Journal of Experimental Botany, Vol. 53, No. 371, pp. 1131-1141,
May 2002
© 2002 Oxford University Press
Original Papers |
Nitrogen storage and remobilization in Brassica napus L. during the growth cycle: effects of methyl jasmonate on nitrate uptake, senescence, growth, and VSP accumulation
1UMR INRA/UCBN 950, Physiologie et Biochimie Végétales, Institut de Recherche en Biologie Appliquée, Esplanade de la Paix, Université de Caen, 14032 Caen Cedex, France
2Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, Ceredigion SY23 3EB, UK
Received 10 September 2001; Accepted 27 December 2001
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Abstract |
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The role of methyl jasmonate (MeJa) in promoting senescence has been described previously in many species, but it has been questioned in monocarpic species whether induced senescence is a result of a potential death hormone like MeJa, or a consequence of an increased metabolic drain resulting from the growth of reproductive tissue. In oilseed rape (Brassica napus L.), a polypeptide of 23 kDa has been recently identified as a putative vegetative storage protein (VSP). This polypeptide could be used as a storage buffer between N losses from senescing leaves putatively promoted by methyl jasmonate that might be produced by flowers, and grain filling which occurs later on, while N uptake is strongly reduced. In order to describe causal relationships during Brassica napus L. plant responses to MeJa treatment, a kinetic experiment was performed to determine the order and the amplitude with which general processes such as growth, photosynthesis, chlorophyll content, N uptake, and N storage under the form of the 23 kDa VSP are affected. One of the most immediate consequences of MeJa treatment was the strong reduction of nitrate uptake within 6 h, relative to control plants. However, this was not a specific effect as K+ uptake was similarly affected. Photosynthesis was reduced later (after 24 h), while chlorophyll content as well as leaf growth also decreased in a similar way. Moreover, this was concomitant with a remobilization of endogenous unlabelled N from senescing leaves to roots. Accumulation of the 23 kDa VSP was induced in the taproot after 24 h of MeJa treatment and was increased 10-fold within 8 d. On the other hand, the reversible effect of a MeJa pretreatment was tested in the long term (i.e. along the growth cycle) using plants previously grown in field conditions induced for flowering. Results show that a MeJa pulse induced a reversible effect on N uptake inhibition. In parallel, protein immunologically related to the 23 kDa VSP was detected in stems with a similar molecular weight (23 kDa), and in flowers and leaves with a molecular weight of 24 kDa. This accumulation was concomitant with the remobilization of both subunits of Rubisco. During stem and pod development, this protein induced by MeJa is fully hydrolysed. The external and intermittent supply of MeJa mimic some of the plant physiological processes previously reported under natural conditions. This suggests that in oilseed rape, methyl jasmonate could be considered as a possible monocarpic senescence factor while accumulation/mobilization of the 23 kDa VSP in taproot could be a marker for the cessation of N uptake and the initiation of a massive leaf senescence.
Key words: Brassica napus, methyl jasmonate, monocarpic senescence, nitrate uptake, nitrogen remobilization, tissular localization, vegetative storage protein.
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Introduction |
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The term ‘death hormone’ has been used (Noodén and Leopold, 1978) when discussing senescence in plants that flower only once (monocarpic plants) such as peas, cereals or soybean. Monocarpic senescence is a dramatic phenomenon, visible in most fields before harvest. Senescence is a highly regulated, ordered series of events involving loss of photosynthesis, breakdown of the CO2-fixing enzyme such as ribulose bisphosphate carboxylase/oxygenase (Rubisco) and other proteins, loss of chlorophyll, removal of amino acids and, in some cases, accumulation of new proteins. Senescence promotes movement of nutrients from the vegetative parts to the fruits or to the seeds. As it has been shown that removal of flowers or fruits prevents senescence, one hypothesis that may explain this phenomenon is that developing fruits produce a senescence factor (Engvild, 1989; Koda, 1994; Smart, 1994; Buchanan-Wollaston, 1997). This so-called death hormone would be exported from the fruits to the vegetative parts of the plant, where it may stop growth, activate senescence, promote remobilization of nutrients, and finally lead to the death of the plant. An alternative hypothesis has been developed to explain monocarpic senescence: the nutrient drain hypothesis of Molisch modified by Kelly and Davies proposes that a pull from the strong sink of young reproductive tissues monopolizes all nutrients, helped by a reduced sink strength of the vegetative parts in the flowering plant (Kelly and Davies, 1988). Engvild reviewed the death hormone hypothesis and described the possibility that jasmonic acid and methyl jasmonate may be the death hormones (Engvild, 1989) since soybean fruit contains a high level of jasmonic acid (Lopez et al., 1987). Moreover, a loss of chlorophyll, the repression of genes related to photosynthesis at the transcriptional and translational levels (Reinbothe et al., 1994) and the degradation of Rubisco (Weidhase et al., 1987a, b; Müller-Uri et al., 1988; Rakwal and Komatsu, 2000; Wierstra and Kloppstech, 2000) are typical symptoms promoted by MeJa treatment in mature leaves. These symptoms resemble those of senescence. Methyl jasmonate can also activate the expression of several genes, leading to the accumulation of their products, which are referred to as jasmonate-induced proteins (JIPs) and vegetative storage proteins (VSPs). Thus, the jasmonate-induced proteins seem to function as stress proteins and may protect and defend plants under stress conditions. For example, the proteinase inhibitors of tomato and potato accumulate in leaves after wounding, either by mechanical damage or injury by insects (Farmer and Ryan, 1990) and are most likely involved in the protection of the remaining tissues against future insect attacks (Sembdner and Parthier, 1993).
Among jasmonate-induced storage proteins, the group of vegetative storage proteins of soybean has been studied intensively in several laboratories. These glycoproteins accumulate in vegetative organs (leaves) during flowering, leaf senescence and dessication periods (Sembdner and Parthier, 1993). In oilseed rape (Brassica napus L.), previous study (Rossato et al., 2001a) has prompted questions about the identity and the transduction pathways of a senescence signal that moves into leaves and thus may be a factor initiating a decrease in N uptake and nutrient depletion of leaves, stem and taproot to sustain grain filling. On the other hand, other previous results (Rossato et al., 2001b) have shown that, in rape, a putative vegetative storage protein (VSP) of 23 kDa could be used as a storage buffer between N losses from senescing leaves putatively promoted by methyl jasmonate (which could be produced by flowers), and grain filling which appears later in the growth cycle. Moreover, it has also been shown that this 23 kDa putative VSP was accumulated at the same rate when MeJa was applied in the nutrient solution or sprayed on the leaves. Because it is well known that methyl jasmonate has strong and diverse effects on plant physiology, a kinetic experiment using MeJa-treated plants was performed in order to determine the order of events, the amplitude with which this hormone affects different physiological processes (from growth, photosynthesis, and chlorophyll content to N uptake and N storage under the form of the 23 kDa VSP) and finally to describe putative causal relationships. A second set of experiments was undertaken using plants taken from the field then pretreated with MeJa in order to see which physiological processes can be restored after MeJa removal, and how it can be related to what can be found in natural field conditions.
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Materials and methods |
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Plant culture
Experiment 1:
Seeds of Brassica napus L. cv Capitol were imbibed for 48 h on tissue paper saturated with 10 mol m-3 CaSO4 and then sown into six culture units of a flowing solution culture (FSC) system incorporating automatic control of concentrations of
, K+ and H+ in solution (Clement et al., 1974; Hatch et al., 1986). Each culture unit contained 200 dm3 of recirculating nutrient solution and 24 culture vessels, each containing three plants. The FSC system was located in a greenhouse, the solution temperature was maintained at 20±0.5 °C and air temperature at 20±2/15±1 °C day/night (09.00–21.00 h) throughout the experiment. The plants were established under natural illumination until day 17 after sowing. Supplementary light (09.00–21.00 h) of 200 µmol m-2 s-1 PAR was provided between days 18–24 by a single 400 W SON-T lamp (Philips Lighting Ltd, Croydon, Surrey, UK) suspended 1.5 m above the surface of each culture unit. On day 24 after sowing, natural light was excluded and thereafter illumination was provided by paired 400 W SON-T and HPI/T lamps (Philips Lighting Ltd, Croydon, Surrey, UK) giving 500 µmol m-2 s-1 PAR at canopy height, over 12 h.
Nutrient concentrations in each culture unit were initially (mmol m-3): , 250; K+, 250; 
, 50; Mg2+, 100; 
, 325; Fe2+, 5.4; with micronutrients as previously described (Clement et al., 1978). All culture units were drained and refilled with fresh nutrient solution of the same composition on day 18 after sowing. Nutrient concentrations were allowed to deplete by plant uptake until day 24 when automatic monitoring (every 27 min) and resupply of nutrients was introduced. Thereafter, the concentrations of K+ and were maintained at 20±2 mmol m-3 in each culture unit by automatic resupply at a rate equal to the depletion of K+ or by plant uptake. All other nutrients were supplied automatically in fixed ratios to the net uptake of K+ (for 1 mol of K, 0.645, 0.057, 0.045, 0.00075, and 0.522 mol of S, Mg, P, Fe, and Ca, respectively, were supplied, with micronutrients as described previously (Clement et al., 1978). Solution pH was maintained at 6.0±0.1 by automatic delivery of H2SO4 or Ca(OH)2 to each culture unit throughout the experiment. A concentration of 20±2 mmol m-3 was maintained automatically in each culture unit from day 24 until the start of the methyl jasmonate (MeJa) treatment (day 26). Net uptake of K+ and was calculated on an hourly basis from the amounts required to maintain the ‘set point’ concentrations in the flowing solutions.
Experiment 2:
Brassica napus L. cv. Capitol plants were taken from a field plot in December when they were at the six leaf stage. The roots were gently rinsed with distilled water before transferring the plants to a hydroponic system (27 seedlings per 15 dm3 plastic tank) in a growth room. The aerated nutrient solution previously described (Rossato et al., 2001a) was renewed every 2 d. CaCO3 was then given in excess at a final concentration of 2 mM in order to maintain the solution pH at 6.5±0.2. Light was provided by high-pressure sodium lamps (250 µmol m-2 s-1 of photosynthetically active radiation at the height of the canopy) for 16 h d-1. The thermoperiod was 20 °C (day) and 15 °C (night). The plants were used for experiments when the lateral roots (partly damaged during collection of the plants from field plots) had been growing for 3 weeks.
Kinetic study of the effects of methyl jasmonate supply (Experiment 1)
On day 26 after germination, methyl jasmonate (100 µM) was applied in the nutrient solution for 8 d. Throughout this period, the composition of nutrient solution was identical to the above-described solution (Experiment 1), except for the N source (K15NO3) which was labelled with a 15N excess of 1.54% at the start of MeJa application. At the beginning of the treatment (‘time zero’) and 6, 10, 24, 48, 96, 144, and 192 h later, net uptake of nitrate and potassium, photosynthetic activity (using a photosynthesis system Li-Cor, Eurosep Instruments), and chlorophyll content (using a SPAD-502 system, Minolta; Manetas et al., 1998) were measured for both control (grown without methyl jasmonate) and treated plants. Plants were then harvested at each date previously mentioned.
Study of the effects of a methyl jasmonate pretreatment along the growth cycle (Experiment 2)
At day 0, plants at the bolting stage C1 (CETIOM source) were pretreated for 3 d with 100 µM methyl jasmonate (that was previously shown to induce 23 kDa protein accumulation within plants) applied in the nutrient solution, then grown in a nutrient solution without MeJa during 73 d (i.e. until the end of the growth cycle stage G5). Throughout this period, the composition of nutrient solution was identical to the above-described solution (Experiment 2) except for the N source (K15NO3) which was labelled with a 15N excess of 1.00%. Control plants (constantly grown without methyl jasmonate) and treated plants were sampled after 0, 3, 10, 17, 24, 39, and 76 d of treatment.
Sampling, chemical analysis and calculation of N remobilization
Roots of plants supplied with K15NO3 were first rinsed with a 1 mM solution of CaSO4 to remove any superficial 15N. At each harvest date, three vessels of three plants each (Experiment 1) or six plants (Experiment 2) were separated into leaves, stems, taproots, lateral roots, flowers, pods, and grains. Each plant fraction was weighed, freeze-dried, re-weighed for dry weight determination, and then ground to a fine powder for isotopic analysis. For SDS-PAGE and Western blot analysis of the soluble proteins, tissues of harvested plants were immediately frozen in liquid N2 and kept at -80 °C until soluble protein extraction.
The total N and 15N in the plant samples was determined with a continuous flow isotope mass spectrometer (Twenty-twenty, PDZ Europa Scientific Ltd, Crewe, UK) linked to a C/N analyser (Roboprep CN, PDZ Europa Scientific Ltd, Crewe, UK). As all the mineral N taken up from the nutrient solutions was 15N-labelled, the cumulated uptake and further translocation into plant parts could be calculated from the excess 15N in each tissue. Natural 15N abundance (0.3663±0.0004%) of atmospheric N2 was used as reference for 15N analysis. Therefore, excess 15N [E (%)] in a given tissue was obtained by E (%)=A (%)-0.3663, where A (%) is tissue 15N abundance given by mass spectrometry. Nitrogen coming from uptake in a given tissue (15N) was calculated by 15N=Ntotx[E (%)/Es (%)], where Ntot is total N in tissue (mg plant-1) and Es (%) is nutrient solution 15N excess (1.54% or 1.00%, for experiments 1 and 2, respectively). Consequently, the patterns of mobilization of unlabelled N (14N absorbed prior to the beginning of the experiment) between plant parts could be used to calculate N remobilization within the plant. The N in growing leaves derived from the mobilization of endogenous unlabelled N was calculated by subtracting from total N (14N+15N), firstly, the 15N content derived from uptake of 15 and, secondly, the initial 14N content found in this tissue at the beginning of the experiment.
Soluble protein extraction and analysis
Soluble proteins were extracted and analysed by SDS-PAGE as previously described (Rossato et al., 2001a). Electrophoretic transfer of proteins from SDS-PAGE gels onto PVDF membrane (Immobilon-P, Proteigene, Saint-Marcel, France) was conducted by semi-dry electroblotting (100 V, 2.5 mA for 30 min, Milli Blot system, Proteigene), according to the protocol described previously (Towbin et al., 1979). After blotting, PVDF membranes were treated with affinity-purified polyclonal anti-23 kDa protein (dilution 1/1000) primary antibodies previously obtained (Rossato et al., 2001b). The antigen–antibody complex was visualized with alkaline phosphatase linked to goat (Ovis L.) anti-rabbit (Oryctolagus cuniculus L.) IgG as described earlier (Blake et al., 1984).
Statistical analysis
All data were processed by analysis of variance and mean separation was performed using Fisher-Snedecor test (Statitcf software).
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Results |
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Kinetic study of the effects of methyl jasmonate supply
Plant growth:
Growth of plants at the vegetative stage treated with methyl jasmonate (MeJa) was not affected during the first 48 h in comparison with control plants (Table 1
). Thereafter, the growth of treated plants was strongly reduced by MeJa as compared with control plants. Thus, after 8 d of treatment, growth of leaves and roots was finally reduced by 69% and 63%, respectively (Table 1).
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Photosynthetic activity and chlorophyll content:
After 10 h, photosynthetic activity strongly decreased from 26.5 to 5.7 µmol CO2 m-2 s-1 between 10 h and 8 d of treatment, respectively (Fig. 1A). By contrast, photosynthesis in control plants was constant during the same period at approximately 32 µmol CO2 m-2 s-1 (Fig. 1A).
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The chlorophyll content in leaves was affected later, after 24 h, decreasing strongly in treated plants, while increasing gradually in control plants (Fig. 1B
). After 8 d of treatment, chlorophyll content in leaves was finally reduced by 84% as compared with control plants (Fig. 1B).
Nitrate and potassium uptake:
Nitrate uptake was strongly repressed within only 6 h of methyl jasmonate treatment (Fig. 2A). Northern blots show that both BnNRT1 and BnNRT2 gene expression, encoding low (LATS) and high (HATS) affinity NO3- transporters, respectively, was repressed (data not shown). After 8 d of treatment, nitrate uptake was finally reduced by 90% as compared with control plants (Fig. 2A). Similar inhibition by MeJa was also found for potassium uptake (Fig. 2B).
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N partitioning in plants:
Partitioning of the 15N taken up into plant tissues is presented in Fig. 3A, B, C. As a consequence of the inhibition of nitrate uptake by MeJa, 15N content of leaves, lateral roots and taproot was reduced by 96, 90 and 95%, respectively, after 8 d of treatment as compared with control plants (Fig. 3A, B, C). Moreover, plants treated with methyl jasmonate showed a significant (P<0.05) preferential partitioning of 15N taken up into lateral roots (32% versus 14.3%) at the expense of aerial tissues (Fig. 3A, B, C). Thus, only 64.8% of 15N taken up was translocated to leaves of treated plants versus 83.1% for control plants (Fig. 3A, B, C). This preferential allocation of absorbed 15N to roots was concomitant with a remobilization of endogenous unlabelled N from leaves to lateral roots and taproot (Fig. 3D, E, F). As a consequence, partitioning of endogenous unlabelled N was modified significantly (P<0.05) in treated plants, decreasing from 80.5% to 76.7% in leaves, while increasing in lateral roots from 18.3% to 20.5% (Fig. 3D, E, F). However, in treated plants, preferential partitioning of 15N taken up and endogenous unlabelled N to taproots was not significantly different from control plants. This small taproot contribution is probably linked to the very low taproot biomass in this experiment linked to the developmental stage and the growth conditions of the plants.
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Accumulation of the 23 kDa protein:
Accumulation of the 23 kDa protein was induced in taproots after only 24 h of methyl jasmonate supply (Fig. 4A). After 8 d of treatment, this protein became the most prominent polypeptide of the taproot soluble proteins and represented 16% of the total soluble proteins in this tissue (Fig. 4A). In addition, accumulation of the 23 kDa protein paralleled those of soluble proteins in taproots (data not shown). Using polyclonal antibodies raised against the 23 kDa protein (Rossato et al., 2001b), an immunologically related protein of 24 kDa was detected in leaves (Fig. 4B). This 24 kDa protein accumulated later in leaves than in taproots (i.e. within 48 h of methyl jasmonate treatment) and reached 13% of the total leaf-soluble proteins after 8 d (Fig. 4B). By contrast, no immunoreactive protein was found in lateral roots of treated plants (data not shown).
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Study of the effects of a methyl jasmonate pretreatment along the growth cycle
Methyl jasmonate pretreatment resulted in further inhibition of cumulative nitrate uptake (calculated from 15N labelling) during the following 7 d (Fig. 5). Thereafter, cumulative 15 uptake was significant although lower than that of control plants. Thus, the inhibition of nitrate uptake by methyl jasmonate seems to be reversible, at least partly, when plants are transferred to a nutrient solution without MeJa. Nevertheless, like control plants, cumulative nitrate uptake of pretreated plants exhibited an inflexion point during flowering (Fig. 5). At the end of the growth cycle stage G5, total 15 taken up was finally 1.6-fold lower for pretreated plants as compared with control plants. In addition, total N content of pretreated stems was strongly reduced when compared to control plants (-61.7% at day 36 after the end of methyl jasmonate supply, Fig. 6). Moreover, N content of pretreated taproots was higher than those of control plants during the first 17 d following MeJa supply, then became lower until the end of the growth cycle (Fig. 6). Methyl jasmonate pretreatment did not affect the N content of leaves and lateral roots when compared to control plants (Fig. 6). Western blots performed with polyclonal antibodies raised against the 23 kDa protein show that, in pretreated plants, an immunologically related protein was detected in stems with a similar molecular weight (23 kDa), and in flowers and leaves with a molecular weight of 24 kDa (Fig. 7). By contrast, no immunoreactive protein was found in lateral roots, pods and grains (data not shown). In taproot and leaves of pretreated plants, the accumulation of this protein was at the highest level 7 d after the end of MeJa supply, and represented 16% and 13% of the total soluble proteins of these tissues, respectively (Fig. 7). This accumulation was concomitant with the remobilization of the two subunits of Rubisco (Fig. 7). In flowers and stem, this protein reached 8% and 4% of the total soluble proteins of these tissues, respectively (Fig. 7). During stem and pod development, this protein was gradually remobilized in taproots, stems and leaves of pretreated plants, to be finally fully hydrolysed at the end of the growth cycle stage G5 (Fig. 7).
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Discussion |
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According to previous studies, it is well known that methyl jasmonate triggers numerous physiological processes (for reviews see Herman et al., 1989; Farmer and Ryan, 1990; Falkenstein et al., 1991; Pelacho and Mingo-Castel, 1991; Staswick, 1990, 1992, 1994; Staswick et al., 1991; Sembdner and Parthier, 1993; Koda, 1994; Creelman and Mullet, 1997). However, the present study clearly shows that given mechanisms are affected at different time scales (Fig. 7
). One of the most immediate consequences was the strong reduction of nitrate and potassium root uptake within 6 h. Other results (data not shown) suggest that uptake, in particular, was down-regulated at the gene level, as two putative transcripts encoding low (LATS, BnNRT1) and high (HATS, BnNRT2) affinity transporters were expressed to a low level in MeJa-treated plants relative to control plants. However, this was probably not a specific effect as 
(data not shown) and also K+ uptake were similarly affected. This large decrease of nutrient uptake by the root triggered by exogenously supplied MeJa suggests that the down-regulation of uptake found at the flowering stage (Rossato et al., 2001a) under normal conditions, could be the result of an increased MeJa synthesis in planta. Photosynthesis was reduced later (after 24 h) and less strongly than uptake (Fig. 8). Chlorophyll content as well as leaf growth decreased in a similar way. These results are consistent with those reported in barley where leaf segments treated with methyl jasmonate exhibited a strong yellowing caused by an intense loss of chlorophyll a and b (Wierstra and Kloppstech, 2000). This yellowing and the decrease of total chlorophyll content were first visible after 24 h of incubation and progressed substantially during incubation. In rice, jasmonate action was also mediated through the inhibition of photosynthesis (Rakwal and Komatsu, 2001). The transpiration rates of plants submitted to MeJa treatment (data not shown) have also been quantified and a very slight reduction was found as previously reported for Hordeum vulgare (Horton, 1991).
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The role of MeJa in promoting senescence has been described previously in many species (Noodén, 1988; Anderson, 1989; Horton, 1991; Sembdner and Parthier, 1993; Koda, 1994; Creelman and Mullet, 1995, 1997; He et al., 2001), but it has been questioned in monocarpic species whether induced senescence is a result of potential death hormone like MeJa, or a consequence of an increased metabolic drain resulting from the growth of reproductive tissue (Engvild, 1989). Because the plants used in this study (Experiment 1) were not induced for flowering and therefore remained at the vegetative stage, no corresponding metabolic drain occurred. However, as shown previously (Rossato et al., 2001b), MeJa increases N cycling within the plant at the expense of leaves, leading to a transient accumulation of N within roots. This fits with results obtained with untreated plants for which N, made available from leaf senescence, is mobilized to taproots and stems acting as a transient N storage buffer if the sink strength of reproductive tissue such as developing seeds is too low (Rossato et al., 2001a). The accumulation of the 23 kDa putative VSP in taproot occurred very early after MeJa supply, i.e. before 24 h (Fig. 8
) and was increased by a 10-fold factor within 8 d. At the same time, the concentration of soluble proteins in taproots was also increased, but to a much lower level (by 139% after 8 days). It follows, from the different ranges of accumulation found for VSP and for soluble proteins, that numerous soluble proteins, other than VSP, must be hydrolysed in the same time.
In order to mimic a transient production of MeJa that may occur under natural conditions, plants induced for flowering were experimentally submitted to a 3 d pulse of methyl jasmonate (Experiment 2). Results showed that VSP accumulation was strongly increased in all tissues analysed and this protein was then fully hydrolysed during the next following week (Fig. 7). Furthermore, this pattern of accumulation/mobilization paralleled the change of total N content in taproot, which reinforces the role of this tissue to act until the end of flowering, as an N storage organ. In addition, the results showed clearly that a MeJa pulse induced a reversible effect on N uptake inhibition. Taken together, these results show a strong relationship between VSP accumulation/remobilization within taproots and the loss/recovery of N uptake activity, and suggest that accumulation/remobilization of taproot VSP could be a marker for the cessation of N uptake and the initiation of a massive leaf senescence. Taken as a whole, the present results suggest that senescence that is directly or indirectly induced by externally supplied MeJa not only decreases photosynthesis and chlorophyll content of leaves but also promotes the hydrolysis of the large and small subunit of Rubisco. These observations match those previously reported in barley and rice where it has been clearly shown that jasmonate causes a reduction of the protein amounts of both subunits of Rubisco (Weidhase et al., 1987b; Reinbothe et al., 1993; Rakwal and Komatsu, 2000). Furthermore, the decrease in the levels of Rubisco and chlorophyll was also closely related with the decline of the photosynthetic activity (Rakwal and Komatsu, 2001). It further emphasizes the direct or indirect role that MeJa can exert on nitrogen metabolism in plants (from uptake, to Rubisco turnover). Studying the implication of this phytohormone on plant defence against pathogen it has been demonstrated that the increased synthesis of nicotine diverted quite a large amount of N from the normal pathways (Baldwin 1988a, b, 1989; Baldwin et al., 1994a, b, 1997). More recently, using Populus plants treated with airborne methyl jasmonate, a large modification of N distribution within plant tissues was shown which was also related to an increased accumulation of VSP (Beardmore et al., 2000). The vegetative plants used, devoid of any significant metabolic drain such as developing seeds, accumulated available N under the form of specific protein such as the 23 kDa VSP, and in tissues (taproot, stem) having potentially a larger life span than leaves. Proteins immunologically related to the 23 kDa VSP were also found in leaves, stems and flowers, but it remained difficult from the present results to determine the degree of homology of these different polypeptides. Whether or not MeJa is produced by Brassica napus L. during the flowering stage is still speculative although there is convincing evidence showing that amongst volatilized compounds from an oilseed rape culture, some like monoterpene cis-3-hexen-1-ol (McEwan and Macfarlane Smith, 1998), are probably derived from the MeJa biosynthetic pathway. Therefore, future work will be necessary to describe this phytohormone synthesis kinetically under field conditions. Nevertheless, the similarity between plant responses during flowering and seed filling, and the effects of exogenously applied MeJa remains intriguing. In both cases, nitrate uptake is inhibited, senescence promoted, mobilization of foliar N to seeds or alternatively to stem and taproot is increased while the 23 kDa VSP follows a typical pattern of accumulation/mobilization. This appears to be sufficient evidence to ascribe a potential death hormone role to MeJa and a marker role to the 23 kDa VSP for the cessation of N uptake and initiation of a massive leaf senescence in Brassica napus L.
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Acknowledgements |
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We are grateful to Patrick Beauclair (UMR INRA/UCBN Physiologie et Biochimie végétales, Université de Caen) for his invaluable help with soluble protein extraction. Part of this work has been funded through a grant from INRA/BBSRC.
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Footnotes |
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3 To whom correspondence should be addressed. Fax: +33 2 31 56 53 60. E-mail: ourry{at}ibba.unicaen.fr
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References |
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Anderson JM. 1989. Membrane derived fatty acids as precursors to second messengers. In: Boss WF, Morre DJ, eds. Second messengers in plant growth and development. New York: Alan R Liss, 181–212.
Baldwin IT. 1988a. The alkaloidal responses of wild tobacco to real and simulated herbivory. Oecologia 77, 378–381.[Web of Science]
Baldwin IT. 1988b. Short-term damage-induced increases in tobacco alkaloids protect plants. Oecologia 75, 367–370.
Baldwin IT. 1989. Mechanism of damage-induced alkaloid production in wild tobacco. Journal of Chemical Ecology 15, 1661–1680.[Web of Science]
Baldwin IT, Karb MJ, Ohnmeiss TE. 1994a. Allocation of 15N from nitrate to nicotine after leaf damage: production and turnover of a damage-induced mobile defense. Ecology 75, 1703–1713.[Web of Science]
Baldwin IT, Schmelz EA, Ohnmeiss TE. 1994b. Wound-induced changes in root and shoot jasmonic acid pools correlate with induced nicotine synthesis in Nicotiana sylvestris Spengazzini and Comes. Journal of Chemical Ecology 20, 2139–2157.[Web of Science]
Baldwin IT, Zhang ZP, Diab N, Ohnmeiss TE, McCloud ES, Lynds GY, Schmelz EA. 1997. Quantification, correlations, and manipulations of wound-induced changes in jasmonic acid and nicotine in Nicotiana sylvestris. Planta 201, 397–404.[Web of Science]
Beardmore T, Wetzel S, Kalous M. 2000. Interactions of airborne methyl jasmonate with vegetative storage protein gene and protein accumulation and biomass partitioning in Populus plants. Canadian Journal of Forest Research 30, 1106–1113.
Blake MS, Johnston KH, Russel-Jones GJ, Gotschlich EC. 1984. A rapid, sensitive method for detection of alkaline phosphatase conjugated anti-antibody on Western blots. Analytical Biochemistry 136, 175–179.[Web of Science][Medline]
Buchanan-Wollaston V. 1997. The molecular biology of leaf senescence. Journal of Experimental Botany 48, 181–199.
Clement CR, Hopper MJ, Canaway RJ, Jones LPH. 1974. A system for measuring the uptake of ions by plants from solutions of controlled composition. Journal of Experimental Botany 25, 81–99.
Clement CR, Hopper MJ, Jones LPH. 1978. The uptake of nitrate by Lolium perenne from flowing nutrient solution. Journal of Experimental Botany 109, 453–464.
Creelman RA, Mullet JE. 1995. Jasmonic acid distribution and action in plants: regulation during development and response to biotic and abiotic stress. Proceedings of the National Academy of Sciences, USA 92, 4114–4119.
Creelman RA, Mullet JE. 1997. Biosynthesis and action of jasmonates in plants. Annual Review of Plant Physiology and Plant Molecular Biology 48, 355–381.[Web of Science][Medline]
Engvild KC. 1989. The death hormone hypothesis. Physiologia Plantarum 77, 282–285.
Falkenstein E, Groth B, Mithofer A, Weiler EW. 1991. Methyl jasmonate and 
-linolenic acid are potent inducers of tendril coiling. Planta 185, 316–322.[Web of Science]
Farmer EE, Ryan CA. 1990. Interplant communication: airborne methyl jasmonate induces synthesis of proteinase inhibitors in plant leaves. Proceedings of the National Academy of Sciences, USA 87, 7713–7716.
Hatch DJ, Hopper MJH, Dhanoa MS. 1986. Measurement of ammonium ions in flowing solution culture and diurnal variation in uptake in Lolium perenne. Journal of Experimental Botany 37, 589–596.
He Y, Tang W, Swain JD, Green AL, Jack TP, Gan S. 2001. Networking senescence-regulating pathways by using Arabidopsis enhancer trap lines. Plant Physiology 126, 707–716.
Herman G, Lehmann J, Peterson A, Sembdner G, Weidhase RA, Parthier B. 1989. Species and tissue specificity of jasmonate-induced abundant proteins. Journal of Plant Physiology 134, 703–709.
Horton RF. 1991. Methyl jasmonate and transpiration in barley. Plant Physiology 96, 1376–1378.
Kelly MO, Davies PJ. 1988. The control of whole plant senescence. CRC Critical Review in Plant Sciences 7, 139–173.
Koda Y. 1994. The role of jasmonic acid and related compounds in the regulation of plant development. International Review of Cytology 135, 155–199.
Lopez R, Dathe W, Brückner C, Miersch O, Sembdner G. 1987. Jasmonic acid in different parts of the developing soybean fruit. Biochemie und Physiologie der Pflanzen 182, 195–201.
Manetas Y, Grammatikopoulos G, Kyparissis A. 1998. The use of portable, non-destructive, SPAD-502 (Minolta) chlorophyll meter with leaves of varying trichome density and anthocyanin content. Journal of Plant Physiology 153, 513–516.
McEwan M, Macfarlane Smith WH. 1998. Identification of volatile organic compounds emitted in the field by oilseed rape (Brassica napus L. spp. oleifera) over the growing season. Clinical and Experimental Allergy 28, 332–338.[Web of Science][Medline]
Müller-Uri F, Parthier B, Nover L. 1988. Jasmonate-induced alteration of gene expression in barley leaf segments analysed by in vivo and in vitro protein synthesis. Planta 176, 241–247.[Web of Science]
Noodén LD. 1988. Abscisic acid, auxin, and other regulators of senescence. In: Noodén LD, Leopold AC, eds. Senescence and aging in plants. San Diego: Academic Press, 329–368.
Noodén LD, Leopold AC. 1978. Phytohormones and the endogenous regulation of senescence and abscission. In: Letham DS, Goodwin PB, Higgins TJV, eds. Phytohormones and related compounds: a comprehensive treatise, Vol. 3. Amsterdam: Elsevier, 329–369.
Pelacho AM, Mingo-Castel AM. 1991. Jasmonic acid induces tuberization of potato stolons cultured in vitro. Plant Physiology 97, 1253–1255.
Rakwal R, Komatsu S. 2000. A role for jasmonate in the rice (Oryza sativa L.) self defense mechanism using proteome analysis. Electrophoresis 21, 2492–2500.[Web of Science][Medline]
Rakwal R, Komatsu S. 2001. Jasmonic acid-induced necrosis and drastic decreases in ribulose-1,5-bisphosphate carboxylase/oxygenase in rice seedlings under light involves reactive oxygen species. Journal of Plant Physiology 158, 679–688.[Web of Science]
Reinbothe S, Mollenhauer B, Reinbothe C. 1994. JIPs and RIPs: the regulation of plant gene expression by jasmonates in response to environmental cues and pathogens. The Plant Cell 6, 1197–1209.[Web of Science][Medline]
Reinbothe S, Reinbothe C, Heintzen C, Seidenbeicher C, Parthier B. 1993. A methyl jasmonate-induced shift in the length of the 5' untranslated region impairs translation of the plastid rbcL transcript in barley. EMBO Journal 12, 1505–1512.[Web of Science][Medline]
Rossato L, Laine P, Ourry A. 2001a. Nitrogen storage and remobilization in Brassica napus L. during the growth cycle: nitrogen fluxes within the plant and changes in soluble protein patterns. Journal of Experimental Botany 52, 1655–1663.
Rossato L, Le Dantec C, Laine P, Ourry A. 2001b. Nitrogen storage and remobilization in Brassica napus L. during the growth cycle: identification, characterization and immunolocalization of a putative taproot storage glycoprotein. Journal of Experimental Botany In press.
Sembdner G, Parthier B. 1993. The biochemistry and the physiological and molecular actions of jasmonates. Annual Review of Plant Physiology and Plant Molecular Biology 44, 569–589.[Web of Science]
Smart CM. 1994. Gene expression during leaf senescence. New Phytologist 126, 419–448.[Web of Science]
Staswick PE. 1990. Novel regulation of vegetative storage protein genes. The Plant Cell 2, 1–6.[Medline]
Staswick PE. 1992. Jasmonate, genes, and fragrant signals. Plant Physiology 99, 804–807.
Staswick PE. 1994. Storage proteins of vegetative plant tissues. Annual Review of Plant Physiology and Plant Molecular Biology 45, 303–322.[Web of Science]
Staswick PE, Huang J, Rhee Y. 1991. Nitrogen and methyl jasmonate induction of soybean vegetative storage protein genes. Plant Physiology 96, 130–136.
Towbin H, Staehelin T, Gordon J. 1979. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proceedings of the National Academy of Sciences, USA 76, 4350–4354.
Weidhase RA, Kramell HM, Lehmann J, Liebisch HW, Lerbs W, Parthier B. 1987a. Methyl jasmonate-induced changes in the polypeptide pattern of senescing barley leaf segments. Plant Science 51, 177–186.
Weidhase RA, Lehmann J, Kramell H, Sembdner G, Parthier B. 1987b. Degradation of ribulose-1–5-bisphosphate-carboxylase and chlorophyll in senescing barley leaf segments triggered by jasmonic acid methyl ester and counteraction by cytokinin. Physiologia Plantarum 69, 161–166.
Wierstra I, Kloppstech K. 2000. Differential effects of methyl jasmonate on the expression of the early light-inducible proteins and other light-regulated genes in barley. Plant Physiology 124, 833–844.
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