Jean-Pierre Gaudillère

Phosphoenolpyruvate carboxylase during grape berry development: protein level, enzyme activity and regulation

The protein level and regulation of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31, involved in malic acid synthesis) was studied during the fruit development of two grape (Vitis vinifera L.) varieties, ‘Cabernet Sauvignon’ and ‘Gora Chirine’, with berries of normal and low organic acid content, respectively. The protein level and in vitro activity were higher in the low-acid variety than in the normal-acid variety for most stages. In vivo PEPC activity, measured using 14 CO2 labelling, was significantly higher in the low-acid variety than in the normal-acid variety about 1 week before and 1 week after veraison (the day which corresponds to the onset of ripening). However, partitioning into malate was the same for both varieties. Antibodies raised against the N-terminal part of SorghumPEPC recognised the grape berry PEPC, indicating the presence of the consensus phosphorylation site involved in PEPC regulation. PEPC phosphorylation status was estimated by studying sensitivity to pH and malate. Grape berry PEPC appeared more sensitive to low pH and malate during ripening (IC50 malate, 0.2–0.7 mM) compared to during the earlier stages of development (IC50 malate, 1.2–2 mM) for both varieties. Therefore, in the normal-acid variety, PEPC seems to participate in controlling malic acid accumulation but does not seem to control the differences in malic acid concentration observed between the two varieties.

Vigour and non-structural carbohydrates in young prune trees

Abstract To determine if differences in vegetative vigour are related to carbon partitioning in trees, three rootstck clones (i.e. Marianna GF8.1, Ishtara® and St. Julien Pixy 2879), inducing vigorous or dwarfing graft combinations with two Prunus domestica L. scion genotypes, were studied. Three years after planting in the orchard, just before bud burst, partitioning of dry matter and carbohydrate content were determined in the different parts of the tree. Dry matter, starch and soluble carbohydrate distribution in Spring were slightly affected by the rootstock. During Year 4 after planting, sorbitol, soluble sugar and starch content were determined monthly in the trunk wood. The relationship between vegetative vigour and carbohydrate content depended on the type of carbohydrate examined and on the season. The kinetics of starch mobilization were modified with the dwarfing rootstock when compared with the other rootstocks. Differences in the kinetics of starch accumulation were observed between the two cultivars which showed the same vigour. The level of carbohydrate reserves in perennial parts did not appear to be related to rootstock-induced modification of vegetative vigour.

Modifications of Etioplasts in Cotyledons during Prolonged Dark Growth of Sugar Beet Seedlings' Identification of Etiolation-Related Plastidial Aminopeptidase Activities

We studied the effects of prolonged dark growth on proplastids and etioplasts in cotyledons of sugar beet (Beta vulgaris L.) seedlings. Differentiation of proplastids into etioplasts occurred between d 4 and d 6 after imbibition, with the typical characteristics of increased synthesis of plastidial proteins, protein and carotenoid accumulation, size increase, development of plastid membranes and of the prolamellar body, and increase of the greening capacity. However, this situation of efficient greening capacity was short-lived. The greening capacity started to decline from d 6 after imbibition. This decline was due in part to reserve depletion and glucose limitation and also to irreversible damage to plastids. Indeed, electron microscopy observations in situ showed some signs of plastidial damage, such as accumulation of plastoglobuli and membrane alterations. The biochemical characterization of purified plastids also showed a decrease of proteins per plastid. Aminopeptidase activities, and to a lesser extent, neutral endopeptidase activities, were found to increase in plastids during this degenerative process. We identified two plastidial aminopeptidases showing a sharp increase of activity at the onset of the degenerative process. One of them, an alanyl aminopeptidase, was shown to be inactivated by exposure to light or addition of exogenous glucose, thus confirming the relationship with prolonged dark growth and indicating a relationship with glucose limitation.

Carbon and nitrogen reserves in prune tree shoots: effect of training system

Abstract To determine if differences in the pruning and training system adopted could modify the concentration of C and N reserves in trees, a Prunus domestica L. scion genotype grafted on Ishtara® rootstock clone was pruned and trained in two different ways. One system induced vigorous vertical shoots, while the other maintained less vigorous bent shoots. Two years after planting in the orchard, just after leaf fall, non-structural carbohydrates, total free amino-acid and soluble protein content were determined in the wood and bark of 1-year-old shoots. The mean concentration and distribution of carbohydrate reserves in wood or bark appeared to be only slightly influenced by the training system. The mean concentration and distribution of soluble proteins was the same for both training systems. The major amino acids in wood and bark were arginine and asparagine. The mean concentration of total amino acids was higher in shoot bark for the bent-training system than for the vertical-training system. However, no similar difference appeared for total amino acids in shoot wood. Therefore, C and N reserves showed little modification with a change in the pruning and training system, under the present experimental conditions.

Purification and Characterization of a Novel Aminopeptidase, Plastidial Alanine-Aminopeptidase, from the Cotyledons of Etiolated Sugar Beet Seedlings

During prolonged dark growth of sugar beet (Beta vulgaris L.) seedlings, etioplasts, rapidly after the proplastid-etioplast transition, undergo a degenerative process characterized by ultrastructural modifications, protein loss, and the decrease of carotenoid and chlorophyll accumulation upon illumination. Two plastidial aminopeptidase activities were identified as early markers of this degenerative process (A. El Amrani, I. Couee, J.-P. Carde, J.-P. Gaudillere, P. Raymond [1994] Plant Physiology 106: 1555–1565). The present study focuses on one of these markers and describes the purification to homogeneity and characterization of plastidial alanine-aminopeptidase. This novel aminopeptidase was found to be a metallo-type naphthylamidase particularly active with alanyl, arginyl, and leucyl substrates. Its plastidial location was confirmed by immunofluorescence with polyclonal antibodies against the purified enzyme. Its physico-chemical and enzymic properties are discussed with respect to other higher plant aminopeptidases and to its potential functions during prolonged dark growth.

Accumulation of plastidial alanine-aminopeptidase in relation to plastid damage in cotyledons of dark-grown sugar beet seedlings

During prolonged dark-growth of sugar beet (Beta vulgaris L.) seedlings, the etioplasts of cotyledons undergo a degenerative process shortly after the proplastid to etioplast transition. Two plastidial aminopeptidase activities were previously identified as early markers of this degenerative process which results in a decrease of greening capacity. One of these activities was shown to be due to one enzyme, plastidial alanine-aminopeptidase, which was purified to homogeneity and characterized. In the present study, the effects of dark development, light, and exogenous glucose on enzyme activity and protein level of plastidial alanine-aminopeptidase were investigated. Changes of enzyme activity during prolonged dark-growth were found to be correlated with changes of protein level. Significant increases of both activity and protein levels occurred at the onset of the degenerative process affecting etioplasts. By contrast, the activity and protein levels of plastidial alanine-aminopeptidase were negatively affected by light and by exogenous glucose, thus suggesting a type of expression that is related to prolonged dark-growth and etioplast ageing.

Analyse qualitative des sucres solubles de la sève xylémienne de Acer platanoides

Summary The qualitative analysis of soluble sugars contained in Acer platanoides xylem sap pointed out, at first, the very limited number of molecular species of this sap. Sucrose was the main sugar. Glucose and fructose were well detected only in autumn and In winter except during the frost period. We found only slight traces of sorbitol, meso-inositol, and oligosaccharides. An unusual molecule was detected in significant quantities. The mass spectro- metric analysis of this molecule allowed us to consider it as a mono-O-methyl- inosrtol, probably quebrachitol. Its presence seemed to be specially pronounced in December, just after the first cold days in autumn.

Variations saisonnières des glucides de réserve chez le prunier: relations avec la vigueur

Summary Storage of non—structural carbohydrates was investigated in the wood of prune trees (Prunus domestica L.). The vegetative vigour of the variety was modified by grafting on different rootstocks, and by using special training systems (pruning and branche bending). Non—structural carbohydrate contents were monitored monthly in the trunk, and at leaf fall in one-year-old shoots. No relationships was observed between vegetative vigour and either carbohydrate content in the perennial parts, or carbohydrate content in the one-year-old shoots. Carbon partitioning between carbohydrate storage and vegetative growth seems to be highly regulated.