The role of temperature in determining the stimulation of CO2 assimilation at elevated carbon dioxide concentration in soybean seedlings

Soybean ( Glycine max cv. Clark) was grown at both ambient (ca 350 μmol mol⁻¹) and elevated (ca 700 μmol mol⁻¹) CO₂ concentration at 5 growth temperatures (constant day/night temperatures of 20, 25, 30, 35 and 40°C) for 17–22 days after sowing to determine the interaction between temperature and CO₂ concentration on photosynthesis (measured as A, the rate of CO₂ assimilation per unit leaf area) at both the single leaf and whole plant level. Single leaves of soybean demonstrated increasingly greater stimulation of A at elevated CO₂ as temperature increased from 25 to 35°C (i.e. optimal growth rates). At 40°C, primary leaves failed to develop and plants eventually died. In contrast, for both whole plant A and total biomass production, increasing temperature resulted in less stimulation by elevated CO₂ concentration. For whole plants, increased CO₂ stimulated leaf area more as growth temperature increased. Differences between the response of A to elevated CO₂ for single leaves and whole plants may be related to increased self-shading experienced by whole plants at elevated CO₂ as temperature increased. Results from the present study suggest that self-shading could limit the response of CO₂ assimilation rate and the growth response of soybean plants if temperature and CO₂ increase concurrently, and illustrate that light may be an important consideration in predicting the relative stimulation of photosynthesis by elevated CO₂ at the whole plant level.     

Canopy photosynthesis of crops and native plant communities exposed to long-term elevated CO2

Abstract. There have been seven studies of canopy photosynthesis of plants grown in elevated atmospheric CO₂: three of seed crops, two of forage crops and two of native plant ecosystems. Growth in elevated CO₂ increased canopy photosynthesis in all cases. The relative effect of CO₂ was correlated with increasing temperature: the least stimulation occurred in tundra vegetation grown at an average temperature near 10°C and the greatest in rice grown at 43°C. In soybean, effects of CO₂ were greater during leaf expansion and pod fill than at other stages of crop maturation. In the longest running experiment with elevated CO₂ treatment to date, monospecific stands of a C₃ sedge, Scirpus olneyi (Grey), and a C₄ grass, Spartina patens (Ait.) Muhl., have been exposed to twice normal ambient CO₂ concentrations for four growing seasons, in open top chambers on a Chesapeake Bay salt marsh. Net ecosystem CO₂ exchange per unit green biomass (NCE_b) increased by an average of 48% throughout the growing season of 1988, the second year of treatment. Elevated CO₂ increased net ecosystem carbon assimilation by 88% in the Scirpus olneyi community and 40% in the Spartina patens community.     

Elevated CO2 and ozone reduce nitrogen acquisition by Pinus halepensis from its mycorrhizal symbiont

The effects of 700 μmol mol⁻¹ CO₂ and 200 nmol mol⁻¹ ozone on photosynthesis in Pinus halepensis seedlings and on N translocation from its mycorrhizal symbiont, Paxillus involutus, were studied under nutrient-poor conditions. After 79 days of exposure, ozone reduced and elevated CO₂ increased net assimilation rate. However, the effect was dependent on daily accumulated exposure. No statistically significant differences in total plant mass accumulation were observed, although ozone-treated plants tended to be smaller. Changes in atmospheric gas concentrations induced changes in allocation of resources: under elevated ozone, shoots showed high priority over roots and had significantly elevated N concentrations. As a result of different shoot N concentration and net carbon assimilation rates, photosynthetic N use efficiency was significantly increased under elevated CO₂ and decreased under ozone. The differences in photosynthesis were mirrored in the growth of the fungus in symbiosis with the pine seedlings. However, exposure to CO₂ and ozone both reduced the symbiosis-mediated N uptake. The results suggest an increased carbon cost of symbiosis-mediated N uptake under elevated CO₂, while under ozone, plant N acquisition is preferentially shifted towards increased root uptake.     

Growth and photosynthetic response of three soybean cultivars to simultaneous increases in growth temperature and CO2

Three soybean ( Glycine max L. Merr.) cultivars (Maple Glen, Clark and CNS) were exposed to three CO₂ concentrations (370, 555 and 740 μmol mol⁻¹) and three growth temperatures (20/15°, 25/20° and 31/26°C, day/night) to determine intraspecific differences in single leaf/whole plant photosynthesis, growth and partitioning, phenology and final biomass. Based on known carboxylation kinetics, a synergistic effect between temperature and CO₂ on growth and photosynthesis was predicted since elevated CO₂ increases photosynthesis by reducing photorespiration and photorespiration increases with temperature. Increasing CO₂ concentrations resulted in a stimulation of single leaf photosynthesis for 40–60 days after emergence (DAE) at 20/15°C in all cultivars and for Maple Glen and CNS at all temperatures. For Clark, however, the onset of flowering at warmer temperatures coincided with the loss of stimulation in single leaf photosynthesis at elevated CO₂ concentrations. Despite the season-long stimulation of single leaf photosynthesis, elevated CO₂ concentrations did not increase whole plant photosynthesis except at the highest growth temperature in Maple Glen and CNS, and there was no synergistic effect on final biomass. Instead, the stimulatory effect of CO₂ on growth was delayed by higher temperatures. Data from this experiment suggest that: (1) intraspecific variation could be used to select for optimum soybean cultivars with future climate change; and (2) the relationship between temperature and CO₂ concentration may be expressed differently at the leaf and whole plant levels and may not solely reflect known changes in carboxylation kinetics.     

Increasing growth temperature reduces the stimulatory effect of elevated CO2 on photosynthesis or biomass in two perennial species

We examined how anticipated changes in CO₂ concentration and temperature interacted to alter plant growth, harvest characteristics and photosynthesis in two cold-adapted herbaceous perennials, alfalfa ( Medicago sativa L. cv. Arc) and orchard grass ( Dactylis glomerata L. cv. Potomac). Plants were grown at two CO₂ concentrations (362 [ambient] and 717 [elevated] μmol mol⁻¹ CO₂) and four constant day/night temperatures of 15, 20, 25 and 30°C in controlled environmental chambers. Elevated CO₂ significantly increased total plant biomass and protein over a wide range of temperatures in both species. Stimulation of photosynthetic rate, however, was eliminated at the highest growth temperature in M. sativa and relative stimulation of plant biomass and protein at high CO₂ declined as temperature increased in both species. Lack of a synergistic effect between temperature and CO₂ was unexpected since elevated CO₂ reduces the amount of carbon lost via photorespiration and photorespiration increases with temperature. Differences between anticipated stimulatory effects of CO₂ and temperature and whole plant single and leaf measurements are discussed. Data from this study suggest that stimulatory effects of atmospheric CO₂ on growth and photosynthesis may decline with anticipated increases in global temperature, limiting the degree of carbon storage in these two perennial species.     

How does the C4 grass Eragrostis pilosa respond to elevated carbon dioxide and infection with the parasitic angiosperm Striga hermonthica?

Eragrostis pilosa (Linn.) P Beauv., a C₄ grass native to east Africa, was grown at both ambient (350 μmol mol⁻¹ and elevated (700 μmol mol⁻¹) CO₂ in either the presence or absence of the obligate, root hemi-parasite Striga hermonthica (Del.) Benth. Biomass of infected grasses was only 50% that of uninfected grasses at both CO₂ concentrations, with stems and reproductive tissues of infected plants being most severely affected. By contrast, CO₂ concentration had no effect on growth of E. pilosa , although rates of photosynthesis were enhanced by 30–40% at elevated CO₂. Infection with S. hermonthica did not affect either rates of photosynthesis or leaf areas of E. pilosa , but did bring about an increase in root:shoot ratio, leaf nitrogen and phosphorus concentration and a decline in leaf starch concentration at both ambient and elevated CO₂. Striga hermonthica had higher rates of photosynthesis and shoot concentrations of soluble sugars at elevated CO₂, but there was no difference in biomass relative to ambient grown plants. Both infection and growth at elevated CO₂ resulted in an increase in the Δ¹³C value of leaf tissue of E. pilosa , with the CO₂ effect being greater. The proportion of host-derived carbon in parasite tissue, as determined from δ¹³C values, was 27% and 39% in ambient and elevated CO₂ grown plants, respectively. In conclusion, infection with S. hermonthica limited growth of E. pilosa , and this limitation was not removed or alleviated by growing the association at elevated CO₂.     

Elevated atmospheric CO2 influences the interaction between the parasitic angiosperm Orobanche minor and its host Trifolium repens

The influence of the root holoparasitic angiosperm Orobanche minor Sm. on the biomass, photosynthesis, carbohydrate and nitrogen content of Trifolium repens L. was determined for plants grown at two CO₂ concentrations (350 and 550 μmol mol⁻¹). Infected plants accumulated less biomass than their uninfected counterparts, although early in the association there was a transient stimulation of growth. Infection also influenced biomass allocation both between tissues (infected plants had lower root:shoot ratios) and within tissues:infected roots were considerably thicker before the point of parasite attachment and thinner below. Higher concentrations of starch were also found in roots above the point of attachment, particularly for plants grown in elevated CO₂. Elevated CO₂ stimulated the growth of T. repens only during the early stages of development. There was a significant interaction between infection and CO₂ on growth, with infected plants showing a greater response, such that elevated CO₂ partly alleviated the effects of the parasite on host growth. Elevated CO₂ did not affect total O. minor biomass per host, the number of individual parasites supported by each host, or their time of attachment to the host root system. Photosynthesis was stimulated by elevated CO₂ but was unaffected by O. minor . There was no evidence of down-regulation of photosynthesis in T. repens grown at elevated CO₂ in either infected or uninfected plants. The data are discussed with regard to the influence of elevated CO₂ on other parasitic angiosperm-host associations and factors which control plant responses to elevated CO₂.     

Canopy development of a model herbaceous community exposed to elevated atmospheric CO2 and soil nutrients

To test the prediction that elevated CO₂ increases the maximum leaf area index (LAI) through a stimulation of photosynthesis, we exposed model herbaceous communities to two levels of CO₂ crossed with two levels of soil fertility. Elevated CO₂ stimulated the initial rate of canopy development and increased cumulative LAI integrated over the growth period, but it had no effect on the maximum LAI. In contrast to CO₂, increased soil nutrient availability caused a substantial increase in maximum LAI. Elevated CO₂ caused a slight increase in leaf area and nitrogen allocated to upper canopy layers and may have stimulated leaf turnover deep in the canopy. Gas exchange measurements of intact communities made near the time of maximum LAI indicated that soil nutrient availability, but not CO₂ enrichment, caused a substantial stimulation of net ecosystem carbon exchange. These data do not support our prediction of a higher maximum LAI by elevated CO₂ because the initial stimulation of LAI diminished by the end of the growth period. However, early in development, leaf area and carbon assimilation of communities may have been greatly enhanced. These results suggest that the rate of canopy development in annual communities may be accelerated with future increases in atmospheric CO₂ but that maximum LAI is set by soil fertility.     

Mycorrhiza formation and elevated CO2 both increase the capacity for sucrose synthesis in source leaves of spruce and aspen

The effects of mycorrhiza formation in combination with elevated CO₂ concentrations on carbon metabolism of Norway spruce ( Picea abies ) seedlings and aspen ( Populus tremula × Populus tremuloides ) plantlets were analysed. Plants were inoculated for 6 wk with the ectomycorrhizal fungi Amanita muscaria and Paxillus involutus (aspen only) in an axenic Petri-dish culture at 350 and 700 μl l⁻¹ CO₂ partial pressure. After mycorrhiza formation, a stimulation of net assimilation rate was accompanied by decreased activities of sucrose synthase, an increased activation state of sucrose-phosphate synthase, decreased fructose-2,6-bisphosphate and starch, and slightly elevated glucose-6-phosphate contents in source leaves of both host species, independent of CO₂ concentration. Exposure to elevated CO₂ generally resulted in higher net assimilation rates, increased starch as well as decreased fructose-2,6-bisphosphate (aspen only) content in source leaves of both mycorrhizal and nonmycorrhizal plants. Our data indicate only slightly improved carbon utilization by mycorrhizal plants at elevated CO₂. They demonstrate however, that both factors which modulate the sink-source properties of plants increase the capacity for sucrose synthesis in source leaves mainly by allosteric enzyme regulation.     

Effect of enhanced atmospheric CO2 on mycorrhizal colonization by Glomus mosseae in Plantago lanceolata and Trifolium repens

Plantago lanceolata L. and Trifolium repens L. were grown for 16 wk in ambient (360 μmol mol⁻¹) and elevated (610 μmol mol⁻¹) atmospheric CO₂. Plants were inoculated with the arbuscular mycorrhizal (AM) fungus Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe and given a phosphorus supply in the form of bonemeal, which would not be immediately available to the plants. Seven sequential harvests were taken to determine whether the effect of elevated CO₂ on mycorrhizal colonization was independent of the effect of CO₂ on plant growth. Plant growth analysis showed that both species grew faster in elevated CO₂ and that P. lanceolata had increased carbon allocation towards the roots. Elevated CO₂ did not affect the percentage of root length colonized (RLC); although total colonized root length was greater, when plant size was taken into account this effect disappeared. This finding was also true for root length colonized by arbuscules. No CO₂ effect was found on hyphal density (colonization intensity) in roots. The P content of plants was increased at elevated CO₂, although both shoot and root tissue P concentration were unchanged. This was again as a result of bigger plants at elevated CO₂. Phosphorus inflow was unaffected by CO₂ concentrations. It is concluded that there is no direct permanent effect of elevated CO₂ on mycorrhizal functioning, as internal mycorrhizal development and the mycorrhizal P uptake mechanism are unaffected. The importance of sequential harvests in experiments is discussed. The direction for future research is highlighted, especially in relation to C storage in the soil.     

Effects of manganese toxicity on photosynthesis of white birch (Betula platyphylla var. japonica) seedlings

The effects of manganese (Mn) toxicity on photosynthesis in white birch ( Betula platyphylla var. japonica ) leaves were examined by the measurement of gas exchange and chlorophyll fluorescence in hydroponically cultured plants. The net photosynthetic rate at saturating light and ambient CO₂ (C_a) of 35 Pa decreased with increasing leaf Mn concentrations. The carboxylation efficiency, derived from the difference in CO₂ assimilation rate at intercellular CO₂ pressures attained at C_a of 13 Pa and O Pa, decreased with greater leaf Mn accumulation. Net photosynthetic rate at saturating light and saturating CO₂ (5%) also declined with leaf Mn accumulation while the maximum quantum yield of O₂ evolution at saturating CO₂ was not affected. The maximum efficiency of PSII photochemistry (F_v/F_m) was little affected by Mn accumulation in white birch leaves over a wide range of leaf Mn concentrations (2–17 mg g₋₁ dry weight). When measured in the steady state of photosynthesis under ambient air at 430 μmol quanta m₋₂ s₋₁, the levels of photochemical quenching (qP) and the excitation capture efficiency of open PSII (F'^v/F'^m) declined with Mn accumulation in leaves. The present results suggest that excess Mn in leaves affects the activities of the CO² reduction cycle rather than the potential efficiency of photochemistry, leading to increases in Q_A reduction state and thermal energy dissipation, and a decrease in quantum yield of PSII in the steady state.     

Elevated CO2 enhances stomatal responses to osmotic stress and abscisic acid in Arabidopsis thaliana

A , carbon assimilation rate
ABA, abscisic acid
Ci , intercellular space CO₂ concentration
g , leaf conductance
WUE, water use efficiency

Carbon dioxide and abscisic acid (ABA) are two major signals triggering stomatal closure. Their putative interaction in stomatal regulation was investigated in well-watered air-grown or double CO₂-grown Arabidopsis thaliana plants, using gas exchange and epidermal strip experiments. With plants grown in normal air, a doubling of the CO₂ concentration resulted in a rapid and transient drop in leaf conductance followed by recovery to the pre-treatment level after about two photoperiods. Despite the fact that plants placed in air or in double CO₂ for 2 d exhibited similar levels of leaf conductance, their stomatal responses to an osmotic stress (0·16–0·24 MPa) were different. The decrease in leaf conductance in response to the osmotic stress was strongly enhanced at elevated CO₂. Similarly, the drop in leaf conductance triggered by 1 μ M ABA applied at the root level was stronger at double CO₂. Identical experiments were performed with plants fully grown at double CO₂. Levels of leaf conductance and carbon assimilation rate measured at double CO₂ were similar for air-grown and elevated CO₂-grown plants. An enhanced response to ABA was still observed at high CO₂ in pre-conditioned plants. It is concluded that: (i) in the absence of stress, elevated CO₂ slightly affects leaf conductance in A. thaliana ; (ii) there is a strong interaction in stomatal responses to CO₂ and ABA which is not modified by growth at elevated CO₂.     

Ecophysiological responses to simulated canopy gaps of two tree species of contrasting shade tolerance in elevated CO2

1. One-year-old seedlings of shade tolerant Acer rubrum and intolerant Betula papyrifera were grown in ambient and twice ambient (elevated) CO₂, and in full sun and 80% shade for 90 days. The shaded seedlings received 30-min sun patches twice during the course of the day. Gas exchange and tissue–water relations were measured at midday in the sun plants and following 20 min of exposure to full sun in the shade plants to determine the effect of elevated CO₂ on constraints to sun-patch utilization in these species.
2. Elevated CO₂ had the largest stimulation of photosynthesis in B. papyrifera sun plants and A. rubrum shade plants.
3. Higher photosynthesis per unit leaf area in sun plants than in shade plants of B. papyrifera was largely owing to differences in leaf morphology. Acer rubrum exhibited sun/shade differences in photosynthesis per unit leaf mass consistent with biochemical acclimation to shade.
4. Betula papyrifera exhibited CO₂ responses that would facilitate tolerance to leaf water deficits in large sun patches, including osmotic adjustment and higher transpiration and stomatal conductance at a given leaf-water potential, whereas A. rubrum exhibited large increases in photosynthetic nitrogen-use efficiency.
5. Results suggest that species of contrasting successional ranks respond differently to elevated CO₂, in ways that are consistent with the habitats in which they typically occur.     

Effects of manganese toxicity on leaf CO2 assimilation of contrasting common bean genotypes

Parameters related to leaf photosynthesis were evaluated in three genotypes of common bean ( Phaseolus vulgaris L.) with contrasting tolerance to Mn toxicity. Two short-term studies in solution culture were used to assess the effect of excess Mn on CO₂ assimilation in mature and immature leaves. Mn toxicity decreased total chlorophyll content only in immature leaves, with a consequent reduction of leaf CO₂ assimilation. Mature leaves that showed brown speckles characteristic of Mn toxicity, did not suffer any detriment in their capacity to assimilate CO₂, at least in a 4-day experiment. Stomatal conductance and transpiration were not affected by the presence of high levels of Mn in leaf tissue. Lower stomatal conductance and transpiration rates were observed only in leaves with advanced chlorosis. Differences among genotypes were detected as increased chlorosis in the more sensitive genotype ZPV-292, followed by A-283 and less chlorosis in the tolerant genotype CALIMA. Since CO₂ assimilation expressed per unit of chlorophyll was not different between high-Mn plants and control plants, we conclude that the negative effect of Mn toxicity on CO₂ assimilation can be explained by a reduction in leaf chlorophyll content.     

Effects of intermittent exposure to high atmospheric CO2 on vegetative growth in soybean

Short-term exposure to high CO₂ increases rates of photosynthesis and growth in soybeans, but with prolonged high CO₂ exposure, these high rates are sometimes not maintained. Growth of soybean (Glycine max (L.) Merrill cv. Fiskeby V) seedlings kept for 25 days at atmospheres of 350 or 1000 μ/l CO₂ was compared with growth of plants given 2, 4 or 6 day alternating exposure to high and low CO₂ levels (13 days of total exposure to each level). Final dry weight of plants increased with number of days in high CO₂ but leaf areas were not greatly affected. Thus dry weight gains per unit leaf area (net assimilation rates) were higher in high CO₂ than in low CO₂ throughout the entire period of the experiment and the pattern of exposure to high CO₂ did not affect the rate of dry weight gain in high CO₂.     

Elevated CO2 and conifer roots: effects on growth, life span and turnover

Elevated CO₂ increases root growth and fine (diam. 2 mm) root growth across a range of species and experimental conditions. However, there is no clear evidence that elevated CO₂ changes the proportion of C allocated to root biomass, measured as either the root:shoot ratio or the fine root:needle ratio. Elevated CO₂ tends to increase mycorrhizal infection, colonization and the amount of extramatrical hyphae, supporting their key role in aiding the plant to more intensively exploit soil resources, providing a route for increased C sequestration. Only two studies have determined the effects of elevated CO₂ on conifer fine-root life span, and there is no clear trend. Elevated CO₂ increases the absolute fine-root turnover rates; however, the standing crop root biomass is also greater, and the effect of elevated CO₂ on relative turnover rates (turnover:biomass) ranges from an increase to a decrease. At the ecosystem level these changes could lead to increased C storage in roots. Increased fine-root production coupled with increased absolute turnover rates could also lead to increases in soil organic C as greater amounts of fine roots die and decompose. Although CO₂ can stimulate fine-root growth, it is not known if this stimulation persists over time. Modeling studies suggest that a doubling of the atmospheric CO₂ concentration initially increases biomass, but this stimulation declines with the response to elevated CO₂ because increases in assimilation are not matched by increases in nutrient supply.     

Effects of prolonged restriction in water supply on photosynthesis, shoot development and storage root yield in sweet potato

Besides the paucity of information on the effects of drought stress on photosynthesis and yield in sweet potato [ Ipomoea batatas (L.) Lam.], available reports are also contradictory. The aim of this study was to shed light on the effects of long-term restricted water supply on shoot development, photosynthesis and storage root yield in field-grown sweet potato. Experiments were conducted under a rainout shelter where effects of restricted water supply were assessed in two varieties (Resisto and A15). Large decreases in stomatal conductance occurred in both varieties after 5 weeks of treatment. However, continued measurements revealed a large varietal difference in persistence of this response and effects on CO₂ assimilation. Although restricted water supply decreased leaf relative water content similarly in both varieties, the negative effects on stomatal conductance disappeared with time in A15 (indicating high drought acclimation capacity) but not in Resisto, thus leading to inhibition of CO₂ assimilation in Resisto. Chlorophyll a fluorescence measurements, and the relationship between stomatal conductance, intercellular CO₂ concentration and CO₂ assimilation rate, indicated that drought stress inhibited photosynthesis primarily through stomatal closure. Although yield loss was considerably larger in Resisto, it was also reduced by up to 60% in A15, even though photosynthesis, expressed on a leaf area basis, was not inhibited in this variety. In A15 yield loss appears to be closely associated with decreased aboveground biomass accumulation, whereas in Resisto, combined effects on biomass accumulation and photosynthesis per unit leaf area are indicated, suggesting that research aimed at improving drought tolerance in sweet potato should consider both these factors.     

Biomass, reproductive output, and physiological responses of rapid-cycling Brassica (Brassica rapa) to ozone and modified root temperature

Brassica rapa L. (rapid-cycling Brassica), was grown in environmentally controlled chambers to determine the interactive effects of ozone (O₃) and increased root temperature (RT) on biomass, reproductive output, and photosynthesis. Plants were grown with or without an average treatment of 63 ppb O₃. RT treatments were 13°C (LRT) and 18°C (HRT). Air temperatures were 25°C/15°C day/night for all RT treatments.
Ozone affected plant biomass more than did root temperature. Plants in O₃ had significantly smaller total plant d. wt, shoot weight, leaf weight, leaf area and leaf number than plants grown without O₃. LRT plants tended to have slightly smaller total plant d. wt, shoot weight, root weight, leaf weight, leaf area, and leaf number than HRT plants. For all variables, LRT plants grown in O₃ had the smallest biomass, and plants grown in HRT without O₃ had the largest biomass.
Ozone reduced both fruit weight and fruit number; LRT also reduced fruit weight but had no effect on fruit number. Ozone reduced photosynthesis but RT had no effect. Conductance and internal CO₂ were unaffected by O₃ or RT.
These studies indicate that plant growth with LRT might be more reduced in the presence of O₃ than growth in plants with HRT, which might be able to compensate for O₃-caused reductions in photosynthesis to avoid decreased biomass and reproductive output.     

Interaction of elevated CO2 and O3 on growth, photosynthesis and respiration of three perennial species grown in low and high nitrogen

Seedlings of three species native to central North America, a C₃ tree, Populus tremuloides Michx., a C₃ grass, Agropyron smithii Rybd., and a C₄ grass, Bouteloua curtipendula Michx., were grown in all eight combinations of two levels each of CO₂, O₃ and nitrogen (N) for 58 days in a controlled environment. Treatment levels consisted of 360 or 674 μmol mol^-1 CO₂, 3 or 92 nmol mol^-1 O₃, and 0.5 or 6.0 m M N. In situ photosynthesis and relative growth rate (RGR) and its determinants were obtained at each of three sequential harvests, and leaf dark respiration was measured at the second and third harvests. In all three species, plants grown in high N had significantly greater whole-plant mass, RGR and photosynthesis than plants grown in low N. Within a N treatment, elevated CO₂ did not significantly enhance any of these parameters nor did it affect leaf respiration. However, plants of all three species grown in elevated CO₂ had lower stomatal conductance compared to ambient CO₂-exposed plants. Seedlings of P. tremuloides (in both N treatments) and B. curtipendula (in high N) had significant ozone-induced reductions in whole-plant mass and RGR in ambient but not under elevated CO_2. This negative O₃ impact on RGR in ambient CO₂ was related to increased leaf dark respiration, decreased photosynthesis and/or decreased leaf area ratio, none of which were noted in high O₃ treatments in the elevated CO₂ environment. In contrast, A. smithii was marginally negatively affected by high O_3.