Exposure to Light of the Abaxial versus Adaxial Side of Detached Kalanchoë blossfeldiana Leaves Affects Anthocyanin Content and Composition Differently.

The accumulation and composition of anthocyanins in leaves of Kalanchoë blossfeldiana, detached and kept for five days under natural light conditions, were investigated. The presence of fifteen derivatives of cyanidin, petunidin, and delphinidin was found. Changes in the content of each anthocyanin in the leaves before and after exposure to light on the abaxial (naturally upper) and adaxial (naturally lower) sides of the leaves were compared. When the adaxial side was exposed to light, the anthocyanin contents of the leaves did not change. In contrast, when the abaxial side of detached leaves was exposed to light, there was enhanced accumulation of delphinidin-rhamnoside-glucoside, cyanidin-rhamnoside-glucoside, cyanidin-glucoside-glucoside, and two unknown derivatives of petunidin and delphinidin. Application of methyl jasmonate (JA-Me) on the abaxial side exposed to light inhibited the accumulation of these anthocyanins. This effect could probably be due to the presence of these anthocyanins in the epidermal cells of K. blossfeldiana leaves and was visible in the microscopic view of its cross-section. These anthocyanins were directly exposed to JA-Me, leading to inhibition of their formation and/or accumulation. The lack of significant effects of JA-Me on anthocyanin mono- and tri-glycosides may indicate that they are mainly present in the mesophyll tissue of the leaf.

A Possible Mode of Action of Methyl Jasmonate to Induce the Secondary Abscission Zone in Stems of Bryophyllum calycinum: Relevance to Plant Hormone Dynamics.

Plants can react to environmental stresses through the abscission of infected, damaged, or senescent organs. A possible mode of action of methyl jasmonate (JA-Me) to induce the formation of the secondary abscission zone (SAZ) in the stems of Bryophyllum calycinum was investigated concerning plant hormone dynamics. Internode segments were prepared mainly from the second or third internode from the top of plants with active elongation. JA-Me applied to the middle of internode segments induced the SAZ formation above and below the treatment after 5-7 days. At 6 to 7 days after JA-Me treatment, the above and below internode pieces adjacent to the SAZ were excised and subjected to comprehensive analyses of plant hormones. The endogenous levels of auxin-related compounds between both sides adjacent to the SAZ were quite different. No differences were observed in the level of jasmonic acid (JA), but the contents of 12-oxo-phytodienoic acid (OPDA), a precursor of JA, and N-jasmonyl-leucine (JA-Leu) substantially decreased on the JA-Me side. Almost no effects of JA-Me on the dynamics of other plant hormones (cytokinins, abscisic acid, and gibberellins) were observed. Similar JA-Me effects on plant hormones and morphology were observed in the last internode of the decapitated growing plants. These suggest that the application of JA-Me induces the SAZ in the internode of B. calycinum by affecting endogenous levels of auxin- and jasmonate-related compounds.

Accumulation of Anthocyanins in Detached Leaves of Kalanchoë blossfeldiana: Relevance to the Effect of Methyl Jasmonate on This Process.

Accumulation of anthocyanins in detached leaves and in excised stems of Kalanchoë blossfeldiana kept under natural light conditions in the presence or absence of methyl jasmonate (JA-Me) was investigated. When the abaxial surface of detached leaves was held lower than the adaxial surface (the normal or natural position) under natural light conditions, anthocyanins were not accumulated on the abaxial side of the leaves. In contrast, when the adaxial surface of detached leaves was held lower than the abaxial surface (inverted position), anthocyanins were highly accumulated on the abaxial side of the leaves. These phenomena were independent of the growth stage of K. blossfeldiana as well as photoperiod. Application of JA-Me in lanolin paste significantly inhibited anthocyanin accumulation induced on the abaxial side of detached leaves held in an inverted position in a dose-dependent manner. Anthocyanin accumulation in the excised stem in response to natural light was also significantly inhibited by JA-Me in lanolin paste. Possible mechanisms of anthocyanin accumulation on the abaxial side of detached K. blossfeldiana leaves held in an inverted position under natural light conditions and the inhibitory effect of JA-Me on this process are described. The accompanying changes in the content of primary metabolites and histological analyses were also described.

Effect of Methyl Jasmonate on the Terpene Trilactones, Flavonoids, and Phenolic Acids in Ginkgo biloba L. Leaves: Relevance to Leaf Senescence.

The present study compared the effects of natural senescence and methyl jasmonate (JA-Me) treatment on the levels of terpene trilactones (TTLs; ginkgolides and bilobalide), phenolic acids, and flavonoids in the primary organs of Ginkgo biloba leaves, leaf blades, and petioles. Levels of the major TTLs, ginkgolides B and C, were significantly higher in the leaf blades of naturally senesced yellow leaves harvested on 20 October compared with green leaves harvested on 9 September. In petioles, a similar effect was found, although the levels of these compounds were almost half as high. These facts indicate the importance of the senescence process on TTL accumulation. Some flavonoids and phenolic acids also showed changes in content related to maturation or senescence. Generally, the application of JA-Me slightly but substantially increased the levels of TTLs in leaf blades irrespective of the difference in its application side on the leaves. Of the flavonoids analyzed, levels of quercetin, rutin, quercetin-4-glucoside, apigenin, and luteolin were dependent on the JA-Me application site, whereas levels of (+) catechin and (-) epicatechin were not. Application of JA-Me increased ferulic acid and p-coumaric acid esters in the petiole but decreased the levels of these compounds in the leaf blade. The content of p-coumaric acid glycosides and caffeic acid esters was only slightly modified by JA-Me. In general, JA-Me application affected leaf senescence by modifying the accumulation of ginkogolides, flavonoids, and phenolic acids. These effects were also found to be different in leaf blades and petioles. Based on JA-Me- and aging-related metabolic changes in endogenous levels of the secondary metabolites in G. biloba leaves, we discussed the results of study in the context of basic research and possible practical application.

Mode of Action of 1-Naphthylphthalamic Acid in Conspicuous Local Stem Swelling of Succulent Plant, Bryophyllum calycinum: Relevance to the Aspects of Its Histological Observation and Comprehensive Analyses of Plant Hormones.

The mode of action of 1-naphthylphthalamic acid (NPA) to induce conspicuous local stem swelling in the area of its application to the growing internode in intact Bryophyllum calycinum was studied based on the aspects of histological observation and comprehensive analyses of plant hormones. Histological analyses revealed that NPA induced an increase in cell size and numerous cell divisions in the cortex and pith, respectively, compared to untreated stem. In the area of NPA application, vascular tissues had significantly wider cambial zones consisting of 5-6 cell layers, whereas phloem and xylem seemed not to be affected. This indicates that stem swelling in the area of NPA application is caused by stimulation of cell division and cell enlargement mainly in the cambial zone, cortex, and pith. Comprehensive analyses of plant hormones revealed that NPA substantially increased endogenous levels of indole-3-acetic acid (IAA) in the swelling area. NPA also increased endogenous levels of cytokinins, jasmonic acid, and its precursor, 12-oxo-phytodienoic acid, but did not increase abscisic acid and gibberellin levels. It was shown, using radiolabeled 14C-IAA, that NPA applied to the middle of internode segments had little effect on polar auxin transport, while 2,3,5-triiodobenzoic acid substantially inhibited it. These results strongly suggest that NPA induces changes in endogenous levels of plant hormones, such as IAA, cytokinins, and jasmonic acid, and their hormonal crosstalk results in a conspicuous local stem swelling. The possible different mode of action of NPA from other polar auxin transport inhibitors in succulent plants is extensively discussed.

Changes in Polar Metabolites Content during Natural and Methyl-Jasmonate-Promoted Senescence of Ginkgo biloba Leaves.

The present study clarified changes in the contents of polar metabolites (amino acids, organic acids, saccharides, cyclitols, and phosphoric acid) in leaf senescence in Ginkgo biloba with or without the application of methyl jasmonate (JA-Me) in comparison with those in naturally senescent leaf blades and petioles. The contents of most amino acids and citric and malic acids were significantly higher in abaxially, and that of myo-inositol was lower in abaxially JA-Me-treated leaves than in adaxially JA-Me-treated and naturally senescent leaves. The levels of succinic and fumaric acids in leaves treated adaxially substantially high, but not in naturally senescent leaves. In contrast, sucrose, glucose, and fructose contents were much lower in leaf blades and petioles treated abaxially with JA-Me than those treated adaxially. The levels of these saccharides were also lower compared with those in naturally senescent leaves. Shikimic acid and quinic acid were present at high levels in leaf blades and petioles of G. biloba. In leaves naturally senescent, their levels were higher compared to green leaves. The shikimic acid content was also higher in the organs of naturally yellow leaves than in those treated with JA-Me. These results strongly suggest that JA-Me applied abaxially significantly enhanced processes of primary metabolism during senescence of G. biloba compared with those applied adaxially. The changes in polar metabolites in relation to natural senescence were also discussed.

Formation of the Secondary Abscission Zone Induced by the Interaction of Methyl Jasmonate and Auxin in Bryophyllum calycinum: Relevance to Auxin Status and Histology.

The interaction of methyl jasmonate (JA-Me) and indole-3-acetic acid (IAA) to induce the formation of the secondary abscission zone in the middle of internode segments of Bryophyllum calycinum was investigated in relation to auxin status and histology. When IAA at 0.1% (w/w, in lanolin) was applied to the segments, the formation of the secondary abscission zone at a few mm above the treatment in the apical direction was observed. On the contrary, IAA at 0.5% (w/w, in lanolin) did not induce the formation of the secondary abscission zone. JA-Me at 0.5% (w/w, in lanolin) applied to the middle of internode segments kept in the normal (natural) or inverted positions also induced the formation of the secondary abscission zone below and above parts of the treatment. IAA at 0.5% applied to the cut surface of the upper part of the segments completely prevented the formation of the secondary abscission zone induced by JA-Me. Simultaneous application of IAA 0.5% with JA-Me 0.5% in the middle part of the internode segments induced the formation of the secondary abscission zone at 10 mm to 12 mm above the treatment. Histological analyses indicated that the formation of the secondary abscission zone was characterized by the presence of newly synthesized cell plates that resulted from periclinal cell division within one layer of mother cells in stems. The effects of IAA (0.1%) and JA-Me (0.5%) on the formation of the secondary abscission zone were histologically similar. Comprehensive analyses of plant hormones revealed that the balance of the endogenous levels of IAA in both sides adjacent to the abscission zone was significantly disturbed when the secondary abscission formation was induced by the application of IAA. These results strongly suggest that an auxin gradient is important in the formation of the secondary abscission zone in the internode segments of B. calycinum, and IAA gradient results from polar IAA transport from the application site. IAA is important in the regulation of formation of the secondary abscission zone induced by JA-Me. Further possible mechanisms of the formation of the secondary abscission zone in the internode segments of B. calycinum are also discussed in the interaction of JA-Me and IAA.

Hormonal regulation of gummosis and composition of gums from bulbs of hyacinth (Hyacinthus orientalis).

Hyacinth (Hyacinthus orientalis) bulbs infected by Fusarium oxysporum showed the symptoms of gummosis. The purpose of this study was to clarify the hormonal regulation of gummosis and composition of gums from hyacinth bulbs. The application of ethephon (2-chloroethylphosphonic acid), an ethylene-releasing compound, at 2% (w/w, in lanolin) induced gummosis in hyacinth bulbs. Methyl jasmonate (JA-Me) at 1.5% (w/w, in lanolin) induced gummosis as well. Simultaneous application of JA-Me and ethephon further enhanced gummosis. Molecular mass distribution of hyacinth gums analyzed by gel permeation chromatography indicated that the gums were mainly homogenous polysaccharides with an average molecular weight of ca. 30kDa. Analysis of the sugar composition of the gums after hydrolysis revealed that the majority were arabinose (ca. 35%) and galactose (ca. 40%) together with small amounts of fucose, rhamnose and uronic acids (ca. 5%, respectively), suggesting that the gums are pectic arabinogalactans. These results indicate that jasmonates (JAs) interact with ethylene to stimulate sugar metabolism, producing pectic arabinogalactans, and vice versa, leading to gummosis. These findings, together with those from our previous studies in tulips (Tulipa gesneriana) and grape hyacinth (Muscari armeniacum), revealed that sugar metabolism and hormonal regulation relating to gummosis are different among species of bulbous plants.

Gummosis in grape hyacinth (Muscari armeniacum) bulbs: hormonal regulation and chemical composition of gums.

The purpose of this study was to investigate the hormonal regulation of gummosis in grape hyacinth (Muscari armeniacum) bulbs, focusing especially on the chemical composition of the gums. The application of ethephon (2-chloroethylphosphonic acid), an ethylene-releasing compound, at 1% and 2% (w/w) in lanolin as well as ethylene induced gummosis in the bulbs within several days. Methyl jasmonate (JA-Me, 0.1-2% in lanolin) alone had no effect on gummosis. However, simultaneous application of JA-Me and ethephon led to extreme stimulation of ethephon-induced gummosis. Ethephon-induced gummosis in the bulbs depended on the maturation stage of the bulbs, increasing from April to July, but decreasing from August to September. Regardless of the presence of JA-Me, the application of ethephon to the inflorescence axis of grape hyacinths did not induce gummosis. Gel permeation chromatography analysis revealed that gums were homogenous polysaccharides with an average molecular mass of ca. 8.3 kDa. Analysis of the sugar composition of the gums after hydrolysis revealed that the molar ratio of Rha:Ara:Gal:GalA:GlcA was 25:10:40:7:15. These results suggest that principal factors of gummosis as well as the chemical composition of gums differ between species of bulbous plants.

Jasmonates are essential factors inducing gummosis in tulips: mode of action of jasmonates focusing on sugar metabolism.

The purpose of this study was to know the mechanism of jasmonates to induce gummosis in tulip (Tulipa gesneriana L. cv. Apeldoorn) shoots, especially on the focus of sugar metabolism. Gummosis in the first internode of tulip plants was induced by the application of methyl jasmonate (JA-Me, 1% w/w in lanolin) and jasmonic acid (JA, 1% w/w in lanolin) 5 days after application and strongly stimulated by the simultaneous application of ethylene-releasing compound, ethephon (2-chloroethylphosphonic acid, 1% w/w in lanolin), although ethephon alone had little effect. JA-Me stimulated ethylene production of the first internodes of tulips, ethylene production increasing up to more than 5 times at day 1 and day 3 after the application. On the other hand, application of ethephon did not increase endogenous levels of jasmonates in tulip stems. Analysis of composition of tulip gums revealed that they were consisted of glucuronoarabinoxylan with an average molecular weight of ca. 700 kDa. JA-Me strongly decreased the total amount of soluble sugars in tulip stems even in 1 day after application, being ca. 50% of initial values 5 days after application, but ethephon did not. However, both JA-Me and ethephon had almost no effect on the neutral sugar compositions of soluble sugars mainly consisting of glucose, mannose and xylose in ratio of 20:2:1 and traces of arabinose. Both JA-Me and ethephon applied exogenously stimulated senescence of tulip shoots shown by the loss of chlorophyll. These results strongly suggest that the essential factor of gummosis in tulips is jasmonates affecting the sugar metabolism in tulip shoots. The mode of action of jasmonates to induce gummosis of tulip shoots is discussed in relation to ethylene production, sugar metabolism and senescence.

Identification of jasmonic acid and its methyl ester as gum-inducing factors in tulips.

The purpose of this study was to identify endogenous factors that induce gummosis and to show their role in gummosis in tulip (Tulipa gesneriana L. cv. Apeldoorn) stems. Using procedures to detect endogenous factors that induce gum in the stem of tulips, jasmonic acid (JA) and methyl jasmonate (JA-Me) were successfully identified using gas-liquid chromatography-mass spectrometry. Total amounts of JA and JA-Me designated as jasmonates in tulip stems were also estimated at about 70-80 ng/g fresh weight, using deuterium-labeled jasmonates as internal standards. The application of JA and JA-Me as lanolin pastes substantially induced gums in tulip stems with ethylene production. The application of ethephon, an ethylene-generating compound, however, induced no gummosis although it slightly affected jasmonate content in tulip stems. These results strongly suggest that JA and JA-Me are endogenous factors that induce gummosis in tulip stems.