Journal of Experimental Botany, Vol. 53, No. 379, pp. 2431-2439,
December 1, 2002
© 2002 Oxford University Press
Contribution of vegetative storage proteins to seasonal nitrogen variations in the young shoots of peach trees (Prunus persica L. Batsch)
Received 4 March 2002; Accepted 21 July 2002
1 INRA Unité Plantes et Systèmes de culture Horticoles, Domaine Saint Paul, Agroparc, 84914 Avignon cedex 9, France
2 INRA Unité de Génétique et d’Amélioration des Fruits et Légumes, Domaine Saint Maurice, Agroparc, 84914 Avignon cedex 9, France
3 To whom correspondence should be addressed. Fax: +33 4 32 72 24 32. E-mail: Gomez{at}avignon.inra.fr
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Abstract |
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Qualitative and quantitative variations in the level of two low molecular weight vegetative storage proteins (VSP 19 kDa and 16.5 kDa) in peach shoots were compared with annual variations in total nitrogen and total soluble proteins. Protein patterns were obtained by SDS–PAGE and silver staining on each of the 12 kinetic samples collected between October 1995 and November 1996. VSP 16.5 kDa and 19 kDa exhibited typical annual VSP variations in both parenchyma and phloem. In wood, VSP 16.5 kDa was only present in November. All N compounds tested were stored in the autumn and their levels fell in the spring. Parenchyma was the principal stem storage tissue for all N compounds tested, even if proteins were more often highly concentrated in phloem and even if wood was the major shoot constituent. In winter, the two VSP accounted for 13% of bark proteins and 11% of wood proteins. Their storage yield, given by the winter/summer (W/S) ratio was higher (18.5) than that of total proteins (4). Between August to March, i.e. during the storage phase, N fractions obtained from VSP (N3) and total soluble proteins minus VSP (N2) accounted, respectively, for only 3% and 21% of total N accumulation in the bark, the remainder being due to the fraction not extracted (N1). A marked drop in all N compound levels characterized the mobilization phase (March to April), particularly for N3 (–84% between March and April) which were mobilized slightly before other N compounds. Although N3 exhibited the best mobilization yield, it represented only 5% of the total N mobilized. So, in spite of a similarity between VSP and N annual variation patterns, there was no tight correlation between their contents in bark. N2 supplied a high proportion of the N used for spring regrowth (40%), but the larger share (55%) came from N1 which was probably made up of free amino acids. Very tight positive correlations have been observed between these two N fractions and the N status. The lower bark total N content measured in August (6.4 mg N g–1 DW) during the assimilation phase (April to August) was equal to the unavailable N fraction, and the bark N mobilization potential (between March and August) was estimated at 6.35 mg N g–1 DW. VSP did not quantitatively represent the main stored N pool. But, because of their high W/S ratio and their early remobilization, they seemed to play an important role in spring regrowth initiation.
Key words: Peach tree, SDS–PAGE, shoot, VSP.
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Introduction |
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In woody plants, spring growth, including flowering, depends on the remobilization of stored nitrogen (N). Some N is stored as proteins in perennial organs (Tromp, 1970). There is no assay available to identify proteins with a storage function, so the criteria developed by O’Kennedy and Titus (1979) is used to define the vegetative storage proteins (VSP): VSP disappear as growth is resumed and constitute the predominant proteins in dormant organs.
First studied in the apple tree (Tromp, 1970; Tromp and Ovaa, 1971; O’Kennedy and Titus, 1979), VSP in woody plants were then widely investigated in softwoods and hardwoods; see reviews by Stepien et al. (1994) and Rowland and Arora (1997). Shoots are considered to be the principal storage area for N (Tromp, 1970).
VSP respond to specific stimuli (photoperiod, chilling accumulation, rest-breaking treatment, and wounding) and may have a physiological role (cold hardiness) and/or a biochemical function (Rowland and Arora, 1997). Indeed, an accumulation of VSP may represent an early event in the initiation and development of vegetative dormancy in woody plants (Coleman et al., 1991). Ar far as is known, no specific enzyme activity has been established for an isolated VSP.
In apple trees, the amount of stored nitrogen correlates with the growth of the next shoot (Tromp and Ovaa, 1973; Titus and Kang, 1982). Tromp (1970) showed that prior to spring growth, there was a marked breakdown of protein, especially in bark. Trees may employ VSP strategically to obtain readily accessible nitrogen during budbreak (Wetzel and Greenwood, 1989). However, the contribution of VSP to the remobilization of stored N remains unclear; studies have shown considerable variability between species and dormant tissues. Indeed, in autumn and winter, the VSP count (expressed as a percentage of soluble proteins) in shoot bark is 15% in Douglas fir (Roberts et al., 1991), 25% in poplar (Langheinrich and Tischner, 1991), and up to 70% in poplar wood (Sauter et al., 1988). Moreover, because a decline in the VSP content is insufficient to explain the spring fall in the total protein content, other proteins may also be mobilized during foliation (Langheinrich, 1993).
Few studies have investigated VSP levels in the shoots of peach trees, except those of Arora (Arora et al., 1992, 1996; Arora and Wisniewski, 1994, 1996). Tree VSP levels have been connected with the photoperiod (16 and 19 kDa) or seasonal changes (60 kDa). However, quantitative data are unavailable on their function as an N reserve for the tree.
The first objective was to confirm the existence of VSP in peach trees and to quantify their tissue distribution in shoots. Secondly, the VSP impact was assessed as a shoot N reserve, by comparison with the total soluble proteins content and the total N content.
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Materials and methods |
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Experimental design
Experiments were conducted between October 1995 and November 1996 in a low density orchard (500 trees ha–1), located at the Montfavet INRA Centre in southern France. The open-centre peach trees (cv. Suncrest grafted on Prunus persicaxPrunus amygdalus cv. GF677) had been planted in 1982. Four trees, enjoying the same exposure to light and wind, were selected from one row. Three main branches were then chosen on each tree for their uniformity of diameter. During the experimental period these trees remained unpruned and unthinned for fruits and flowers. Other agricultural practices were those of a traditional commercial orchard: undisturbed bare soil, application of herbicide, and fertilization monitored by leaf analysis.
In 1996, bud-burst started on 19 March and full blossoming was observed on 25 March. The fruits were harvested between 26 August and 31 August. No shoot growth was observed after that date. The trees were bare by 15 November, both in 1995 and 1996. According to these observations, 12 series of samples were collected: on 4 October and 25 November in 1995, and on 25 January, 19 March, 24 April, 31 May, 27 June, 25 July, 26 August, 4 September, 18 October, and 21 November in 1996. At each date, one shoot that had started growing in 1995 was harvested from each of the 12 selected branches. A piece of wood, located about 3 cm above the shoot base (or insertion point), and about approximately 15 cm long, was cut and immediately stored in liquid nitrogen. The shoot diameter varied from 1 cm to 1.5 cm.
Sample processing
The samples were stored at –80 °C until freeze-drying. The two wounded extremities were then eliminated and the wood was debarked. The bark was separated into ‘phloem’ and ‘parenchyma’ fractions. The first fraction was obtained as a white powder by gently scraping off the cambium and phloem tissues. The second fraction consisted of cortical parenchyma, sclerenchyma fibres, and suberin; it was ground to a powder in a stainless steel Dangoumeau grinder under liquid nitrogen. The dry weight (DW) of each fraction was measured. The residual humidity of the powder (about 3%) was measured by redrying (75 °C for 24 h) a 500 mg aliquot at the time of each determination.
Total N and nitrate analysis
The total N content (mg g–1 DW) of each sample was measured on 5 mg of plant material powder using an automated CN analyser (Carlo Erba analyzer ANA1500, Thermo Finnigan, Les Ulis France) according to the ANCA-MS technique. The nitrate content of parenchyma and phloem (mg g–1 DW) was determined on water extracts of the dried materials in January and July in an autoanalyser by colorimetry of nitrite after reduction by cadmium (Aquatec 5400 analyser, Tecator, Höganäs Sweden).
Protein extraction and electrophoresis
The soluble protein content (mg g–1 DW) was measured in each sample. Proteins were extracted using the method developed by Faurobert (1997). This N fraction will henceforth referred to as ‘proteins’, even though membrane proteins were not extracted. The plant material powder (50, 100 and 180 mg DW, respectively, for phloem, parenchyma, and wood) was mixed with 3 ml extraction buffer (50 mM Tris–HCl, 5 mM DTT, 0.3% PEG, 20 mM DIECA, 50 mM ascorbic acid, and 40 mg of PVP, pH 7.5 at 4 °C). The mixture was centrifuged at 20 000 g for 15 min at 4 °C. Proteins in the supernatant were quantified using the Bio-Rad protein assay kit (Richmond, CA), which is based on the Bradford method (Bradford, 1976). The supernatant was collected and mixed with 4 vols of cold acetone containing 0.07% ß-mercaptoethanol, and incubated overnight at –20 °C. Proteins were pelleted by centrifugation at 1500 g for 10 min at 4 °C. The pellet was washed with cold acetone, dried at room temperature in a speed-vac, resuspended in Laemmli lysis buffer (Laemmli, 1970) to obtain 1 µg µl–1 protein solution, and then boiled for 5 min to denature the proteins.
SDS–PAGE was performed as described by Laemmli (1970) with a Protean II electrophoresis unit (Bio-Rad) using a 4% stacking gel and a 12% running acrylamide gel. Precisely 12.5 µg of proteins was loaded on each lane. The gels were silver-stained according to the technique described by Rabilloud et al. (1988).
Image analysis
Gels were digitalized with an Epson GT-12000 scanner at 300 dots per inch. The integrated intensity of protein bands was measured manually using Bio-Rad Melanie II software and then plotted relative to the total intensity of the proteins in the lane. By applying the approximation that the intensity of a band was correlated to its protein content (Langheinrich and Tischner, 1991; Noquet et al., 2001), it was possible to plot the annual kinetic of each band’s contribution to the total protein content. Bands with the annual variation characteristics of VSP (accumulation in autumn and winter, marked decrease in spring and very low content until the end of the summer) were defined as VSP.
Measurements
Based on the relative proportions of the three shoot tissues, it is possible to infer the composition of N compounds for an entire shoot and for bark throughout the year. The results concerned the annual kinetics of N, and the protein and VSP contents in shoot tissue (parenchyma, phloem and wood), the shoot and bark.
Protein-N and VSP-N were estimated by dividing the amount of protein by 6.25 (Côté and Dawson, 1986). Bark total N was fractionated into three parts: N1, N2, and N3, defined as non-extracted N (N1), protein-N non-VSP-N (N2 = protein-N–VSP-N), and the VSP-N (N3).
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Electrophoretic analysis of proteins
Figure 1A illustrates the annual phloem protein pattern, showing the January and June protein patterns for parenchyma and wood. Good repeatability was observed at all time points in each tissue. Nearly 40 protein bands were clearly separated in each lane, and their apparent MW ranged from 10–140 kDa. The two VSP, 19 kDa and 16.5 kDa, exhibited major annual variations. Their levels began to rise from the autumn until January, and then sharply decreased before full blossoming in April. Between April and August their levels remained low. A few polypeptides such as PP15 showed inversed annual variations (maximum in summer, minimum in winter). Most of the other bands appeared to be almost invariable (see PP41 as an example). Very similar protein patterns were obtained for parenchyma and phloem. The same annual variation was observed for VSP 19 kDa in wood. However, VSP 16.5 kDa was only detected in November in this tissue.
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Figure 1B shows the kinetics of the relative intensities of the bands studied in phloem. The results obtained with 12 repeated gels are shown as a percentage of the total proteins extracted. As may be seen from the standard deviation bars drawn for VSP bands, considerable variability was seen as a function of the date and, depending on the sample at the same date, especially during the winter. The VSP sum ranged from 7–22% of the total protein content in January, depending on the shoot. On average, VSP 19 kDa and VSP 16.5 kDa accounted for up to 13% of total phloem proteins in winter and 4% in summer. Similar winter/summer VSP ratios were obtained in the parenchyma and wood. The other bands did not vary so widely: whereas polypeptide PP41 remained constant (about 2% of proteins), polypeptide PP15 rose lightly from 1.5% in winter to 2% in summer.
Kinetics of nitrogen, proteins and VSP in shoots
Total nitrogen, total extracted proteins and VSP were quantified for each tissue of the shoot (Fig. 2). Throughout the year, bark tissues (parenchyma and phloem) exhibited higher levels of N compounds than wood. The protein content was particularly low following wood extraction. Whereas the N content was always higher in phloem than in parenchyma, protein levels were only higher in phloem during the winter. The kinetics of the total nitrogen and total proteins extracted were similar to those of the VSP contents: following an increase in the autumn which lasted until January, the nitrogen and protein contents abruptly decreased in March and April. Levels remained low from April to August and then started to rise again.
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Figure 3 illustrates the annual variations of nitrogen, protein and VSP in shoots, and shows the relative contributions of each tissue to the total content. The relative weights of the three fractions were nearly constant all year long. Although the parenchyma represented only 23% (annual mean) ±5 (SD) shoot DW, it made the largest contribution to the shoot protein content (generally more than 60%). Phloem (5% ±2 shoot DW), because of its high protein content, and wood because of its weight (72% ±6 shoot DW), each made up 10–20% of the protein content. In shoots, the most pronounced mean decrease, between March and April, concerned VSP (–85%) and proteins (–67%). N was less mobilized, with –29%. Moreover for VSP, the decrease appeared earlier than for the protein and N contents. Increases in the autumn (between August and November) reached 57%, 166%, and 300%, respectively, for the N, protein and VSP contents. The maximum amplitude of the VSP content was much greater (from 6.5 mg g–1 DW in January to 0.35 mg g–1 DW in June) than for the protein content (from 5.9 mg g–1 DW in March to 1.4 mg g–1 DW in June) and the N content (5.4 mg g–1 DW in March to 3.1 mg g–1 DW in June). Figure 3 shows a winter/summer (W/S) ratio of 18.5, 4.3, and 1.7 for VSP, proteins and N, respectively.
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Comparison of variations in different N-pools in the bark
Because of the low nitrogen content in wood, the problem encountered in obtaining a protein extract and the absence of VSP 16.5 kDa (only present in November), calculations of the nitrogen-pools arising from VSP (N3), total soluble protein minus VSP (N2) and all other the nitrogen compounds (N1) were restricted to the bark. Figure 4 shows that N1 comprised the principal N fraction, of more than 80% (autumn–winter) and up to 90% (spring–summer). Nitrate contributed to this N1 pool but levels remained stable, at around 0.25 mg g–1 DW content in both January and July. The fraction N2 varied mainly from September (16% N) to March (20% N), after which it remained constant during spring and summer. It then represented about 10% of the total N content. Nitrogen from the VSP fraction (N3) accounted for 2–3% of the N content during the storage period (autumn and winter) and less than 1% from April to August.
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According to Fig. 4, three different periods could be distinguished: from August to March, March to April and April to August. Table 1 shows the percentage variations of the different N pools during these periods. From August to March, each N pool increased, with a higher percentage for N2 (171%) and particularly for N3 (376%). However, N2 and N3 accounted, respectively, for only 3% and 21% of the total N accumulation in the bark, the remainder being due to the non-extracted fraction (N1). Levels of all N fractions fell in March and April. The major decline in N3 (–84%) represented, in fact, only 12% of mobilized N-protein in the spring (–69%), i.e. 5% of mobilized total N, comprised principally (55%) of N1 (–34%). From April to August, only N3 began to show a slight increase whereas total N fell by 23%. Moreover, in terms of the W/S ratio, it was markedly higher for N3 (15) than for other N fractions (<5).
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The annual kinetics of total N, N-protein, N1, and N2 were well correlated (Fig. 5). A strong correlation existed between total N and N-protein, even though N-protein constituted only a small proportion of total N. Based on r2, 72% of the variability of N-protein (y) could be explained by N (x). The best correlations were observed between N and N1 (r2=0.95) and between N-protein and N2 (r2=0.95). The weakest correlations were obtained between N3 and other N fractions (r2<0.31), indicating a specific pattern of variation for N-VSP.
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Characterization, quantification and potential function of VSP
The electrophoretic results showed three stem protein patterns: a large number of polypeptides which did not change during the year (PP41), some which were more present in summer (PP15) and others which increased markedly in the autumn and dropped sharply near flowering (VSP19, VSP16.5). The latter exhibited annual kinetics characteristic of VSP (O’Kennedy and Titus, 1979) and were probably those VSP which had been detected by Arora et al. (1992). The large number of replicates (12) showed that VSP accumulation varied considerably between shoots. Part of this variability may have been due to the spatial position of the shoot on the tree. The winter VSP content in peach trees (13% of protein in bark, 11% in wood) was in line with the 16% found by Roberts et al. (1991) in softwood bark and the 25% seen by Langheinrich and Tischner (1991) in poplar bark, but was lower than that found by Sauter et al. (1988) in poplar wood (70%).
The most abundant VSP (19 kDa) exhibited the same kinetics in the three tissues. In winter, it accounted for up to 6% of the total proteins extracted from phloem and parenchyma and nearly 11% in wood (data not shown). This polypeptide was detected throughout the year; its smallest contribution to the protein content was about 1.5% in July. VSP 16.5 kDa and VSP 19 kDa are also present in apricot bark (M Faurobert and L Gomez, unpublished results). As shown by-two dimensional electrophoresis, several polypeptides with different isoelectric points, but similar MW, were implicated in the variations seen in each VSP band. Some could remain constant throughout the year, but further investigations are required using this technique.
In bark tissue, the two VSP identified had similar annual kinetics. Contrary to VSP 19 kDa, VSP 16.5 kDa appeared in wood only in November. This protein had disappeared in January but could still be detected in December, in line with the observations of Arora et al. (1992). In view of the lack of precise knowledge on this protein, it is difficult to confer a physiological meaning on this late and transient accumulation. Nevertheless, accumulation of this protein in wood is not related to the VSP function in spring growth, but its kinetics may be linked to N translocation within the tree during winter. Further studies are required to support this hypothesis. Arora et al. (1992) had also described a 60 kDa dehydrin as a VSP in bark, which may play a role in cold acclimation. This protein was also detected, but its seasonal variations were too imprecise for it to be considered a putative VSP. Another VSP (61 kDa) found only in buds (Lang, 1994), was not detected during this study.
In soybean and trees, VSP gene expression is influenced by diverse developmental or external stimuli: source–sink status, N availability wounding or drought, light, jasmonic acid, and sucrose (Staswick, 1994; Rowland and Arora, 1997). Since the two VSP, 16.5 kDa and 19 kDa, are present all year in the bark, their synthesis is not strictly connected with a seasonal storage phenomenon. In peach trees, Rowland and Arora (1997) suggested a relationship between the 16.5 kDa protein and endodormancy, and between the 60 kDa and 19 kDa proteins and resistance to frost. No data are as yet available on a causal relationship between these VSP and cold hardiness. Certain VSP are glycosylated (Langheinrich, 1993; Stepien and Martin, 1992). The sugar moiety is expected to provide better thermostability during winter (Stepien et al., 1994) which may contribute to cold hardiness. In alfalfa, a ß-amylase (57 kDa) acts as a VSP (Gana et al., 1998). In soybeans, 
and ß VSP genes exhibit sequence homology with the tomato acid phosphatase (Staswick, 1994), but these VSP do not provide effective phosphatase activity in situ. In woody plants, the increased activities of certain enzymes (dehydrogenase, lipase, catalase) have been related to seasonal changes or chilling accumulation (Rowland and Arora, 1997). However, contrary to herbaceous plants, no enzymatic role has been proposed for VSP with a known MW. Moreover, no mechanism concerning their higher levels of synthesis in the autumn is known.
Annual kinetics of N compounds
These results confirm the annual variations of N (Stassen et al., 1981) and protein (Arora et al., 1992) contents in peach trees, characterized by accumulation in the autumn and mobilization in the spring. N compounds exhibit very varied seasonal amplitudes, reaching annual maximum and minimum levels at different times. Changes to the VSP content can only explain a small part of N variations. Thus, in shoots, VSP mobilization which mainly occurs between March and April, represents only 13% of the mobilized proteins. Even though VSP have the best storage yield (shoot WS ratio =18.5), a larger quantity of N is stored in another way.
As has long been known for apple trees (Tromp and Ovaa, 1971; O’Kennedy et al., 1975), peach bark also stores more proteins than wood over the year: annual means of 17% and 83% of stem protein are extracted from wood and bark, respectively. Annual changes are mainly due to the parenchyma, which constitutes the principal stem storage tissue, even if proteins are often more highly concentrated in phloem and wood is the major shoot constituent (annual mean of 72% shoot DW).
The biochemical difference between bark and wood may be related to the presence of more protein-filled vacuoles (protein bodies) in bark (Stepien et al., 1994). The structural fraction of the total protein content is higher in wood, where living cells are limited to the functional xylem, the external part of this tissue. As reported by Clausen and Apel (1991) in poplar trees, it was considered that the wood proteins extracted arose from this limited zone, where cell metabolism changes with the phenological stage, whereas total N came from the entire wood. Protein storage near the xylem and phloem appears to facilitate their mobilization (Staswick, 1994).
Different N storage forms in the bark
Working on bark has taken account of the living fraction of the shoot. The existence of three phases for N metabolism has allowed the annual kinetics of N fractions (N1, N2, and N3) to be connected to phenological stages.
(1) Protein storage phase (from August to March): Although the bark N content rises until flowering, the most intense storage period runs from September to when leaves fall in November. During this period, storage in the perennial tissue arises as a result of N-protein translocation from leaves (Titus and Kang, 1982) or as a reduction in the NO3 absorbed by the roots (Gojon et al., 1994). N3 levels rise markedly, but this does not account wholly for the N-protein increase. These findings confirm that bark stores other proteins. Such an accumulation may result from strong synthesis, stimulated by a higher availability of basal constituents (nitrate, free amino acids and carbohydrates), and/or a lack of use of synthesized proteins in the absence of sinks. However, this protein increase accounts for only 24% of the total N accumulation, the remainder being due to the rise in N1. No major, concomitant decrease in non-structural carbohydrates (MO Jordan et al., unpublished results) could lead to an overestimation of N-protein levels. Nitrogen from nitrate, free amino acids and structural proteins are the constituents of N1. At the end of the summer, growth has been achieved and it is probable that structural protein levels remain stable until the spring regrowth. The nitrate content always remains low, in agreement with the findings of Gojon et al. (1994). In fact, according to Bussi et al. (1989), nitrate reduction and amino acid synthesis occur in peach roots. One explanation for the N1 increase may be linked to the free amino acid storage previously described for peach trees by Stassen et al. (1981). Until leaf fall, amino acids are essentially relocated from senescing leaves. The rise in N1, observed after leaf fall, may be due to relocation events within the perennial part of the tree.
(2) Mobilization phase (from March to April): This phase is characterized by a rapid fall in N components between March and April, which signals their translocation to the sinks (buds). Bark cells process N forms differently, with, in particular, an early mobilization of VSP. However, N3 levels start to decline in January, while N1 and N2 continue to rise very slightly. Recently, in soybean, Staswick et al. (2001) showed that VSP were reduced by antisense and played little if any direct role in overall plant productivity under typical growth conditions. In this perennial plant model, VSP are not the main N storage form but the first N reserve mobilized. Their role could be linked to growth initiation and may be more qualitative than quantitative. VSP may be preferentially used to initiate spring regrowth. The other proteins (N2) are rapidly mobilized before the tree becomes autotrophic. N1 is quantitatively the most higher mobilized fraction (–2.4 mg N g–1 DW). This probably means that free amino acids are used for protein synthesis, before or after translocation towards growing areas. From around bud-break, Stassen et al. (1981) observed a drop in the arginine and asparagine contents in the shoots and roots of peach trees.
(3) Assimilation phase (from April to August): During this period, only the N1 fraction seems to be used for vegetative and/or fruit production. The very low and constant N2 fraction may constitute house-keeping proteins, i.e. those associated with the basal cell metabolism. The rise in N3, even if it is moderate, shows that VSP are the first N fraction to be stored. During this period, the bark apparently works as a transit area for nitrogen components and, in particular, for the free amino acids synthesized in the roots and carried in sap. The decline in N1 is probably due to a reduction in amino acid synthesis or the higher requirements of sinks. Moreover, intensive bark protein synthesis is possible even if the protein content remains low: many proteins may be synthesized and relocated to the sinks. The lower total N content in bark, measured in August (6.4 mg N g–1 DW), may be equal to the unavailable N fraction. It is proposed that the more marked difference in N content observed between March and August (6.35 mg N g–1 DW), constitutes the N mobilization potential of the bark. This potential may be affected by the N fertilization level.
Very strong correlations may enable an approximate evaluation of the N1 content and the N-protein content from the total N content. Experiments using different levels of fertilization are necessary to confirm the regression parameters and, ultimately, to see whether a simple total N determination may be sufficient to evaluate approximately the N reserve throughout the year. A weak or non-existent correlation between N3 and the other N fractions confirms the particular kinetics of the VSP content. In total shoots, VSP are characterized by a WS ratio which is at least 3-fold that of the other N fractions. Their low quantitative importance explains why their annual kinetics do not affect the strong correlation between N-protein and N total contents.
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The bark or more precisely the parenchyma, constitutes the principal site of storage in shoots. The three N fractions evidenced (N1, N2 and N3) play a role in N storage. VSP are characterized by early, and marked, mobilization in the spring, and by the greatest annual variations (ratio W/S=15), but they do not quantitatively represent the principal stored N pool, which may be made up of free amino acids. However, VSP appear to play an important role in initiating of spring regrowth. Rapidly exhausted, they are replaced by other soluble proteins. Free amino acids then permit a rapid return of protein synthesis.
Future studies are planned on peach trees to see how N fertilization affects N management, to determine the role of N fractions in spring regrowth and to clarify the correlations between different N forms in different organs of the tree.
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Acknowledgements |
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The authors would like to thank E Rubio, D Bancel and N Tassard for their technical assistance, MO Jordan for critical reading of the manuscript, and V Hawken for revising the English.
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References |
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