A morphometric study on the effects of ethylene treatment in promoting abscission of tobacco flower pedicels.

The ultrastructural changes observed in ethylene-induced abscission of tobacco flower pedicels (Nicotiana tabacum L. ;Little Turkish') were studied by the techniques of morphometric analysis. The surface area of the membranes, relative volume of the organelles, and the number of organelles were determined for both ethylene-treated and control cells. In pedicels exposed to ethylene for 4.5 to 5 hours, abscission was evident within the separation zone. The most significant change in cell structure was observed in the surface area of the rough endoplasmic reticulum which more than doubled with ethylene treatment of the tissue.

Peroxidases in Tobacco Abscission Zone Tissue: II. Time Course Studies of Peroxidase Activity during Ethylene-induced Abscission.

Ethylene-induced abscission in flower pedicels of Nicotiana tabacum L. cv. Little Turkish causes a progressive increase in peroxidase activity during the first 4 hours of a 5-hour time course ethylene treatment period, with decrease in peroxidase activity occurring between 4 hours and 5 hours, when the supernatant extracts of abscission zone segments are tested spectrophotometrically for peroxidase activity, using guaiacol and hydrogen peroxide. Nonethylene-treated tissue has a much lower level of peroxidase activity over the same time course period. In ethylene-treated tissue the decline in break-strength correlates with the beginning of increase in peroxidase activity (3 hours). When the abscission zone area of the pedicel is further divided into proximal, abscission zone, and distal portions, respectively, the ethylene-treated tissue has the highest peroxidase activity in the abscission zone portion, with the maximum peak occurring at 4 hours and decreasing between 4 hours and 5 hours. Acrylamide gel electrophoresis of enzyme breis from ethylene-treated aand nonethylene-treated plants reveals that no new peroxidase isozymes are formed in response to ethylene, indicating an increase in the amount of one or in both of the two already existing isozyme banding patterns. The measurement of protein in the proximal, abscission zone, and distal segments, over a 5-hour ethylene treatment period, indicates that it is being translocated in a distal to proximal direction in the abscission zone pedicel. The possible participatory role for peroxidase in ethylene-induced tobacco flower pedicel abscission are discussed.

Fine structure of abscission zones : IV. Effect of ethylene on the ultrastructure of abscission cells of tobacco flower pedicels.

The effect of ethylene on abscission of flower pedicels of tobacco plants has been investigated. For the first 2 h of exposure to C2H4, the pedicels bend rather than break in response to applied force, but after 2.5 h exposure they break at the abscission zone under an applied force of 40 g. The break strength of the abscission zone decreases exponentially with time to 5 g at 5 h after beginning of the C2H4 treatment. An examination of the tissue at the fine structural level 2 h after exposure to C2H4 reveals the accumulation of rough endoplasmic reticulum (RER) in the abscission cells. Rough ER becomes increasingly abundant by 3-5 h exposure of the tissue to C2H4. There is approximately a 30 fold increase in RER by 5 h of exposure, as compared to untreated tissue.Loss in the integrity of the membranes of microbodies occurs by 5 h exposure of the tissue to C2H4. As cell wall degradation proceeds, fibrous material, vesicular structures, and electron dense bodies-the latter often appearing striated-develop in the disintegrating wall. Little change is seen in the structure of nuclei, mitochondria, chloroplasts and in the crystalloid cores of microbodies during the first 5 h of exposure of the tissue to C2H4. However, disorganization of cytoplasmic components does occur in cells where cell wall breakdown is at an advanced stage.

Regulation of Auxin Levels in Coleus blumei by Ethylene.

An investigation of the effects of ethylene pretreatment on several facets of auxin metabolism in Coleus blumei Benth "Scarlet Rainbow" revealed a number of changes presumably induced by the gas. Transport of indoleacetic acid-1-(14)C in excised segments of the uppermost internode was inhibited by about 50%. Decarboxylation of indoleacetic acid-1-(14)C by enzyme breis was not affected by the pretreatment. Levels of extractable native auxin in upper leaf and apical bud tissue of the pretreated plants were approximately one-half of those present in untreated plants. The rate of formation of auxin from tryptophan by enzyme breis from pretreated plants was approximately one-half that occurring in incubation mixtures containing the enzyme system from untreated plants. The conjugation of indoleacetic acid-1-(14)C in a form characterized chromatographically as indoleacetylaspartic acid was increased 2-fold in the upper stem region of plants pretreated with ethylene.

Fine structure of abscission zones : II. Cell-wall changes in abscising pedicels of tobacco and tomato flowers.

The fine structure of the abscission zones of Lycopersicon esculentum and Nicotiana tabacum flower pedicels was studied, with special reference to structural changes in the walls of cells during the abscission process. The separation of cells appeared to be initiated primarily in the middle-lamella region of the cell walls. Disintegration of the primary wall, which usually followed breakdown of the middlelamella region, also occurred concurrently with the lysis of the middle-lamella region. During cell-wall degradation, the walls appeared to swell and became highly flexible. The walls of at least some cells in the zone of separation invaginated during the advanced stages of cell-wall disintegration, and ultimately collapsed. Cell-wall changes in abscising pedicels are almost identical to those which occur in abscising cotton and Coleus leaves, as described by BORNMAN (1967).

Fine structure of abscission zones : III. Cytoplasmic changes in abscising pedicels of tobacco and tomato flowers.

Changes in the cells comprising the abscission zones of Lycopersicon esculentum and Nicotiana tabacum flower pedicels were observed, apparently associated with the separation of the flowers from the plants. Special attention is devoted to changes which occurred in the fine structure of intracellular components of the abscission cells. Microbodies with crystalloid cores in cells comprising the abscission tissue of the pedicels were observed to undergo structural changes. These changes occurred prior to the cell-wall degradation which resulted in cell separation. At the time of cell separation, the number of microbodies present in the abscission cells was greatly diminished. The changes in the fine structure of the crystalloid core may represent the release of latent enzymes into an active form which then induces cell separation in the abscission tissue. Other structures in the cells in which changes occurred are the endoplasmic reticulum (ER) and the nucleolus. Prior to, and during cell-wall disintegration, an abundance of rough ER was observed within the abscission cells. The occurrence of rough ER may represent an increase in the synthesis of protein for export from the cells of the abscission region and/or for the synthesis of certain enzymes associated with the degradation of cell walls in the abscission zone itself. In some abscission cells, prior to the cell-separation phase, nucleoli were observed which appeared as though they had segregated spatially. In such cases, the amorphous zone was the only nucleolar component observed. The possible significance of these observations is discussed.

Fine structure of abscission zones : I. Abscission zones of the pedicels of tobacco and tomato flowers at anthesis.

Electron micrographs of the zone of separation in flower pedicels of Lycopersicon esculentum and Nicotiana tabacum are presented with particular reference to the indentation of epidermal tissue in the abscission zone, subcellular organelles, and the cell wall. The indentation or groove which delineates the abscission zone extends some distance into the pedicel with branchings off the main groove. These branches are approximately 20 mµ in width while the main groove averages approximately 200 mµ in width. Invaginations of the plasmalemma are observed with considerable frequency. within these invaginations one observes a material of about the same density as the cell wall except that it is more fibrillar. Plasmodesmata are also observed, with considerable branching into middle lamellae of cells comprising the abscission zones. Microbodies with crystalloid cores appear with considerable frequency in cells of the abscission zone. The crystalloids appear to be cubical in shape and are composed of parallel sheets of osmiophilic material. The sheets average about 6 mµ in thickness and are spaced at 4 mµ intervals. The microbodies with crystalloid cores are observed to be characteristically of two size groupings. In tobacco the microbodies average 900 mµ and 1,500 mµ in profile. In tomato they average 300 mµ and 500 mµ. Chloroplasts contain a granular component which is membrane-enclosed. The component is large in comparison with the plastid in which it occurs, averaging 1.2-1.4 µ in diameter in chloroplasts ranging from 1.6 µ to 2.2 µ in diameter. The inner membrane of the chloroplast is highly invaginated, and DNA- and phytoferritin-like materials are observed within the plastids. Microtubules with an average diameter of 20 mµ are observed adjacent and parallel to the plasmalemma, primarily in the corners of the cells. Micrographs of other normally occurring cytoplasmic inclusions are also presented.

Metabolism of tryptophan in petioles of coleus.

Auxin precursors retard abscission when applied to debladed petioles of Coleus blumei Benth. The d and l forms of tryptophan are equally effective in retarding abscission. Tryptamine is more effective than is tryptophan. Both compounds apparently are converted to auxin through an aldehyde intermediate. The evidence presented suggests that a major pathway of tryptophan metabolism proceeds through tryptamine, as can be demonstrated by the use of amine oxidase inhibitors in the petiole tissue. Cell free preparations of the tissues metabolize tryptophan-1-(14)C with the release of carbon dioxide. The rate of tryptophan mtabolism in abscission tissue is 5 times that in distal petiole tissue. Radioactivity is associated with basic indole conversion products as well as with neutral and acidic fractions. The radioactivity is most concentrated in the neutral fraction. The results indicate that the Coleus petiole itself is capable of producing auxin.