Gianfranca Rossi

Effect of benzyladenine on the development of plastids and microbodies in excised watermelon cotyledons.

Excised watermelon (Citrullus vulgaris Schrad.) cotyledons were grown in the dark in the presence of 0.1 mM benzyladenine (BA). Under these conditions reserve breakdown and organelle differentiation progress very slowly. Treatment with BA accelerates, breakdown of reserves and stimulates development of organelles. Electron micrographs of cells from treated cotyledons show a larger number of plastids with a more developed inner membrane system. The levels of plastid pigments and enzymes are increased while starch content is reduced. Glyoxysomal enzyme levels are increased by BA during the first three days of development and their decline is accelerated thereafter. Also the activity of hydroxypyruvate reductase (EC 1.1.1.81.), a peroxisomal enzyme, is increased, but this increase is not followed by a decay phase. In water controls, hydroxypyruvate reductase bands together with glyoxysomal enzymes after equilibrium centrifugation in a sucrose gradient. In treated cotyledons the equilibrium position of glyoxysomal enzymes is uchanged while that of hydroxypyruvate reductase is shifted to a lower density.

Development of male and female flower in Asparagus officinalis. Search for point of transition from hermaphroditic to unisexual developmental pathway

Asparagus officinalis is a dioecious plant. The flowers start to develop as hermaphrodites and later become unisexual. In female flowers the stamens degenerate, while in male flowers the ovary stops growing without degenerating. We have examined young asparagus flowers using SEM and optical microscopy in order to determine the exact moment of transition from hermaphroditic to unisexual development. We defined 13 stages of development, starting from flower primordia up to completely mature flowers and labelled them with numbers from -6 to 7. The first five stages are fully hermaphroditic: a difference between sexes becomes visible at stage — 1 when the style begins to develop in female flowers. Degeneration of stamens in female flowers starts somewhat later. At the stage of transition, some differences between sexes also appear in the bidimensional polypeptide pattern of flowers. RNase activity shows a distinct peak at this stage (in female flowers only), probably related to stamen degeneration.

Benzyladenine induces the appearance of LHCP-mRNA and of the relevant protein in dark-grown excised watermelon cotyledons.

Cotyledons were excised from imbibed watermelon seeds, grown for 4 days in darkness on water or 10 μM benzyladenine (BA) and then tested for the presence of the light-harvesting chlorophyll a/b protein (LHCP) and its mRNA. LHCP was assayed immunologically by western blotting of SDS gels: the protein was present in plastids, but it was not recovered with the thylakoid fraction. Antibodies directed against LHCP precipitated a 32 kDa polypeptide from translation products of poly(A) RNA of cotyledons only if these had been grown on BA. Taken together the data suggest that in absence of light cytokinins are necessary for the maintenance of a detectable level of LHCP-mRNA as well as for synthesis of the protein.

Variations in carbohydrate and lipid content and in osmotic potential of watermelon cotyledons treated with benzyladenine

Abstract Benzyladenine (BA)-stimulated growth of excised watermelon cotyledons is accompanied by a stimulation of the conversion of reserve lipids to soluble sugars. After the 3rd day of development most of the carbohydrate is found as starch in the controls and as soluble sugars in treated cotyledons. As a result of these two effects the amount of soluble sugars per cotyledon is always higher in BA-treated cotyledons. Osmotic potential (measured as variation in fresh weight in presence of mannitol solutions or as osmolarity of cell sap) is, however, always lower in BA-treated cotyledons. These data suggest that BA increases the capacity of water uptake not only by build-up of solutes, but also by a direct influence on cell walls.

Evidence for two classes of responses of watermelon cotyledons to benzyladenine

Abstract Watermelon ( Citrullus vulgaris Schrad.) cotyledons were excised after 24 h of imbibition and cultured for 5 days on filter paper saturated with benzyladenine (BA) solution at concentrations ranging from 10 −8 to 10 −4 M. Alternatively the cotyledons were exposed to 10 −4 M BA for 5 min or 4 h and then cultured in water. We examined the effects of these treatments on several parameters: growth, carotenoid level, lipid disappearance and representative enzymes for plastids, mitochondria and microbodies. Two groups of responses to BA could be distinguished. Growth and mitochondrial enzyme levels showed a dose-response curve with maximal stimulation at the highest concentration tested (10 −4 M). A 5-min treatment with 10 −4 M BA yielded a very small response. Other responses (increase in carotenoids and in marker enzymes for plastids and microbodies) showed a sharp optimum at 10 −5 M concentration in the dose-response curve. A 5-min treatment with 10 −4 M BA was sufficient to yield a large response. Irradiation of the cotyledons with white light mimicked several responses to BA belonging to the second group, but none belonging to the first group.

I Lomasomi: Loro Probabili Rapporti con la Crescita per Distensione della Parete Cellulare

Abstract « Lomasomes »: their probable role in the expansion growth of the cell wall. — It was previously reported that lomasomes are present in higher plant cells. In a preliminary comunication Authors described the morfological characteristics of the lomasomes and their position in the cell. It was shown that lomasomes vary in organization which appears to be granular or vescicular and in distribution along the cell wall. Both these characters seemed directly related with the level of cellular differentiation. In this paper it has been reported that the number of lomasomes per cell was sharply decreased when Avena Coleoptiles were illuminated per 3 hours. During this period of time also the growth of the Coleoptiles was inhibited about 8%. These data seem favorable to the hypothesis that lomasomes are tigtly involved in the mechanisms controlling the expansion growth of the cell wall.

Benzyladenine stimulates the development of mitochondria in watermelon cotyledons

Abstract The development of mitochondria was followed for 5 days in watermelon cotyledons excised from the seedling 24 h after soaking and cultured in the dark on filter paper saturated with water or with a 10−4 M benzyladenine (BA) solution. Treatment with BA increases the total mass of mitochondria as shown by electron microscopy and by measurements of mitochondrial protein after density gradient centrifugation. Enzyme levels are increased to the same extent as total mitochondrial protein so that specific activities in mitochondria from treated cotyledons are the same as those in mitochondria from water controls. Only the specific activity of cytochrome oxidase and the specific content of cytochrome a 3 are higher in mitochondria of BA-treated cotyledons. Mitochondria isolated from cotyledons treated with BA seem to be in a better state of preservation than those isolated from water controls.

Osservazioni Sulla Morfologia Dei Lomasomi

Abstract Morphological organization of the lomasomes. — From a study on morphological organization of the lomasomes, in Avena coleoptile cells, it has been reported: 1) lomasomes form is quite variable, nevertheless, usually they resemble to a cone having a flattened tip and the base line against the cellular wall. 2) the external border of the lomasoma towards the cytoplasm is represented by the plasma membrane; such external profile results very sinuous and deep invaginations are present. 3) the internal structure is characterized by the presence of an unstructurated matrix containing spherical vescicles, flattened vescicles and tubules all showing an higher density to the electron radiations than the cytoplasmic matrix. Both vescicles and tubules are delimited by a single membrane.

Expansion growth is not the main driving force for cytokinin-promoted development of cotyledons

Abstract Excised watermelon cotyledons were treated with benzyladenine (BA) or fusicoccin (FC), both at 10 −5 M concentration. The effects of the two growth regulators were compared by monitoring the time course of the following parameters: growth (increase in fresh wt.), carotenoid and lipid content and the activities of cytochrome oxidase and hydroxypyruvate reductase (HPR). The treatment with FC induces a large growth response, of the same order of magnitude as that observed with BA. All other developmental responses that are strongly stimulated by BA are little or not-at-all affected by FC. These results indicate that growth by itself is not sufficient to entrain the other developmental responses induced by BA. On the other hand BA can partially enhance carotenoid content and HPR activity even when expansion growth is completely suppressed by osmotic stress.

Effect of benzyladenine on proteolytic activity of excised watermelon cotyledons

Abstract Excised watermelon cotyledons were grown in presence or absence of 10−4 M benzyladenine (BA). The time course of proteolytic activity was assayed with hemoglobin and azocasein. Total protease activity shows a large peak at pH 4.4 (with hemoglobin) or 5.4 (with azocasein) and a smaller one at pH 8.5. BA does not enhance proteolytic activity. On the other hand electron microscopical data show an accelerated breakdown of protein bodies in BA-treated cotyledons. BA increases autolytic activity (release of free aminoacids without exogenous substrate) in a 17 500 × g pellet and in a band that sediments to equilibrium at 1.28–1.30 g/cm3 on a sucrose gradient. This band is most probably formed by protein body fragments. It is likely that BA enhances the action of proteases that are sequestered inside protein bodies and are inaccessible to exogenous substrates.