Free access
Issue
Ann. For. Sci.
Volume 67, Number 6, September 2010
Article Number 609
Number of page(s) 8
DOI http://dx.doi.org/10.1051/forest/2010022
Published online 08 July 2010

© INRA, EDP Sciences, 2010

1. INTRODUCTION

Along with carbon (C), nitrogen (N) is the most significant resource for plant functioning. In temperate ecosystems, soil nitrogen is a limiting element for plant growth because its concentration is kept low due to losses by leaching and microbial consumption (Tischner, 2000). N uptake occurs predominantly in roots by nitrate () and/or ammonium () absorption (Pate, 1973). In forest trees, N assimilation occurs mainly in roots while it occurs both in leaves and roots of herbaceous plants (Martin and Plassard, 1997). Whereas nitrates typically predominate in agricultural soils presenting high N turnover, neutral pH and good aeration, ammonium plays an important role in forest soils (Glass and Siddiqi, 1995). As is the case in herbaceous plants (Lee and Drew, 1989), nitrate uptake by roots of most tree species is strongly inhibited when ammonium is available (Gessler et al., 1998; Kreuzwieser et al., 1997; Rennenberg et al., 1996). However, in some species like ash and oak, ammonium has a positive effect on nitrate uptake (Stadler et al., 1993). In trees, ammonium uptake generally shows a seasonal course with maximum uptake rates occurring in mid-summer (Gessler et al., 1998). Nitrogen uptake during bud break in spring is very low because the N demand can be almost completely satisfied by N remobilized from storage tissues (Millard, 1994; Millard and Proe, 1992). Later, during the fall, a large part of leaf N is stored in shoots as vegetative storage proteins (VSPs) that constitute the predominant proteins in dormant organs, especially in the woody compartment (Rowland and Arora, 1997; Stepien et al., 1994). In summer, bark apparently works as a transit area for nitrogen components, in particular, for the free amino acids synthesized in roots and carried in sap (Gomez and Faurobert, 2002).

The woody compartment is known to be an important source of C loss by respiration (Damesin et al., 2002; Granier et al., 2000), but stems are also able to assimilate C (Kharouk et al., 1995; Pearson and Lawrence, 1958; Pfanz et al., 2002; Pilarski, 1989; Wiebe, 1975). This photosynthesis process occurs particularly in the bark chlorenchyma and mainly consists in refixation of C released by respiration (Cernusak and Marshall, 2000; Damesin, 2003). Refixation of CO2 in young stems may compensate for 60–100% of the potential respiratory C loss and may sometimes exceed CO2 release (Berveiller et al., 2007a; Damesin, 2003; Wittmann et al., 2001). The other processes potentially involved in the CO2 release under illumination, i.e. photorespiration and mitochondrial respiration, do not play a major role in stem carbon exchange (Wittmann et al., 2006). Although of a lower magnitude than in leaves, some ecophysiological characteristics of stem photosynthesis are similar to those observed in leaves, such as the positive response of gas exchange to light and temperature (Wittmann and Pfanz, 2007; Wittmann et al., 2001), the positive response of electron transport rate to light (Manetas, 2004), and the presence of Rubisco (Berveiller and Damesin, 2008; Berveiller et al., 2007b; Schaedle and Brayman, 1986). Over a whole season, current-year stems of European beech (Fagus sylvatica) are able to assimilate the equivalent of 40% of the C lost by respiration (Damesin, 2003).

A large number of papers studying the interaction between C- and N-metabolisms in leaves are available. The pathways of the two elements depend on each other and both pathways are regulated by each other. The plant allocates N to tissue maintenance and protein turnover (Ryan, 1991) and to leaf photosynthesis, especially to Rubisco and thylakoïds. Strong correlations exist between C assimilation rate or Rubisco activity and leaf N concentration (Evans, 1989). Phosphoenolpyruvate (PEP) carboxylase is an important cross point between the two metabolisms as it is involved in supplying oxaloacetate to the Krebs cycle, thus ensuring a continuous replenishment of C4-dicarboxylic acid, which is necessary for nitrogen assimilation and amino acid biosynthesis (Huppe and Turpin, 1994). We have recently shown a very high activity of PEP carboxylase in young stems in comparison to leaves of adult European beeches (Berveiller and Damesin, 2008; Berveiller et al., 2007b). Stem photosynthesis is positively correlated to nitrogen concentration in an interspecific study (Berveiller et al., 2007a). Contrary to leaves, interactions between C- and N-metabolisms in woody stems are poorly documented, with the exception of the role of woody tissues as a N storage compartment during winter. In this context, we conducted an experiment on young European beeches planted in pots with three contrasted levels of N fertilization. Our goal was to determine if N had an impact on stem carbon functioning, especially on CO2 exchange and carboxylating enzymes.

2. MATERIALS AND METHODS

2.1. Site description, plant materials and nitrogen supplies

The study was conducted on the experimental field of the University of Paris-Sud (48° 42′ N, 02° 10′ E), 25 km south-west of Paris, France. A set of nine four-year-old European beech trees (Fagus sylvatica L.) were planted outdoors in 90 L pots in December 2005. The soil substrate consisted of a mixture of 50% (v/v) peat and 50% (v/v) sand; its characteristics are detailed in Table I. It was a sandy soil, poor in nitrogen, with a high C/N ratio and rather neutral-basic. All trees were watered every two weeks (depending on rainfall) in 2006 without any nitrogen supply.

Table I

Characteristics of the soil substrate used when planting beech saplings, before any N fertilization. Measurements were achieved from soil aliquots from five pots for the texture and from soil aliquots from nine pots for each other characteristic. Data are means ±1 SE.

In March 2007, i.e. one month before budburst, all trees were fertilized (“Opta Fond Plus” (BHS, Vemar, France) NPK 10–15–20 + 25% SO3 + trace elements, 20 g per pot). At the same time, three trees received an additional supply 7.5 g fertilizer per pot (“Optacote 40N” (BHS, Vemar, France) NPK 40–0–0) and three other trees 20 g per pot of “Optacote 40N”. “Opta Fond Plus” and “Optacote 40N” consist of coated and compacted granules that slowly release nitrogen into the soil during six month. In this way, three treatments (with three trees per treatment) were achieved (expressed in grams of each element):

  • Treatment T1: NPK 2–3–4,

  • Treatment T2: NPK 5–3–4,

  • Treatment T3: NPK 10–3–4.

All measurements detailed below were carried out on whole current-year stems, as the chlorophyll cells likely to refixate CO2 are known to be present even up to the pith of current-year stems of Fagus sylvatica (Berveiller et al., 2007a).

Table II

Mass-based N concentrations (mg g−1 DW) and leaf or stem mass per area (LMA and SMA, g DW m−2) of leaves and current-year stems of Fagus sylvatica L. in each N treatment (T1, T2, and T3). Data are means ±1 SE with n = 3. For each organ (leaves or stems), letters (a, b, and c) indicate significant differences between N treatments.

2.2. Gas-exchange measurements

Gas-exchange measurements were performed in the laboratory with a LI-6400 portable photosynthesis system (LI-COR Biosciences, Lincoln, NE) equipped with the conifer chamber Model 6400-05. In mid-June 2007, three current-year stems of each tree were harvested. Detached stems were immediately recut under distilled water in the section of tissue produced the previous year and the current-year parts of stem were oriented in the chamber so that the top of the stems faced the light source (2 × 1000 W HQI, Osram, Munich, Germany). Measurements were made at 20 °C, with a CO2 concentration of 390 μmol mol−1, 60% relative humidity and 1400 or 0 μmol m−2 s−1 of photosynthetically active radiation (PAR) to determine CO2 efflux rates in the light (Rl) and in the dark, i.e., dark respiration rate (Rd) (for further details of method used for gas exchange see Berveiller et al., 2007a). Assuming that a light-induced inhibition of respiration is unlikely to occur (Wittmann et al., 2006), stem gross photosynthesis (Pg) may be calculated as: (1)From gas exchange measurements, the refixation rate was calculated as follows: (2)After gas-exchange measurements, total surface areas of stems (not projected surface areas) were measured to express CO2 efflux in terms of area units. Stem tissues were then dried at 60 °C for 48 h, weighed and ground for nitrogen concentration measurements (see Sect. 2.3).

2.3. Nitrogen concentration

At the end of June 2007, three leaves and three current-year stems of each tree were sampled and dried at 60 °C for 48 h. Dry mass (DW) of each sample (sampled in mid-June for gas exchange measurement, see above, and at the end of June) was determined. The samples were then cut into small pieces and ground to a powder (MM200, RETSCH, Haan, Germany). The nitrogen concentration of the powder was determined with an elemental analyzer at the “Service Central d’Analyse du CNRS” (Vernaison, France).

2.4. Protein extraction

Frozen materials (500 mg) of leaves and whole stem segments were weighed rapidly, powdered with a pestle in a pre-chilled mortar under liquid N2 and ground in an extraction buffer containing 100 mM HEPES (pH 8), 10 mM MgCl2, 7 % (w/w) polyethylene glycol 20000, 10 mM dithiothreitol, 10% (v/v) glycerol, three antiproteases (20 μM 4-amidinophenylmethanesulfonyl fluoride (p-APMSF), 1 μM pepstatin, 1 μM leupeptin) and an antiphosphatase cocktail (Sigma). During grinding, polyvinylpolypyrrolidone (PVPP) was added. The homogenates were centrifuged for 25 min at 4 °C in a microcentrifuge (5 804 R; EPPENDORF, Germany) at 15 000 g. An aliquot of the supernatant was used for protein determination. The remainder was passed through a desalting column (Sephadex G25TM Pharmacia; AMERSHAM, Sweden) for enzyme assays.

2.5. Enzyme assays

According to Berveiller et al. (2007b), PEP carboxylase activity was measured at pH 7.8 in a buffer containing 100 mM HEPES, 10 mM MgCl2, 5 mM NaHCO3, 0.2 mM NADH, and 3 U mL−1 malate dehydrogenase. This assay was initiated by the addition of 5 mM phosphoenolpyruvate (Tietz and Wild, 1991; Uedan and Sugiyama, 1976).

Rubisco initial and total activities were measured in a buffer containing 100 mM HEPES (pH 8) , 20 mM MgCl2, 25 mM NaHCO3, 3.5 mM ATP, 0.2 mM NADH, 5 mM creatine phosphate, 5 U mL−1 glyceraldehyde-3-phosphate dehydrogenase, 5 U mL−1 3-phosphoglycerokinase, 5 U mL−1 creatine phosphokinase. The assays were initiated by addition of 0.5 mM ribulose-1,5-bisphosphate for the Rubisco initial activity and after 15 min incubation of the extract at 30 °C for the Rubisco total activity.

PEP carboxylase and Rubisco activities were determined by UV spectrometry following NADH oxidation at 340 nm. Activities were expressed in μmol CO2 or consumed per min and per mg proteins for Rubisco and PEP carboxylase respectively. Soluble protein concentrations were determined by the Bradford assay (Bio-Rad, Hercules, CA, USA) using bovine serum albumin as a standard (Bradford, 1976).

2.6. Statistical analysis

Statistical analyses were performed with Statistica (Statsoft Inc., Tulsa, OK). The statistical significance of N concentration, gas exchange, and enzyme activities between the three nitrogen treatments was calculated with analysis of variance (ANOVA). Treatment effects were considered significant if P < 0.05 (Fisher’s LSD test). A non-linear relationship was fitted between gross photosynthetic rate and nitrogen concentration using the following relationship: (3)where Pg is the gross photosynthetic rate, N is the total nitrogen concentration, and a, b, and c, the three parameters obtained by ordinary least-squares estimation.

3. RESULTS

3.1. Effect of N treatment on the N status of above-ground parts

In treatment T1, beech trees accumulated nitrogen in above-ground parts with values reaching 10.2 mg g−1 dry weight (DW) in leaves and 5.67 mg g−1 DW in current-year stems (Tab. II). Additional supplies of nitrogen in soil (in treatments T2 and T3) resulted in a significant increase of nitrogen concentration in the leaves and current-year stems, expressed either in mass-based or area-based units (for stems). The nitrogen concentration reached 17.2 and 10.1 mg g−1 DW in treatment T2 and 14.1 and 8.00 mg g−1 DW in treatment T3, for leaves and current-year stems, respectively. No significant differences were observed between treatments T2 and T3 either for leaves or for current-year stems.

3.2. Effect of N treatment on the gas exchange of current-year stems

Whatever the treatment, we observed a reduction of stem CO2 efflux upon illumination, i.e. 60%, 74%, and 68% in treatments T1, T2 and T3, respectively (Fig. 1). Upon illumination, Rl ranged from –0.71 μmol CO2 m−2 s−1 (treatment T1) to –0.95 μmol CO2 m−2 s−1 (treatment T3) and no significant difference was observed between treatments. In the treatment T1, dark respiration rate (Rd) and gross photosynthesis rate (Pg) were equal to 1.79 and 1.07 μmol m−2 s−1, respectively. The additional supply of nitrogen (treatments T2 and T3) resulted in a significant increase of gas exchange intensity. Rd increased rates by 83% and 66% between treatments T1 and T2 and between T1 and T3, respectively; Pg increased rates by 126% and 90% between treatments T1 and T2 and between T1 and T3, respectively.

thumbnail Figure 1

Gas exchange of current-year stems of Fagus sylvatica L. measured as CO2 efflux in the dark (dark respiration rate, Rd), CO2 efflux under saturated light (Rl) and calculated rates of gross photosynthesis (Pg), for each N treatment (T1, T2, and T3). Data are means ±1 SE, with n = 3. Letters (a, b, and c) indicate significant differences between N treatments, for each parameter (Rd, Rl and Pg).

3.3. Relationships between gross photosynthesis rate, respiration rate and N concentration in current-year stems

Pooling data from all treatments (expressed in area-based units), the linear relationship observed between gross photosynthesis rate and dark respiration rate was significant (r2 = 0.88; P < 0.0001, Fig. 2). Carbon refixation by stem photosynthesis represented 80% of the total respiration rate. Expressed in mass-based units, the relationship was also significant with r2 = 0.84 (P < 0.0001, data not shown). Considering each treatment separately, the following refixation rates were obtained: 58.5% ± 4.66 for T1, 74.3% ± 1.85 for T2 and 68.2 ± 2.17 for T3.

thumbnail Figure 2

Relationship between area-based gross photosynthesis (Pg) and area-based dark respiration rate (Rd) of current-year stems of Fagus sylvatica L. trees subjected to different nitrogen treatments. Each value corresponds to one of the three current-year stems sampled on each of the nine trees (three per N treatment).

Gross photosynthesis rates were highly related with the nitrogen concentrations values in current-year stems of European beech (r2 = 0.83; P < 0.0001, Fig. 3). The response of gross photosynthesis to nitrogen increase followed a hyperbolic-type model. From this non-linear model, the maximum gross photosynthetic rate for saturating nitrogen concentration was calculated as 9.73 μmol CO2 kg−1DW s−1. By extrapolating, the mass-based hyperbole crossed the x-axis at 4.4 mg N g−1 DW (0.3 mmol N g−1 DW).

3.4. Effect of N concentration on soluble protein content and enzyme activities

The total soluble protein content increased with increasing tissue N concentration (Fig. 4). The amount of protein ranged from 6.0 mg g−1 DW for treatment T1 to 10 mg g−1 DW for treatment T2 for stems and from 19 mg g−1 DW in treatment T1 to 34 mg g−1 DW in treatment T2 for leaves.

Whatever the stem N concentration, Rubisco total activity was always higher in leaves than in current-year stems (by a factor of about 10–12 times, Fig. 5a). In average, activity in leaves ranged from 10.2 to 11 μmol CO2 min−1 mg−1 prot and from 0.85 to 0.91 μmol CO2 min−1 mg−1 prot in current-year stems. No significant effect of stem N concentration was observed on Rubisco activities, either in leaves or in current-year stems.

thumbnail Figure 3

Relationship between mass-based gross photosynthesis (Pg) and mass-based nitrogen concentration of current-year stems of Fagus sylvatica L. trees subjected to different nitrogen treatment. Each value corresponds to one of the three current-year stems sampled on each of nine trees (three per treatment).

thumbnail Figure 4

Relationship between mass-based total soluble protein content and mass-based nitrogen concentration of leaves (black-filled circles) and current-year stems (open circles) of Fagus sylvatica L. Each point corresponds to the mean value of each N treatment and mean value of protein content (n = 3 for each ones). The standard errors of the ordinate and abscissa values are shown as vertical and horizontal bars, respectively.

PEP carboxylase activity was from 1.9 to 3.4 times higher in current-year stems than in leaves (Fig. 5b). The PEP carboxylase activity in leaves ranged from 1.4 μmol min−1 mg−1 prot in treatment T3 to 1.8 μmol min−1 mg−1 prot in treatment T2. In current-year stems, PEP carboxylase activity significantly increased with N concentration, ranging from 3.4 μmol min−1 mg−1 prot at 5.67 mg N g−1 DW to 5.4 μmol min−1 mg−1 prot at 10.1 mg N g−1 DW.

thumbnail Figure 5

Relationships between Rubisco total activity and mass-based nitrogen concentration (a) and between PEP carboxylase total activity and mass-based nitrogen concentration (b) of leaves (black-filled circles) and current-year stems (open circles) of Fagus sylvatica L. Each point corresponds to the mean value of a N treatment and mean value of enzyme activity (n = 3 for each ones). The standard errors of the ordinate and abscissa values are shown as vertical and horizontal bars, respectively.

4. DISCUSSION

Carbon (C) and nitrogen (N) management in perennial species like European beech is complex because of the use and remobilization of N reserves, especially during bud-burst period or fall, and in case of stress (Millard, 1994). It is generally difficult to obtain a large and significant effect of a soil N treatment on the metabolism and growth of trees. In our study, all trees were initially planted in a sandy soil having a low N content, without fertilizer addition throughout 2006. This certainly resulted in a decrease of C- and N- reserves of trees before the 2007 experiment. Although N treatments did not affect the structure of leaves or stems (no significant difference in leaf mass per area or stem mass per area with treatments), we observed a positive effect of N supply on the N status of trees with a significant increase in total N concentrations in both leaves and current-year stems (Tab. II). The N concentrations measured in leaves and stems for the treatment T2 were similar to values commonly observed on current-year stems of adult trees at the beginning of summer, i.e. 20 mg g−1 DW and 10 mg g−1 DW for leaves and current year stems, respectively (Berveiller et al., 2007a; Damesin, 2003). In both stems and leaves, the N concentrations similarly increased between treatments T1 and T2 (×1.7) or between T1 and T3 (×1.4). The amount of N supplied by treatment T3 seemed to be excessive because N accumulation in leaves and stems was lower for trees of the treatment T3 than for trees of the treatment T2. The N source of the fertilizer that we used is mainly composed of urea that releases ammonium () through urease enzyme activity. Because absorption by roots uses an /H+ antiport channel, soil pH decreases when the soil is watered only with ammonium solution (Volk et al., 1992). As a result, a high fertilization level (case of the treatment T3) probably resulted in a decrease in pH, thus limiting the N uptake by plant roots and N accumulation in leaves and current-year stems.

The reduction of stem CO2 efflux when a stem is transferred from dark to light is now well known and is mainly due to C assimilation by stem photosynthesis (Pfanz et al., 2002). Stem photosynthesis allows young stems of Fagus sylvatica to compensate for 60–100% of their CO2 loss by respiration, which corresponds to the refixation rate (Berveiller et al., 2007a; Damesin, 2003; Wittmann et al., 2001). Considering data from all N treatments of this study, similar results were found with a linear relationship between gross photosynthesis rate and dark respiration rate. This suggests that, like in leaves, products of stem photosynthesis are partially and directly used as substrates for respiration to sustain protein turn-over (McCree, 1969). An alternative explanation comes from the results obtained by Cernusak and Marshall (2000) who showed (1) a strong relationship between stem internal CO2 concentration measured in the dark and gross photosynthesis measured at saturated photon flux density in PAR (Pg) on young stems of Western White Pines, but (2) no significant correlation between stem internal CO2 concentration measured in the light and Pg. These authors suggested that stems allocate their photosynthetic capacity to refixation depending on the dark respiration rates which lead to an efficient use of N for recycling respired CO2. This hypothesis is supported by our measurements which show that gross photosynthesis increases with increasing N concentration of the whole current-year stem. The relationship between light-saturated photosynthetic rate and N concentration is widely documented for leaves (Evans, 1989; Field and Mooney, 1986). This relationship exists between stems of various tree species (Berveiller et al., 2007a; 2007b) and this study shows that it is valid at an intraspecific level. This relationship is generally linear, but in our study, the mass-based and area-based relationships were found to be hyperbolic. By calculating the photosynthetic nitrogen use efficiency (PNUE, calculated as C assimilation rate to N concentration ratio) on a projected area basis, we showed that it increased from 18.8 to 22.7 and 24.4 μmol CO2 mol−1N s−1, in treatments T1, T2 and T3 respectively. These PNUE values are close to previous measurements performed on current-year stems of adult trees (Berveiller et al., 2007a) and fairly similar to values commonly observed in conifer needles (e.g. Gower et al., 1993) but much lower than in broad-leaved trees (Poorter and Evans, 1998). A low N investment in stem photosynthesis and conifer needle could be the consequence of the long lifespan of these tissues that invest more N in compounds required for longevity and defense (Field and Mooney, 1986; Hikosaka et al., 1998).

The photosynthetic capacity is generally related to N concentration primarily because the proteins of the Calvin cycle and thylakoïds represent the majority of leaf nitrogen (Evans, 1989). Rubisco is the major carboxylating enzyme in leaves and its activity is highly correlated to leaf N concentration (Sage et al., 1987). Whereas soluble protein content in beech leaves and stems doubled in response to the rise in tissue N concentration (Fig. 4), the N treatment had no significant effect on Rubisco either in stems or leaves (Fig. 5a). The rise in soluble protein content might at least partly explain the relationship observed between respiration and nitrogen concentration (data not shown). It has been reported that allocation of N to Rubisco increases in leaves of herbaceous species with increasing soil N availability (Evans, 1989; Makino et al., 1994), but not in trees (Bauer et al., 2001; Ripullone et al., 2003). The activation level of Rubisco being very similar in stems and leaves (69% and 71% respectively, data not shown), it could be calculated that Rubisco activities mirrored the concentrations of Rubisco in both tissues. As a result, Rubisco concentrations were twenty times lower in stems than in leaves and did not change with nitrogen availability. Thus, N seems to be allocated to other proteins than Rubisco in stems compared to leaves, without invalidating the positive impact of N availability on stem respiration that is directly associated to protein synthesis and protein turnover (Ryan, 1991). A likely stem N sink candidate is PEP carboxylase. As shown in previous studies (Berveiller and Damesin, 2008; Berveiller et al., 2007a; 2007b), PEP carboxylase activity was higher in stems than in leaves, suggesting that this enzyme is involved in stem photosynthesis (Fig. 5b). Whereas no significant differences were observed in leaf PEP carboxylase activities among nitrogen treatments, the PEP carboxylase activity increased up to × 1.6 in stems depending on tissue N concentration. This observation is comparable to the positive relationship observed between stem gross photosynthesis and N concentration (Fig. 3). Leaf and root PEP carboxylase is known to be stimulated by nitrogen supply because the enzyme is involved in supplying carbon skeletons to the Krebs cycle (anaplerotic carbon fixation), thus ensuring a continuous replenishment of C4-dicarboxylic acid, which is necessary for nitrogen assimilation and amino acid biosynthesis (Huppe and Turpin, 1994). Our previous work showed that the stem PEP carboxylase is similar to the leaf PEP carboxylase of Fagus sylvatica, particularly regarding the biochemical characteristics of the enzyme (Berveiller et al., 2007b). In leaves, PEP carboxylase is activated under illumination, following its phosphorylation by a light-sensitive PEP carboxylase kinase (see e.g. Li et al., 1996). To confirm the important role of the enzyme in stem carbon assimilation, it would be interesting to examine if the stem PEP carboxylase can be activated under illumination, as is the case in leaves.

5. CONCLUSION

As commonly observed in leaves, an increase in soil nitrogen supply had a positive impact on stem gas exchange and stem carbon refixation. The current-year stem seems to invest more N in CO2 refixation when more N is available, especially through PEP carboxylase whose activity increased. PEP carboxylase is probably involved in carbon refixation in stems and could concurrently supply N assimilation with carbon skeletons. Further investigations using 13C as a tracer could be helpful to quantify the contribution of assimilated carbon to the various functions of the stem.

Acknowledgments

This research was financed by the “Programme National ACI/FNS ECCO, PNBC” (convention No. 0429 FNS). The ESE laboratory is supported by the University of Paris-Sud, the Centre National de la Recherche Scientifique (CNRS), and AgroParisTech. Two anonymous referees are thanked for constructive comments.

References

  • Bauer G.A., Berntson G.M. and Bazzaz F.A., 2001. Regenerating temperate forests under elevated CO2 and nitrogen deposition: comparing biochemical and stomatal limitation of photosynthesis. New Phytol. 152: 249–266. [CrossRef] (In the text)
  • Berveiller D. and Damesin C., 2008. Carbon assimilation by tree stems: potential involvement of phosphoenolpyruvate carboxylase. Trees-Struct. Func. 22: 149–157. [CrossRef] (In the text)
  • Berveiller D., Kierzkowski D. and Damesin C., 2007. Interspecific variability of stem photosynthesis among tree species. Tree Physiol. 27: 53–61. [PubMed] (In the text)
  • Berveiller D., Vidal J., Degrouard J., Ambard-Bretteville F., Jaillard D. and Damesin C., 2007. Tree stem phosphoenolpyruvate carboxylase (PEPC): lack of biochemical and localization evidence for a C4-like photosynthesis system. New Phytol. 176: 775–781. [CrossRef] [PubMed] (In the text)
  • Bradford M.M., 1976. A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principe of protein-dye binding. Anal. Biochem. 72: 248–254. [CrossRef] [PubMed] (In the text)
  • Cernusak L.A. and Marshall J.D., 2000. Photosynthetic refixation in branches of Western Pine. Funct. Ecol. 14: 300–311. [CrossRef] (In the text)
  • Damesin C., 2003. Respiration and photosynthesis characteristics of current-year stems of Fagus sylvatica: from the seasonal pattern to an annual balance. New Phytol. 158: 465–475. [CrossRef] (In the text)
  • Damesin C., Ceschia E., Le Goff N., Ottorini J.-M. and Dufrêne E., 2002. Stem and branch respiration of beech: from tree measurements to estimations at the stand level. New Phytol. 158: 159–172. [CrossRef] (In the text)
  • Evans J.R., 1989. Photosynthesis and nitrogen relationships in leaves of C3 plants. Oecologia 78: 9–19. [CrossRef] (In the text)
  • Field C. and Mooney H.A., The photosynthesis-nitrogen relationship in wild plants, 1986. In: Givnish T.J. (Ed.), On the economy of plant form and function, Cambridge University Press, Cambridge, pp. 25–55. (In the text)
  • Gessler A., Schneider S., Von Sengbusch D., Weber P., Hanemann U., Huber C., Rothe A., Kreutzer K. and Rennenberg H., 1998. Field and laboratory experiments on net uptake of nitrate and ammonium by the roots of spruce (Picea abies) and beech (Fagus sylvatica) trees. New Phytol. 138: 275–285. [CrossRef] (In the text)
  • Glass A.D.M. and Siddiqi M.Y., Nitrogen absorption by plant roots, 1995. In: Strivastava H.S. and Singh R.P. (Eds.), Nitrogen Nutrition in Higher Plants, Associated Publishing Co., New Delhi, pp. 21–56. (In the text)
  • Gomez L. and Faurobert M., 2002. Contribution of vegetative storage proteins to seasonal nitrogen variations in the young shoots of peach trees (Prunus persica L. Batsch). J. Exp. Bot. 53: 2431–2439. [CrossRef] [PubMed] (In the text)
  • Gower S.T., Reich P.B. and Son Y., 1993. Canopy dynamics and aboveground production of five tree species with different leaf longevities. Tree Physiol. 12: 327–345. [PubMed] (In the text)
  • Granier A., Ceschia E., Damesin C., Dufrene E., Epron D., Gross P., Lebaube S., Le Dantec V., Le Goff N., Lemoine D., Lucot E., Ottorini J.-M., Pontailler J.-Y. and Saugier B., 2000. The carbon balance of a young Beech forest. Funct. Ecol. 14: 312–325. [CrossRef] (In the text)
  • Hikosaka K., Hanba Y.T., Hirose T. and Terashima I., 1998. Photosynthetic nitrogen-use efficiency in leaves of woody and herbaceous species. Funct. Ecol. 12: 896–905. [CrossRef] (In the text)
  • Huppe H.C. and Turpin D.H., 1994. Integration of Carbon and Nitrogen Metabolism in Plant and Algal Cells. Annu. Rev. Plant Physiol. Plant Mol. Biol. 45: 577–607. [CrossRef] (In the text)
  • Kharouk V.I., Middleton E.M., Spencer S.L., Rock B.N. and Williams D.L., 1995. Aspen bark photosynthesis and its significance to remote sensing and carbon budget estimates in the boreal ecosystem. Water Air Soil Pollut. 82: 483–497. [CrossRef] (In the text)
  • Kreuzwieser J., Herschbach C., Stulen I., Wiersema P., Vaalburg W. and Rennenberg H., 1997. Interactions of Formula + and L-glutamate with Formula transport processes of non-mycorrhizal Fagus sylvatica roots. J. Exp. Bot. 48: 1431–1438. [CrossRef] (In the text)
  • Lee R.B. and Drew M.C., 1989. Rapid, reversible inhibition of nitrate influx in barley by ammonium. J. Exp. Bot. 40: 741–752. [CrossRef] (In the text)
  • Li B., Zhang X.Q. and Chollet R., 1996. Phosphoenolpyruvate carboxylase kinase in tobacco leaves is activated by light in a similar but not identical way as in maize. Plant Physiol. 111: 497–505. [PubMed] (In the text)
  • Makino A., Nakano H. and Mae T., 1994. Responses of Ribulose-1,5-Bisphosphate Carboxylase, Cytochrome-F, and Sucrose synthesis enzymes in rice leaves to leaf Nitrogen and their relationships to photosynthesis. Plant Physiol. 105: 173–179. [PubMed] (In the text)
  • Manetas Y., 2004. Probing corticular photosynthesis through in vivo chlorophyll fluorescence measurements: evidence that high internal CO2 levels suppress electron flow and increase the risk of photoinhibition. Physiol. Plant. 120: 509–517. [CrossRef] [PubMed] (In the text)
  • Martin F. and Plassard C., Assimilation de l’azote par les symbioses ectomycorhiziennes, 1997. In: Morot-Gaudry J.-F. (Ed.), Assimilation de l’azote des chez les plantes, INRA, Paris, pp. 179–193. (In the text)
  • McCree K.J., An equation for the rate of respiration of white clover plants grown under controlled conditions, Proceedings of the technical meeting IBP, Centre for Agricultural Publishing and Documentation, Wageningen, Trebon (CSK), 1969, pp. 221–229. (In the text)
  • Millard P., 1994. Measurements of the remobilization of nitrogen for spring leaf growth of trees under field conditions. Tree Physiol. 14: 1049–1054. [PubMed] (In the text)
  • Millard P. and Proe M.F., 1992. Storage and internal cycling of N in relation to seasonal growth of sitka spruce. Tree Physiol. 10: 33–43. [PubMed] (In the text)
  • Pate J.S., 1973. Uptake, assimilation and transport of nitrogen compounds by plants. Soil Biology and Biochemistry 5: 109–119. [NASA ADS] [CrossRef] [EDP Sciences] [MathSciNet] [PubMed] (In the text)
  • Pearson L.C. and Lawrence D.B., 1958. Photosynthesis in aspen bark. Am. J. Bot. 45: 383–327. [CrossRef] (In the text)
  • Pfanz H., Aschan G., Langenfeld-Heyser R., Wittmann C. and Loose M., 2002. Ecology and ecophysiology of tree stems: corticular and wood photosynthesis. Naturwissenschaften 89: 147–162. [CrossRef] [PubMed] (In the text)
  • Pilarski J., 1989. Photosynthesis in shoots and leaves of lilac (Syringae vulgaris L.). Bull. Pol. Acad. Sci. Biol. Sci. 37: 261–269. (In the text)
  • Poorter H. and Evans J.R., 1998. Photosynthetic nitrogen-use efficiency of species that differ inherently in specific leaf area. Oecologia 116: 26–37. [CrossRef] (In the text)
  • Rennenberg H., Schneider S. and Weber P., 1996. Analysis of uptake and allocation of nitrogen and sulphur compounds by trees in the field. J. Exp. Bot. 47: 1491–1498. [CrossRef] (In the text)
  • Ripullone F., Grassi G., Lauteri M. and Borghetti M., 2003. Photosynthesis-nitrogen relationships: interpretation of different patterns between Pseudotsuga menziesii and Populus x euroamericana in a mini-stand experiment. Tree Physiol. 23: 137–144. [PubMed] (In the text)
  • Rowland L.J. and Arora R., 1997. Proteins related to endodormancy (rest) in woody perennials. Plant Sci. 126: 119–144. [CrossRef] (In the text)
  • Ryan M.G., 1991. Effects of climate on change on plant respiration. Ecol. Appl. 1: 157–167. [CrossRef] (In the text)
  • Sage R.F., Pearcy R.W. and Seemann J.R., 1987. The nitrogen use efficiency of C3 and C4 Plants: 3. Leaf nitrogen effects on the activity of carboxylating enzymes in Chenopodium album (L.) and Amaranthus retroflexus (L.). Plant Physiol. 85: 355–359. [CrossRef] [PubMed] (In the text)
  • Schaedle M. and Brayman A., 1986. Ribulose-1,5-bisphosphate carboxylase activity of Populus tremuloides Michx. bark tissues. Tree Physiol. 1: 53–56. [PubMed] (In the text)
  • Stadler J., Gebauer G. and Schulze E.D., 1993. The influence of ammonium on nitrate uptake and assimilation in 2-year-old ash and oak trees - a tracer study with 15N. Isotopenpraxis 29: 85–92. (In the text)
  • Stepien V., Sauter J.J. and Martin F., 1994. Vegetative storage proteins in woody-plants. Plant Physiol. Biochem. 32: 185–192. (In the text)
  • Tietz S. and Wild A., 1991. Investigations on the phosphoenolpyruvate carboxylase activity of spruce needles relative to the occurrence of novel forest decline. J. Plant Physiol. 137: 327–331. (In the text)
  • Tischner R., 2000. Nitrate uptake and reduction in higher and lower plants. Plant Cell Environ. 23: 1005–1024. [NASA ADS] [CrossRef] [EDP Sciences] [MathSciNet] [PubMed] (In the text)
  • Uedan K. and Sugiyama T., 1976. Purification and characterization of phosphoenolpyruvate carboxylase from maize leaves. Plant Physiol. 57: 906–910. [CrossRef] [PubMed] (In the text)
  • Volk R., Chaillou S., Mariotti A. and Morotgaudry J.F., 1992. Beneficial effects of concurrent ammonium and nitrate nutrition on the growth of Phaseolus vulgaris – a N15 study. Plant Physiol. Biochem. 30: 487–493. (In the text)
  • Wiebe H.H., 1975. Photosynthesis in wood. Physiol Plant 332: 45–46. (In the text)
  • Wittmann C., Aschan G. and Pfanz H., 2001. Leaf and twig photosynthesis of young beech (Fagus sylvatica) and aspen (Populus tremula) trees grown under different light regime. Basic Appl. Biol. 2: 145–154. [CrossRef] (In the text)
  • Wittmann C. and Pfanz H., 2007. Temperature dependency of bark photosynthesis in beech (Fagus sylvatica L.) and birch (Betula pendula Roth.) trees. J. Exp. Bot. 58: 4293–4306. [CrossRef] [PubMed] (In the text)
  • Wittmann C., Pfanz H., Loreto F., Centritto M., Pietrini F. and Alessio G., 2006. Stem CO2 release under illumination: corticular photosynthesis, photorespiration or inhibition of mitochondrial respiration? Plant Cell Environ. 29: 1149–1158. [CrossRef] [PubMed] (In the text)

All Tables

Table I

Characteristics of the soil substrate used when planting beech saplings, before any N fertilization. Measurements were achieved from soil aliquots from five pots for the texture and from soil aliquots from nine pots for each other characteristic. Data are means ±1 SE.

Table II

Mass-based N concentrations (mg g−1 DW) and leaf or stem mass per area (LMA and SMA, g DW m−2) of leaves and current-year stems of Fagus sylvatica L. in each N treatment (T1, T2, and T3). Data are means ±1 SE with n = 3. For each organ (leaves or stems), letters (a, b, and c) indicate significant differences between N treatments.

All Figures

thumbnail Figure 1

Gas exchange of current-year stems of Fagus sylvatica L. measured as CO2 efflux in the dark (dark respiration rate, Rd), CO2 efflux under saturated light (Rl) and calculated rates of gross photosynthesis (Pg), for each N treatment (T1, T2, and T3). Data are means ±1 SE, with n = 3. Letters (a, b, and c) indicate significant differences between N treatments, for each parameter (Rd, Rl and Pg).

In the text
thumbnail Figure 2

Relationship between area-based gross photosynthesis (Pg) and area-based dark respiration rate (Rd) of current-year stems of Fagus sylvatica L. trees subjected to different nitrogen treatments. Each value corresponds to one of the three current-year stems sampled on each of the nine trees (three per N treatment).

In the text
thumbnail Figure 3

Relationship between mass-based gross photosynthesis (Pg) and mass-based nitrogen concentration of current-year stems of Fagus sylvatica L. trees subjected to different nitrogen treatment. Each value corresponds to one of the three current-year stems sampled on each of nine trees (three per treatment).

In the text
thumbnail Figure 4

Relationship between mass-based total soluble protein content and mass-based nitrogen concentration of leaves (black-filled circles) and current-year stems (open circles) of Fagus sylvatica L. Each point corresponds to the mean value of each N treatment and mean value of protein content (n = 3 for each ones). The standard errors of the ordinate and abscissa values are shown as vertical and horizontal bars, respectively.

In the text
thumbnail Figure 5

Relationships between Rubisco total activity and mass-based nitrogen concentration (a) and between PEP carboxylase total activity and mass-based nitrogen concentration (b) of leaves (black-filled circles) and current-year stems (open circles) of Fagus sylvatica L. Each point corresponds to the mean value of a N treatment and mean value of enzyme activity (n = 3 for each ones). The standard errors of the ordinate and abscissa values are shown as vertical and horizontal bars, respectively.

In the text