Sucrose-phosphate synthase responds differently to source-sink relations and to photosynthetic rates: Lolium perenne L. growing at elevated pCO2 in the field

Lolium perenne, a main component species in managed grassland, is well adapted to defoliation, fertilization, and regrowth cycles; and hence, to changes in the assimilatory carbon source‐sink ratio. In the Swiss Free Air CO₂ Enrichment experiment the source‐sink ratio is (i) increased by elevated partial pressure of CO₂ (p_CO2), (ii) decreased by enhanced carbon use under high N fertilization, and (iii) gradually increased during regrowth after defoliation. Since sucrose synthesis plays a central role in leaf carbohydrate metabolism in this fructan‐accumulating species, we investigated how sucrose‐phosphate synthase (SPS) responds to the differing assimilatory carbon fluxes and source‐sink ratios in the field. Assimilatory carbon flux, as estimated by leaf gas exchange, strongly depended on p_CO2. Surprisingly, the SPS content per leaf area did not increase with p_CO2, but increased with N fertilization. During later regrowth, when a dense canopy had formed, the SPS content decreased; in particular, SPS was decreased at high N under elevated p_CO2. Further, the higher assimilatory carbon flux through SPS at elevated p_CO2 was accompanied by a higher activation state of SPS. The SPS content correlated very strongly with the ratio of free sucrose to free amino acid in leaves, which represents the carbon source‐sink ratio. Hence, SPS content in L. perenne appears to be regulated by the current, strongly nitrogen‐dependent, source‐sink relation.     

Partitioning of P and the activity of root acid phosphatase in white clover (Trifolium repens L.) are modified by increased atmospheric CO2 and P fertilisation

The growth response of white clover (Trifolium repens L.) to the expected increase in atmospheric partial pressure of CO₂ (p_CO2) may depend on P availability. A decrease in the rate of transpiration due to increased p_CO2 may reduce the amount of P transported to the shoot, thereby causing a change in the partitioning of P between the root and shoot. To test these hypotheses, four concentrations of P in the nutrient solution, combined with two p_CO2 treatments, were applied to nodulated white clover plants. Compared to ambient p_CO2 (35 Pa), twice ambient p_CO2 (70 Pa) reduced the rate of transpiration but did not impair the total P uptake per plant. However, at twice ambient p_CO2 and a moderate to high supply of P, concentrations of structural P and soluble P (Pi) were lower in the leaves and higher in the roots. The activity of root acid phosphatase was lower at twice ambient p_CO2 than at ambient p_CO2; it depended on the Pi concentration in the roots. At the highest P concentration, twice ambient p_CO2 stimulated photosynthesis and the growth rate of the plant without affecting the concentration of nonstructural carbohydrates in the leaves. However, at the lower P concentrations, plants at twice ambient p_CO2 lost their stimulation of photosynthesis in the afternoon, they accumulated nonstructural carbohydrates in the leaves and their growth rate was not stimulated; indicating C-sink limitation of growth. P nutrition will be crucial to the growth of white clover under the expected future conditions of increased p_CO2. This revised version was published online in June 2006 with corrections to the Cover Date.     

Acclimation of Photosynthesis to Elevated CO2 under Low-Nitrogen Nutrition Is Affected by the Capacity for Assimilate Utilization. Perennial Ryegrass under Free-Air CO2 Enrichment

Acclimation of photosynthesis to elevated CO₂ has previously been shown to be more pronounced when N supply is poor. Is this a direct effect of N or an indirect effect of N by limiting the development of sinks for photoassimilate? This question was tested by growing a perennial ryegrass (Lolium perenne) in the field under elevated (60 Pa) and current (36 Pa) partial pressures of CO₂ (pCO2) at low and high levels of N fertilization. Cutting of this herbage crop at 4- to 8-week intervals removed about 80% of the canopy, therefore decreasing the ratio of photosynthetic area to sinks for photoassimilate. Leaf photosynthesis, in vivo carboxylation capacity, carbohydrate, N, ribulose-1,5-bisphosphate carboxylase/oxygenase, sedoheptulose-1,7-bisphosphatase, and chloroplastic fructose-1,6-bisphosphatase levels were determined for mature lamina during two consecutive summers. Just before the cut, when the canopy was relatively large, growth at elevated pCO₂ and low N resulted in significant decreases in carboxylation capacity and the amount of ribulose-1,5-bisphosphate carboxylase/oxygenase protein. In high N there were no significant decreases in carboxylation capacity or proteins, but chloroplastic fructose-1,6-bisphosphatase protein levels increased significantly. Elevated pCO₂ resulted in a marked and significant increase in leaf carbohydrate content at low N, but had no effect at high N. This acclimation at low N was absent after the harvest, when the canopy size was small. These results suggest that acclimation under low N is caused by limitation of sink development rather than being a direct effect of N supply on photosynthesis.     

Elevated pCO2 Affects C and N Metabolism in Wild Type and Transgenic Tobacco Exhibiting Altered C/N Balance in Metabolite Analysis

Abstract: Diurnal changes in starch, sugar and amino acid concentrations in source leaves, sink leaves and roots of tobacco plants were determined. In addition to wild type tobacco, transformed plants deficient in root nitrate reductase and exhibiting decreased rates of growth were employed. Further, the growth rates of tobacco plants were modulated by exposure to elevated pCO₂. From the diurnal alterations in metabolite concentrations, the daily turnover of starch and amino N was estimated in order to: (i) elucidate whether turnover rates can be related to growth rates, and (ii) identify individual amino compounds with the potential to indicate nitrogen fluxes and the C/N status of plants. Elevated pCO₂ increased growth rates and daily turnover of starch in both wild type and transformed plants, indicating enhanced rates of photosynthesis. In wild type plants, elevated pCO₂ increased the turnover of amino N, notably glutamine and alanine, in mature source leaves, indicating enhanced nitrate reduction. By contrast, amino N turnover in source leaves of transformed plants was not affected by elevated pCO₂, although nitrate reduction was presumably enhanced. Apparently, export of amino N was increased from the source leaves of transformed plants. This assumption was supported by a significantly increased turnover of amino N in young sink leaves compared to mature source leaves, indicating a preference for acropetal amino N allocation and import into the young leaves of the transformed plants. Further, elevated pCO₂ increased the allocation of leaf‐derived amino N to the roots of transformed plants. This led to increased levels of amino compounds during the entire day, notably glutamate, but did not affect root growth of the transformed plants. The suitability of individual amino compounds as markers for major N fluxes, such as nitrate reduction, photorespiration, and amino N export and import is discussed.     

Export of carbon from leaf blades of Poa alpina L. at elevated CO2 and two nutrient regimes

The hypothesis was tested that, in plants of the alpine meadow grass (Poa alpina L.) exposed to elevated CO2, net photosynthesis and export from source leaves is reduced as a result of feedback from sinks. Nutrient supply was used as one way of reducing photosynthesis and export. Single plants were grown in sand culture under specified controlled environmental conditions for a period of 50 d at two levels of nitrogen and phosphorus ('low': 0.2 mol m^-3 N, 0.04 mol m^-3 P; 'high': 2.5 mol m^-3 N, 0.5 mol m^-3 P). Compartmentation within, and export of carbon from, individual youngest fully expanded leaves of acclimated plants was determined using ¹⁴C feeding and efflux plus mass balance calculations of carbohydrate export. Independent of treatment, the bulk of soluble carbohydrate (65-75%) was present as fructan, with most of the remainder being sucrose. Depending on nutrient supply, CO2 could alter export from source leaves either by a reduction in the amount of sucrose present in a readily available pool for transport, or by altering the rate constant describing phloem loading.Key words: Poa alpina L., phloem transport, carbohydrate, compartmentation, export, elevated CO2, nutrients.      

Soil mineral nitrogen availability was unaffected by elevated atmospheric pCO2 in a four year old field experiment (Swiss FACE)

The effect of elevated (60 Pa) atmospheric carbon dioxide partial pressure (p_CO2) and N fertilisation on the availability of mineral N and on N transformation in the soil of a Lolium perenne L. monoculture was investigated in the Swiss FACE (Free Air Carbon dioxide Enrichment) experiment. The apparent availability of nitrate and ammonium for plants was estimated during a representative, vegetative re-growth period at weekly intervals from the sorption of the minerals to mixed-bed ion-exchange resin bags at a soil depth of 5 cm. N mineralisation was measured using sequential coring and in situ exposure of soil cores in the top 10 cm of the soil before and after the first cut in spring 1997. High amounts of mineral N were bound to the ion exchange resin during the first week of re-growth. This was probably the combined result of the fertiliser application, the weak demand for N by the newly cut sward and presumably high rates of root decay and exudation after cutting the sward. During the first 2 weeks after the application of fertiliser N at the first cut, there was a dramatic reduction in available N; N remained low during the subsequent weeks of re-growth in all treatments. Overall, nitrate was the predominant form of mineral N that bound to the resin for the duration of the experiment. Apparently, there was always more nitrate than ammonium available to the plants in the high N fertilisation treatment for the whole re-growth period. Apparent N availability was affected significantly by elevated p_CO2 only in the first week after the cut; under high N fertilisation, elevated p_CO2 increased the amount of mineral N that was apparently available to the plants. Elevated p_CO2 did not affect apparent net transformation of N, loss of N or uptake of N by plants. The present data are consistent with earlier results and suggest that the amount of N available to plants from soil resources does not generally increase under elevated atmospheric p_CO2. Thus, a possible limiting effect of N on primary production could become more stringent under elevated atmospheric p_CO2 as the demand of the plant for N increases. This revised version was published online in June 2006 with corrections to the Cover Date.     

Nitrate supply and plant development influence nitrogen uptake and allocation under elevated CO<Subscript>2</Subscript> in durum wheat grown hydroponically

Growth in elevated CO₂ often leads to decreased plant nitrogen contents and down-regulation of photosynthetic capacity. Here, we investigated whether elevated CO₂ limits nitrogen uptake when nutrient movement to roots is unrestricted, and the dependence of this limitation on nitrogen supply and plant development in durum wheat (Triticum durum Desf.). Plants were grown hydroponically at two N supplies and ambient and elevated CO₂ concentrations. Elevated CO₂ decreased nitrate uptake per unit root mass with low N supply at early grain filling, but not at anthesis. This decrease was not associated with higher nitrate or amino acid, or lower non-structural carbohydrate contents in roots. At anthesis, elevated CO₂ decreased the nitrogen content of roots with both levels of N and that of aboveground organs with high N. With low N, elevated CO₂ increased N allocation to aboveground plant organs and nitrogen concentration per unit flag leaf area at anthesis, and per unit aboveground dry mass at both growth stages. The results from the hydroponic experiment suggest that elevated CO₂ restricts nitrate uptake late in development, high N supply overriding this restriction. Increased nitrogen allocation to young leaves at low N supply could alleviate photosynthetic acclimation to elevated CO₂.     

Photosynthetic acclimation to elevated CO2 in Phaseolus vulgaris L. is altered by growth response to nitrogen supply

Abstract Long‐term exposure of plants to elevated CO₂ often leads to downward photosynthetic acclimation. Nitrogen (N) deficiency could potentially exacerbate this response by reducing growth rate and the sink for photosynthates, but this has not always been observed. Experimentally, the interpretation of N effects on CO₂ responses can be confounded by increasing severity of tissue N deficiency over time when N supply is not adjusted as demand increases. In this study, N supply ranged from sub‐ to supra‐optimal (20–540 kgN ha^–l equivalent), and relatively stable levels of tissue N concentration were obtained in all treatments by varying twice‐weekly application rates in proportion to plant growth. The effects of N on photosynthesis and growth of beans (Phaseolus vulgaris L.) raised at ambient (35 Pa) and three elevated (70, 105, 140 Pa) CO₂ partial pressures (pCO₂) were evaluated. Averaging across N treatments, leaf total non‐structural carbohydrates (TNC) were 2.5‐ to 3‐fold higher and leaf N concentrations were 31–35% lower at elevated compared to ambient pCO₂. Light‐saturated net CO₂ assimilation rates measured at growth pCO₂ (A_satg) were significantly higher (26–40% depending on N supply) in plants grown at elevated compared to ambient pCO₂. When measured at a common pCO₂ of 35 Pa, the A_sat of plants grown at elevated CO₂ was 15–29% less than that of plants grown at 35 Pa, indicative of downward photosynthetic acclimation. The magnitude of downward photosynthetic acclimation to elevated CO₂ was greater in plants grown at high (180 and 540 kgN ha^–l) compared to low (20 and 60 kgN ha^–l) N supply, and this was associated with a higher A_sat at growth pCO₂, higher leaf area ratio (leaf area/total biomass), and higher TNC in leaves of high‐N plants. Our results indicate that the effect of N on acclimation to CO₂ will depend on the balance between supply and demand for N during the growing period, and the effect this has on biomass allocation and source‐sink C balance at the whole‐plant level.     

Nutrient and genotypic effects on CO2-responsiveness: photosynthetic regulation in Leucadendron species of a nutrient-poor environment

Four South African Leucadendron congenerics with divergent soil N and P preferences were grown as juveniles at contrasting nutrient concentrations at ambient (350 mol mol^-1) and elevated (700 mol mol^-1) atmospheric CO2 levels. Photosynthetic parameters were related to leaf nutrient and carbohydrate status to reveal controls of carbon uptake rate. In all species, elevated CO2 depressed both the maximum Rubisco catalytic activity (Vc,max, by 19-44%) and maximum electron transport rate (Jmax, by 13-39%), indicating significant photosynthetic acclimation of both measures. Even so, all species had increased maximum light-saturated rate of net CO2 uptake (Amax)) at the elevated growth CO2 level, due to higher intercellular CO2 concentration (ci). Leaf nitrogen concentration was central to photosynthetic performance, correlating with Amax, Vc,max and Jmax, Vc,max and Jmax were linearly co-correlated, revealing a relatively invariable Jmax:Vc,max ratio, probably due to N resource optimization between light harvesting (RuBP regeneration) and carboxylation. Leaf total non-structural carbohydrate concentration (primarily starch) increased in high CO2, and was correlated with the reduction in Vc,max and Jmax. Apparent feedback control of Vc,max and Jmax was thus surprisingly consistent across all species, and may regulate carbon exchange in response to end-product fluctuation. If so, elevated CO2 may have emulated an excess end-product condition, triggering both Vc,max and Jmax down-regulation. In Leucadendron, a general physiological mechanism seems to control excess carbohydrate formation, and photosynthetic responsiveness to elevated CO2, independently of genotype and nutrient concentration. This mechanism may underlie photosynthetic acclimation to source:sink imbalances resulting from such diverse conditions as elevated CO2, low sink strength, low carbohydrate export, and nutrient limitation.     

Soil 13C–15N dynamics in an N2-fixing clover system under long-term exposure to elevated atmospheric CO2

Reduced soil N availability under elevated CO₂ may limit the plant's capacity to increase photosynthesis and thus the potential for increased soil C input. Plant productivity and soil C input should be less constrained by available soil N in an N₂‐fixing system. We studied the effects of Trifolium repens (an N₂‐fixing legume) and Lolium perenne on soil N and C sequestration in response to 9 years of elevated CO₂ under FACE conditions. ¹⁵N‐labeled fertilizer was applied at a rate of 140 and 560 kg N ha^?1 yr^?1 and the CO₂ concentration was increased to 60 Pa pCO₂ using ¹³C‐depleted CO₂. The total soil C content was unaffected by elevated CO₂, species and rate of ¹⁵N fertilization. However, under elevated CO₂, the total amount of newly sequestered soil C was significantly higher under T. repens than under L. perenne. The fraction of fertilizer‐N (f_N) of the total soil N pool was significantly lower under T. repens than under L. perenne. The rate of N fertilization, but not elevated CO₂, had a significant effect on f_N values of the total soil N pool. The fractions of newly sequestered C (f_C) differed strongly among intra‐aggregate soil organic matter fractions, but were unaffected by plant species and the rate of N fertilization. Under elevated CO₂, the ratio of fertilizer‐N per unit of new C decreased under T. repens compared with L. perenne. The L. perenne system sequestered more ¹⁵N fertilizer than T. repens: 179 vs. 101 kg N ha^?1 for the low rate of N fertilization and 393 vs. 319 kg N ha^?1 for the high N‐fertilization rate. As the loss of fertilizer‐¹⁵N contributed to the ¹⁵N‐isotope dilution under T. repens, the input of fixed N into the soil could not be estimated. Although N₂ fixation was an important source of N in the T. repens system, there was no significant increase in total soil C compared with a non‐N₂‐fixing L. perenne system. This suggests that N₂ fixation and the availability of N are not the main factors controlling soil C sequestration in a T. repens system.     

Does nitrogen nutrition restrict the CO2 response of fertile grassland lacking legumes?

The extent of the response of plant growth to atmospheric CO₂ enrichment depends on the availability of resources other than CO₂. An important growth-limiting resource under field conditions is nitrogen (N). N may, therefore, influence the CO₂ response of plants. The effect of elevated CO₂ (60 Pa) partial pressure (pCO₂) on the N nutrition of field-grown Lolium perenne swards, cultivated alone or in association with Trifolium repens, was investigated using free air carbon dioxide enrichment (FACE) technology over 3 years. The established grassland ecosystems were treated with two N fertilization levels and were defoliated at two frequencies. Under elevated pCO₂, the above-ground plant material of the L. perenne monoculture showed a consistent and significant decline in N concentration which, in general, led to a lower total annual N yield. Despite the decline in the critical N concentration (minimum N concentration required for non-N-limited biomass production) under elevated pCO₂, the index of N nutrition (ratio of actual N concentration and critical N concentration) was lower under elevated pCO₂ than under ambient pCO₂ in frequently defoliated L. perenne monocultures. Thus, we suggest that reduced N yield under elevated pCO₂ was evoked indirectly by a reduction of plant-available N. For L. perenne grown in association with T. repens and exposed to elevated pCO₂, there was an increase in the contribution of symbiotically fixed N to the total N yield of the grass. This can be explained by an increased apparent transfer of N from the associated N₂-fixing legume species to the non-fixing grass. The total annual N yield of the mixed grass/legume swards increased under elevated pCO₂. All the additional N yielded was due to symbiotically fixed N. Through the presence of an N₂-fixing plant species more symbiotically fixed N was introduced into the system and consequently helped to overcome N limitation under elevated pCO₂. Received: 11 November 1996 / Accepted: 20 May 1997     

The impact of long-term elevated CO2 on C and N retention in stable SOM pools

Elevated atmospheric CO₂ frequently increases plant production and concomitant soil C inputs, which may cause additional soil C sequestration. However, whether the increase in plant production and additional soil C sequestration under elevated CO₂ can be sustained in the long-term is unclear. One approach to study C–N interactions under elevated CO₂ is provided by a theoretical framework that centers on the concept of progressive nitrogen limitation (PNL). The PNL concept hinges on the idea that N becomes less available with time under elevated CO₂. One possible mechanism underlying this reduction in N availability is that N is retained in long-lived soil organic matter (SOM), thereby limiting plant production and the potential for soil C sequestration. The long-term nature of the PNL concept necessitates the testing of mechanisms in field experiments exposed to elevated CO₂ over long periods of time. The impact of elevated CO₂ and ¹⁵N fertilization on L. perenne and T. repens monocultures has been studied in the Swiss FACE experiment for ten consecutive years. We applied a biological fractionation technique using long-term incubations with repetitive leaching to determine how elevated CO₂ affects the accumulation of N and C into more stable SOM pools. Elevated CO₂ significantly stimulated retention of fertilizer-N in the stable pools of the soils covered with L. perenne receiving low and high N fertilization rates by 18 and 22%, respectively, and by 45% in the soils covered by T. repens receiving the low N fertilization rate. However, elevated CO₂ did not significantly increase stable soil C formation. The increase in N retention under elevated CO₂ provides direct evidence that elevated CO₂ increases stable N formation as proposed by the PNL concept. In the Swiss FACE experiment, however, plant production increased under elevated CO₂, indicating that the additional N supply through fertilization prohibited PNL for plant production at this site. Therefore, it remains unresolved why elevated CO₂ did not increase labile and stable C accumulation in these systems.     

Carbon source–sink limitations differ between two species with contrasting growth strategies

Understanding how carbon source and sink strengths limit plant growth is a critical knowledge gap that hinders efforts to maximize crop yield. We investigated how differences in growth rate arise from source–sink limitations, using a model system comparing a fast‐growing domesticated annual barley (Hordeum vulgare cv. NFC Tipple) with a slow‐growing wild perennial relative (Hordeum bulbosum). Source strength was manipulated by growing plants at sub‐ambient and elevated CO₂ concentrations ([CO₂]). Limitations on vegetative growth imposed by source and sink were diagnosed by measuring relative growth rate, developmental plasticity, photosynthesis and major carbon and nitrogen metabolite pools. Growth was sink limited in the annual but source limited in the perennial. RGR and carbon acquisition were higher in the annual, but photosynthesis responded weakly to elevated [CO₂] indicating that source strength was near maximal at current [CO₂]. In contrast, photosynthetic rate and sink development responded strongly to elevated [CO₂] in the perennial, indicating significant source limitation. Sink limitation was avoided in the perennial by high sink plasticity: a marked increase in tillering and root:shoot ratio at elevated [CO₂], and lower non‐structural carbohydrate accumulation. Alleviating sink limitation during vegetative development could be important for maximizing growth of elite cereals under future elevated [CO₂].     

Comparison of photosynthetic acclimation to elevated CO2 and limited nitrogen supply in soybean

Plants grown at elevated CO₂ often acclimate such that their photosynthetic capacities are reduced relative to ambient CO₂-grown plants. Reductions in synthesis of photosynthetic enzymes could result either from reduced photosynthetic gene expression or from reduced availability of nitrogen-containing substrates for enzyme synthesis. Increased carbohydrate concentrations resulting from increased photosynthetic carbon fixation at elevated CO₂ concentrations have been suggested to reduce the expression of photosynthetic genes. However, recent studies have also suggested that nitrogen uptake may be depressed by elevated CO₂, or at least that it is not increased enough to keep pace with increased carbohydrate production. This response could induce a nitrogen limitation in elevated-CO₂ plants that might account for the reduction in photosynthetic enzyme synthesis. If CO₂ acclimation were a response to limited nitrogen uptake, the effects of elevated CO₂ and limiting nitrogen supply on photosynthesis and nitrogen allocation should be similar. To test this hypothesis we grew non-nodulating soybeans at two levels each of nitrogen and CO₂ concentration and measured leaf nitrogen contents, photosynthetic capacities and Rubisco contents. Both low nitrogen and elevated CO₂ reduced nitrogen as a percentage of total leaf dry mass but only low nitrogen supply produced significant decreases in nitrogen as a percentage of leaf structural dry mass. The primary effect of elevated CO₂ was to increase non-structural carbohydrate storage rather than to decrease nitrogen content. Both low nitrogen supply and elevated CO₂ also decreased carboxylation capacity (V_cmax) and Rubisco content per unit leaf area. However, when V_cmax and Rubisco content were expressed per unit nitrogen, low nitrogen supply generally caused them to increase whereas elevated CO₂ generally caused them to decrease. Finally, elevated CO₂ significantly increased the ratio of RuBP regeneration capacity to V_cmax whereas neither nitrogen supply nor plant age had a significant effect on this parameter. We conclude that reductions in photosynthetic enzyme synthesis in elevated CO₂ appear not to result from limited nitrogen supply but instead may result from feedback inhibition by increased carbohydrate contents.     

Growth response of Trifolium repens L. and Lolium perenne L. as monocultures and bi-species mixture to free air CO2 enrichment and management

Trifolium repens L. and Lolium perenne L. were grown in monocultures and bi-species mixture in a Free Air Carbon Dioxide Enrichment (FACE) experiment at elevated (60 Pa) and ambient (35 Pa) CO₂ partial pressure (pCO₂) for three years. The effects of defoliation frequencies (4 and 7 cuts in 1993; 4 and 8 cuts in 1994/95) and nitrogen fertilization (10 and 42 g m^–2 y^–1 N in 1993; 14 and 56 g m^–2 y^–1 N in 1994/95) on the growth response to pCO₂ were investigated. There were significant interspecific differences in the CO₂ responses during the first two years, while in the third growing season, these interspecific differences disappeared. Yield of T. repens in monocultures increased in the first two years by 20% when grown at elevated pCO₂. This CO₂ response was independent of defoliation frequency and nitrogen fertilization. In the third year, the CO₂ response of T. repens declined to 11%. In contrast, yield of L. perenne monocultures increased by only 7% on average over three years at elevated pCO₂. The yield response of L. perenne to CO₂ changed according to defoliation frequency and nitrogen fertilization, mainly in the second and third year. The ratio of root/yield of L. perenne increased under elevated pCO₂, low N fertilizer rate, and frequent defoliation, but it remained unchanged in T. repens. We suggest that the more abundant root growth of L. perenne was related to increased N limitation under elevated pCO₂. The consequence of these interspecific differences in the CO₂ response was a higher proportion of T. repens in the mixed swards at elevated pCO₂. This was evident in all combinations of defoliation and nitrogen treatments. However, the proportion of the species was more strongly affected by N fertilization and defoliation frequency than by elevated pCO₂. Based on these results, we conclude that the species proportion in managed grassland may change as the CO₂ concentration increases. However, an adapted management could, at least partially, counteract such CO₂ induced changes in the proportion of the species. Since the availability of mineral N in the soil may be important for the species’ responses to elevated pCO₂, more long-term studies, particularly of processes in the soil, are required to predict the entire ecosystem response.     

Does deep soil N availability sustain long‐term ecosystem responses to elevated CO2?

A scrub‐oak woodland has maintained higher aboveground biomass accumulation after 11 years of atmospheric CO₂ enrichment (ambient +350 μmol CO₂ mol^?1), despite the expectation of strong nitrogen (N) limitation at the site. We hypothesized that changes in plant available N and exploitation of deep sources of inorganic N in soils have sustained greater growth at elevated CO₂. We employed a suite of assays performed in the sixth and 11th year of a CO₂ enrichment experiment designed to assess soil N dynamics and N availability in the entire soil profile. In the 11th year, we found no differences in gross N flux, but significantly greater microbial respiration (P≤0.01) at elevated CO₂. Elevated CO₂ lowered extractable inorganic N concentrations (P=0.096) considering the whole soil profile (0–190 cm). Conversely, potential net N mineralization, although not significant in considering the entire profile (P=0.460), tended to be greater at elevated CO₂. Ion‐exchange resins placed in the soil profile for approximately 1 year revealed that potential N availability at the water table was almost 3 × greater than found elsewhere in the profile, and we found direct evidence using a ¹⁵N tracer study that plants took up N from the water table. Increased microbial respiration and shorter mean residence times of inorganic N at shallower depths suggests that enhanced SOM decomposition may promote a sustained supply of inorganic N at elevated CO₂. Deep soil N availability at the water table is considerable, and provides a readily available source of N for plant uptake. Increased plant growth at elevated CO₂ in this ecosystem may be sustained through greater inorganic N supply from shallow soils and N uptake from deep soil.     

The effects of elevated CO2 concentrations on the root growth of Lolium perenne and Trifolium repens grown in a FACE* system

Lolium perenne and Trifolium repens were grown in a Free Air CO₂ Enrichment (FACE) system at elevated (600 μimol mol-¹) and ambient (340 μmol mol-¹) carbon dioxide concentrations during a whole growing season. Using a root ingrowth bag technique the extent to which CO₂ enrichment influenced the growth of L, perenne and T. repens roots under two contrasting nutrient regimes was examined. Root ingrowth bags were inserted for a fixed time into the soil in order to trap roots. It was also possible to follow the mortality of roots in bags inserted for different time intervals. Root ingrowth of both L. perenne and T. repens increased under elevated CO₂ conditions. In L. perenne, root ingrowth decreased with increasing nutrient fertilizer level, but for T. repens the root ingrowth was not affected by the nutrient application rate. Besides biomass measurements, root length estimates were made for T, repens. These showed an increase under elevated CO₂ concentrations. Root decomposition appeared to decrease under elevated CO₂ concentrations. A possible explanation for this effect is the observed changes in tissue composition, such as the increase in the carbon: nitrogen ratio in roots of L. perenne at elevated CO₂ concentrations.     

Responses of net ecosystem CO2 exchange in managed grassland to long-term CO2 enrichment, N fertilization and plant species

The effects of elevated pCO₂ on net ecosystem CO₂ exchange were investigated in managed Lolium perenne (perennial ryegrass) and Trifolium repens (white clover) monocultures that had been exposed continuously to elevated pCO₂ (60 Pa) for nine growing seasons using Free Air CO₂ Enrichment (FACE) technology. Two levels of nitrogen (N) fertilization were applied. Midday net ecosystem CO₂ exchange (mNEE) and night-time ecosystem respiration (NER) were measured in three growing seasons using an open-flow chamber system. The annual net ecosystem carbon (C) input resulting from the net CO₂ fluxes was estimated for one growing season. In both monocultures and at both levels of N supply, elevated pCO₂ stimulated mNEE by up to 32%, the exact amount depending on intercepted PAR. The response of mNEE to elevated pCO₂ was larger than that of harvestable biomass. Elevated pCO₂ increased NER by up to 39% in both species at both levels of N supply. NER, which was affected by mNEE of the preceding day, was higher in T. repens than in L. perenne. High N increased NER compared to low N supply. According to treatment, the annual net ecosystem C input ranged between 210 and 631 g C m⁻² year⁻¹ and was not significantly affected by the level of pCO₂. Low N supply led to a higher net C input than high N supply. We demonstrated that at the ecosystem level, there was a long-term stimulation in the net C assimilation during daytime by elevated pCO₂. However, because NER was also stimulated, net ecosystem C input was not significantly increased at elevated pCO₂. The annual net ecosystem C input was primarily affected by the amount of N supplied.     

Ten years of free-air CO2 enrichment altered the mobilization of N from soil in Lolium perenne L. swards

Effects of free‐air carbon dioxide enrichment (FACE, 60 Pa pCO₂) on plant growth as compared with ambient pCO₂ (36 Pa) were studied in swards of Lolium perenne L. (perennial ryegrass) at two levels of N fertilization (14 and 56 g m^?2 a^?1) from 1993 to 2002. The objectives were to determine how plant growth responded to the availability of C and N in the long term and how the supply of N to the plant from the two sources of N in the soil, soil organic matter (SOM) and mineral fertilizer, varied over time. In three field experiments, ¹⁵N‐labelled fertilizer was used to distinguish the sources of available N. In 1993, harvestable biomass under elevated pCO₂ was 7% higher than under ambient pCO₂. This relative pCO₂ response increased to 32% in 2002 at high N, but remained low at low N. Between 1993 and 2002, the proportions and amounts of N in harvestable biomass derived from SOM (excluding remobilized fertilizer) were, at high N, increasingly higher at elevated pCO₂ than at ambient pCO₂. Two factorial experiments confirmed that at high N, but not at low N, a higher proportion of N in harvestable biomass was derived from soil (including remobilized fertilizer) following 7 and 9 years of elevated pCO₂, when compared with ambient pCO₂. It is suggested that N availability in the soil initially limited the pCO₂ response of harvestable biomass. At high N, the limitation of plant growth decreased over time as a result of the stimulated mobilization of N from soil, especially from SOM. Consequently, harvestable biomass increasingly responded to elevated pCO₂. The underlying mechanisms which contributed to the increased mobilization of N from SOM under elevated pCO₂ are discussed. This study demonstrated that there are feedback mechanisms in the soil which are only revealed during long‐term field experiments. Such investigations are thus, a prerequisite for understanding the responses of ecosystems to elevated pCO₂ and N supply.