protein content decreased during the senescence of detached leaves. Benzimidazole (10-3M) and kinetin (0.5xl0-5M) treatment arrested the increase of the protease activity and the deorease in the protein content during detached leaf senescence. It was indicative that protease protein was more stable than the bulk of other proteins in senescing leaves.     

Relationship between ammonium accumulation and senescence of detached rice leaves caused by excess copper

The possibility that ammonium (NH ₄ ⁺ ) accumulation is linked to the senescence of detached rice (Oryza sativa) leaves induced by copper (Cu) was investigated. CuSO₄ was effective in promoting senescence of detached rice leaves. Both CuSO₄ and CuCl₂ induced NH ₄ ⁺ accumulation in detached rice leaves, indicating that NH ₄ ⁺ accumulation is induced by copper. Sulfate salts of Mg, Mn, Zn, and Fe were ineffective in inducing NH ₄ ⁺ accumulation in detached rice leaves. The senescence of detached rice leaves induced by Cu was found to be prior to NH ₄ ⁺ accumulation. Free radical scavengers, such as glutathione and thiourea, inhibited senescence caused by Cu and at the same time inhibited Cu-induced NH ₄ ⁺ accumulation. The current results suggest that NH ₄ ⁺ accumulation is not associated with senescence induced by Cu, but is part of the overall expression of oxidative damage caused by an excess of Cu. Evidence was presented to show that copper-induced ammonium accumulation in detached rice leaves is attributed to a decrease in glutamine synthetase activity and an increase in reduction of nitrate.     

H2O2 metabolism during senescence of rice leaves: changes in enzyme activities in light and darkness

The possible role of H₂O₂ metabolism on light-regulated senescence of detached rice leaves was investigated. Light retards senescence but at the same time accumulates more H₂O₂. Light treatment resulted in an increase in malondialdehyde level in detached rice leaves but no membrane leakage was observed in light-treated detached leaves. It seems that there was no direct relationship between lipid peroxidation and deterioration in membrane integrity. The results obtained suggest that retardation of senescence by light is closely related to high activities of superoxide dismutase and ascorbate peroxidase.     

The role of proteolytic enzymes in protein degradation during senescence of rice leaves

The role of proteolytic enzymes in protein degradation of detached and intact leaves of rice seedling ( Oryza sativa L. cv. Taiching Native 1) during senescence and of mature leaves during reproductive development was investigated. The amount of soluble protein decreased by about 50% in 2, 4, and 15 days for detached, intact and mature leaves, respectively. Three proteolytic enzyme activities were monitored with pH optima of 4.5 for hemoglobin-digesting proteinase, 5.5 for carboxypeptidase and 8.0 for aminopeptidase. No azocoll-digesting proteinase activity could be detected in rice leaves. Dialysis did not alter the activities of any of the three proteolytic enzymes. Acid proteinase activity and aminopeptidase activity were highly unstable during storage of the enzyme extracts at 4°C. Proteolysis was stimulated by inclusion of meroaptoethanal either in the extraction medium or the assay medium.
Acid proteinase, carboxypeptidase and aminopeptidase were all present in detached, intact and mature leaves throughout senescence. There seems to be a direct correlation between protein degradation and increases of acid proteinase and carboxypeptidase activity in seedling leaves (detached and intact) during senescence. In senescing (detached and intact) leaves of seedlings the acid proteinase activity developed first, while that of carboxypeptidase developed later. Acid proteinase and carboxypeptidase may play major roles in protein degradation of leaves from seedlings during senscence. During reproductive development, protein degradation was associated with decreases in the activities of acid proteinase, carboxypeptidase and aminopeptidase in mature leaves suggesting that the enzymes were less important for protein degradation in this system. Hence, the role of protelytic enzymes in protein degradation during senescence of rice leaves appears to depend largely on the leaf system used.     

Effects of ABA on Senescence and Cellulase Activity of Detached Rice Leaves

The relationship of cellulase to detached leaf senescence of rice seedlings was investigated by examining the effect of ABA and 6-BA on changes in the level of cellulase of leaf segments during senescence. It was shown that the rise in cellulase activity increased with declining chlorophyll content, which was used as the senescence indicator during the senescence of detached rice leaves caused by ABA. The action of ABA took place only after a 48h lag period. ABA enhanced the cellulase secretion and increased the permeability of plasma membrane. A high level of cellulase activity in cell wall closely related to membrane permeability changes. The action of cellulase in the cell wall may cause depolymerization of β-1, 4-glucan in situ, thus speeding senescence. The 6-BA reverses completely or partly the increase in cellulase activity and tile permeability caused by ABA during the first two day, }) ut it antagonized hardly any of the ABA effect from the third day on, suggesting the onset of an irreversible stage in the senescence of detached rice leaves.     

Senescence of Rice Leaves : VII. PROLINE ACCUMULATION IN SENESCING EXCISED LEAVES

Proline content increased greatly in detached rice (Oryza sativa cv. Taichung Native 1) leaves during senescence. There was a slight but significant increase in proline level after one day of incubation, and, subsequently, proline accumulated relatively rapidly. By 4 days after excision, the level of proline had increased 30- to 50-fold, which is similar to the level seen in the water-stressed detached rice leaves. It is unlikely that the proline accumulation in detached leaves is to be derived solely from protein hydrolysis, since the addition of l-glutamic acid increased the proline level during senescence. The proline analog, 3,4-dehydroproline, did not affect the level of proline during senescence. It seems that accumulation of proline may, at least in part, result from an increased rate of synthesis, possibly due to a disruption of the normal feedback inhibition of proline synthesis. Potassium cyanide and 2,4-dinitrophenol strongly inhibited proline accumulation, indicating that some energy compound(s) may participate in proline accumulation during senescence of excised rice leaves.     

Physiology of Rice Tungro Virus Disease: Involvement of Abscisic Acid-Like Substance in Susceptible Host-Virus Interactions

Stunting was severe in susceptible rice (Oryza saliva L.) cultivar ‘Taichung Native 1’ infected with tungro virus (RTV) compared to less-susceptible cultivar ‘IR 20’. The senescence of detached leaves of RTV-infected susceptible cultivar incubated in water in dark was accelerated compared to the healthy leaves as measured by the loss of total chlorophyll content. The transpiration rate of RTV-infected leaves of the susceptible cultivar was much lower than the healthy and RTV-infected leaves of the less-susceptible cultivar. Partially purified extracts obtained from RTV-infected leaves effectively inhibited GA-induced α-amylase synthesis in barley endosperms, and rice seedling growth, and they accelerated senescence of detached rice leaves. In all the three bibassays the ABA-like activity was significantly greater in the extracts from the RTV-infected susceptible cultivar than in extracts from the less-susceptible cultivar.     

钙离子对细胞分裂素延缓水稻叶片衰老的抑制作用

单独使用细胞分裂素 (BA和 Zeatin,1 0 -9～ 1 0 -5 mol/ L和 Ca2 (1 0 -3 mol/ L)处理水稻离体叶片时 ,二者均对叶片衰老有延缓作用。但当用 Ca2 和细胞分裂素同时处理叶片时 ,细胞分裂素延缓衰老的作用受到 Ca2 的明显抑制。进一步研究表明 ,细胞分裂素和 Ca2 并未协同刺激水稻离体叶片的乙烯生成 ,这样排除了通过乙烯促进叶片衰老的可能性。用可提高细胞质 Ca2 浓度的钙通道载体 A2 31 87处理叶片时 ,可延缓叶片衰老 ;而用可降低胞质 Ca2 浓度的试剂 ,如 EGTA、La Cl3 、Verapamil、氯丙嗪等(1 0 -3 mol/ L)处理叶片时 ,可促进叶片衰老 ,进而排除了细胞分裂素促进 Ca2 的吸收而加快衰老的可能性。