Gilles Pétel

Dormancy in vegetative buds of peach : relation between carbohydrate absorption potentials and carbohydrate concentration in the bud during dormancy and its release

The absorption of sucrose and sorbitol by the bud and the stem of Prunus persica was evaluated during the rest period. The utilization of p-chloromercuribenzenesulfonic acid (PCMBS), an inhibitor of sucrose transporter, allowed estimation of the active absorption of sucrose in different isolated organs (bud, buds underlying tissues and stem). It was assumed that the sink capacities of tissues depended on their potential to absorb carbohydrates by active transport. Endogenous carbohydrate levels in the tissues were also measured. In the bud, the qualitative and quantitative distribution of carbohydrates could be related to the absorption potential of the bud. During dormancy, the bud exhibited a low absorption potential and increased its sucrose concentration by starch hydrolysis. Soluble sugars accumulated during winter. During dormancy release, the bud was able to absorb carbohydrates allowing carbon storage. The data presented led to the proposal of a scheme of events occurring during bud dormancy and its release.

Differential expression of four members of the H+-ATPase gene family during dormancy of vegetative buds of peach trees

Abstract. Vegetative-bud dormancy in peach (Prunus persica L. Batsch) trees is known to be correlated, at least partially, with properties of the underlying bud tissues during winter. Variations in the activity and amount of plasma-membrane H+-ATPase were observed. A full-length cDNA, PPA2 (Prunus persica H+-ATPase 2) and three partial cDNAs (PPA1, PPA3 and PPA4) for the plasma-membrane H+-ATPase from peach trees were isolated by reverse transcription (RT)-coupled rapid amplification of cDNA ends (RACE) polymerase chain reaction (PCR). The accumulation of plasma membrane H+-ATPase transcripts was then studied in vegetative buds during dormancy and breaking of dormancy. Competitive RT-PCR analysis revealed that, during dormancy, the plasma membrane H+-ATPase transcripts were higher in the tissues underlying the buds than in the buds themselves. After dormancy release, the level of PPA1, 2, 3 mRNA increased, whereas the level of PPA4 decreased in the buds. When trees were kept in a greenhouse (i.e. sheltered from chilling), no accumulation of PPA mRNA could be detected. These results suggest that there is a differential accumulation of H+-ATPase mRNA between the bud and the underlying bud tissues during dormancy, and that chilling could act as a decisive factor.

Plasma membrane H+-ATPase, succinate and isocitrate dehydrogenases activities of vessel-associated cells in walnut trees

Summary In winter and spring, walnut trees exhibit variations of sugar content in the vascular sap. According to their location, the vessel-associated cells (VACs, also called contact cells) could control nutrient exchanges between the storage parenchyma and the xylem vessels. According to the literature, the recovery of sap (influx) occurs at the VAC/vessel interface via an H+/sugar symport that depends on the transmembrane pH gradient generated by the plasma membrane H+-ATPase. The aim of this study was to investigate the ATPase activity, using a perfusion technique that allows the use of several effectors: carbonyl cyanide m-chlorophenylhydrazone (CCCP) and fusicoccin (FC). During winter, the uncoupler CCCP revealed a low pH gradient between the xylem vessels and the vessel-associated cells. Under these conditions, FC, an activator of the H+-ATPase, had no effect on the pH of the perfusion solution, suggesting that the enzyme could be lightly active. In contrast, close to bud break, a high pH gradient was revealed by the use of CCCP, and an acidification of the perfusion solution was observed in the presence of FC. Moreover, cytochemical investigation showed high activity of two respiratory enzymes located in mitochondria: NAD-dependent isocitrate dehydrogenase and succinate dehydrogenase. The hypothesis is that in spring this high respiratory activity of VACs provides a consequent increase in available ATP that can be utilized by the plasma membrane H+-ATPase.

Localization in sucrose gradients of pyrophosphatase activities in the microsomal fraction of Jerusalem artichoke (Helianthus tuberosus L.) tubers

Summary Three distinct Mg 2+ -PPases have been detected in the microsomal vesicles from Jerusalem artichoke. Separation of the different membranes on a linear sucrose gradient showed that these activities were associated with tonoplast (d = 1.11-1.12), Golgi (d = 1.14-1.15) and plasma membrane (d = 1.17-1.18). PPase activity located in the plasmalemma enriched fractions seemed to be different from ATPase activity, considering the effects of the well known effectors of the plasmalemma ATPase type.

Photoperiod and temperature effect on growth of strawberry plant (Fragaria × ananassa Duch.): development of a morphological test to assess the dormancy induction

At the end of summer, the diminution of photoperiod and temperature cause a decrease of vegetative growth and the dormancy of strawberry plants. Although the decrease in vegetative growth can be measured morphologically, no test is able to evaluate the decrease in growth potential (i.e. during the dormancy induction) nor its possible influence on vegetative growth. On the one hand, to estimate this influence biometrically, we have correlated photoperiod and temperature decreases with the vegetative growth decrease of some strawberry cultivars observed in the field. Results have confirmed the major role of photoperiod, temperature and the effect of growth potential decrease on vegetative growth. Moreover, the results showed that the decrease of vegetative growth was an early event at the end of summer which depended upon strawberry cultivar. On the other hand, we have measured petiole length under standard climatic conditions in a growth chamber, after natural summer and autumnal exposures. Observations of strawberry plants under these conditions revealed a decrease of their growth potential which also depended upon strawberry cultivar. Results also confirmed the possible action of growth potential decrease on the vegetative growth at the end of summer. Consequently, the observation of strawberry plants under standard conditions can be used, as a test, to assess the exact moment of dormancy induction.

Saccharose and sorbitol transporters from plasmalemma membrane vesicles of peach tree leaves

The mechanisms of saccharose and sorbitol transport in Prunus persica leaves were investigated in plasma membrane vesicles purified by aqueous 2-phase partitioning and equilibrated in pH 7.5 buffer containing K+. The imposition of an artificial proton motive force energized an active uptake of both saccharose and sorbitol. The maximum uptake rate of saccharose was 2.5 times higher than that of sorbitol. Saccharose and sorbitol uptake exhibited saturation kinetics suggesting they were carrier-mediated. Apparent Km for the saccharose and the sorbitol uptake were 0.36 and 0.67 mM, respectively. Active absorption of saccharose was completely inhibited by a non-permeant thiol reagent, PCMBS, contrary to sorbitol absorption. These results suggested that saccharose and sorbitol were transported at least by two different carriers.

Effect of exogeneous methyl oleate on the time course of some parameters of Streptomyces hygroscopicus NRRL B-1865 culture

Addition of methyl oleate to a Streptomyces hygroscopicus NRRL B-1865 culture modified the metabolic properties of this strain. This addition decreased the pH of the medium, increased the valine uptake of the cells and reduced their consumption of glucose until the beginning of antibiotic biosynthesis, which was delayed. At the same time, an increase in growth (× 1.8) and a marked improvement in antibiotic production (× 20) could be observed. The use of labelled methyl oleate showed that methyl oleate was not a precursor of antibiotics produced by S. hygroscopicus NRRL B-1865. It is suggested that methyl oleate addition may cause some alteration in membrane permeability, inducing an increase in H+ extrusion and stimulating the accumulation of branched amino acids, known to be direct precursors of polyether antibiotics.

Determination of the growth potential of strawberry plants (Fragaria × ananassa Duch.) by morphological and nucleotide measurements in relation to chilling

Early strawberry fruit production requires that non-dormant strawberry plants be covered during winter. Two methods were used to study the breaking of dormancy of Elsanta strawberry plants following chilling at temperatures below 8 °C. First, a morphological test using the petiole lengths of plants transferred from a nursery to a growth chamber was used. Second, the nucleotide contents before and after incubation of samples in an adenosine solution were measured by bioluminescence. The results show that these two methods can reveal changes in the growth potential of strawberry plants, and, therefore, the release from dormancy. Key words: Dormancy, strawberry plant, nucleotide, growth potential, Fragaria

Study of H+-nutrient co-transport in peach-tree and the approach to their involvement in the expression of vegetative bud growth capability

Summary Sucrose and sorbitol are the predominant forms of photosynthetically reduced carbon transported in Prunus persica . The mechanisms of sucrose and sorbitol transport were investigated in plasma membrane vesicles isolated from peach tree leaves. The maximum uptake was obtained for sucrose (2.5 times higher than sorbitol uptake). Active absorption of sucrose was completely inhibited by a non-permeant thiol reagent, PCMBS, contrary to sorbitol absorption. These results suggested that sucrose and sorbitol were transported at least by two different carriers. The absorption of exogenous sucrose and sorbitol in the bud and the underlying territories of the bud and their distribution in the single node cuttings were evaluated during the vegetative rest period. The utilization of PCMBS, an inhibitor of sucrose transporter, allowed for the estimation of the active absorption of sucrose in different isolated territories. It was assumed that the sink capacities of tissues depend on their potentialities to absorb carbohydrates by H + /symport. The presence of unlabelled sucrose during PCMBS pretreatment decreased the inhibition exerted by PCMBS: it suggested that the inhibitor was bound specifically on the sucrose transporter. The «active» absorption related to the transporter could therefore be distinguished from the total absorption, including diffusion. The evolution of exogenous sucrose and sorbitol was evaluated during the vegetative rest period, in isolated territories and in single node cuttings. In natural conditions, three periods were disclosed: 1) from September to October, the bud, intrinsically able to grow, was inhibited by the stem; 2) from November to December the bud was in a state of deep dormancy, and it was intrinsically unable to absorb nutrients; and 3) January was characterized by the reacquisition of bud growth potential and bud burst. Concerning sorbitol, a similar evolution was observed but it seemed that sorbitol was principally utilized a few days before budburst.

Changes in Parameters of the Plasmalemma ATPase during Peach Vegetative Bud Dormancy

Plant dormancy and dormancy breaking depend, at least partially, on close relationships between buds and tissues underlying bud (bud stands). In Prunus persica, the dormancy was related to high nutrient absorption in bud stands linked to high plasmalemma ATPase (EC 3.6.1.3) activity. Two plasmalemma fractions was isolated from peach vegetative buds and bud stands using aqueous phase partitioning and ultracentrifugation. Results of markers enzyme assays indicated that both plasmalemma enriched fractions obtained were highly purified. During the dormancy period plasma membrane ATPase amount and activity were higher in bud stands than in buds. Moreover, assays performed at different temperatures (4, 18, 30 °C) indicated modifications of kinetic parameters (Km, Vm) in both tissues during dormancy release. In buds, from November to February, Km declined at 4°C and increased at 30 °C whereas no changes was measured at 18 °C and Vm increased at all temperature. In bud stands, no changes of Km was measured at 4 °C and 18 °C whereas an increase occurred at 30 °C and Vm decreased at all temperature. According to the results, it can be postulated that dormancy release in peach-tree could be related to modifications of plasma membrane ATPase properties, in buds and bud stands, during winter time.