Gender-specific responses of Populus tremuloides to atmospheric CO2 enrichment


  • Xianzhong Wang,

    Corresponding author
    1. Department of Evolution, Ecology and Organismal Biology, The Ohio State University, 1735 Neil Avenue, Columbus, OH 43210–1293, USA
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  • Peter S. Curtis

    1. Department of Evolution, Ecology and Organismal Biology, The Ohio State University, 1735 Neil Avenue, Columbus, OH 43210–1293, USA
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Author for correspondence: Xianzhong Wang Tel: +1 845 365 8995 Fax: +1 845 365 8150


  •  Dioecious species represent an important component of terrestrial ecosystems, but little is known about gender-specific responses to elevated atmospheric CO2.
  •  In an open-top chamber experiment carried out in Michigan, USA, the physiological and growth responses were studied of male and female Populus tremuloides to elevated CO2 and soil nitrogen concentrations.
  •  Male trees had a higher net photosynthetic rate than female trees, but the difference was greater at elevated (25%) than at ambient (13%) CO2. Leaf dark respiration, averaged across the growing season, tended to be higher in males than in females, and increased significantly in male and female trees with CO2 enrichment. Female trees had higher total biomass than male trees grown in low-nitrogen soil and at ambient CO2, but not in other treatments. Elevated CO2 increased the total biomass of males by 58–66% and of females by 22–70%.
  •  Differing physiological and growth responses to CO2 enrichment by male and female trees should be taken into consideration when predicting the effects of global environmental changes on forest ecosystem structure and functioning.


Dioecious species are an important component of terrestrial ecosystems, representing more than 14 000 angiosperm species (Renner & Ricklefs, 1995). Many dominant woody species in forest ecosystems, including members of Salicaceae and Aceraceae, are dioecious. Male and female individuals of dioecious species have been shown to differ physiologically and ecologically from one another at current concentrations of atmospheric carbon dioxide (CO2). For example, female Acer negundo had a higher photosynthetic rate than males (Dawson & Ehleringer, 1993), while the opposite was true for Salix arctica (Jones et al., 1999). Female A. negundo, S. arctica and Thalictrum fendleri significantly outnumbered males in mesic or high-nutrient sites, while males were more numerous in xeric or low-nutrient sites (Freeman et al., 1976; Dawson & Bliss, 1989; Dawson & Ehleringer, 1993). Grant & Mitton (1979) found that the ratio of female to male Populus tremuloides clones on the Front Range in Colorado was 1.27 below 2450 m of elevation, but was only 0.56 above 2900 m.

The physiological and ecological differences between genders were hypothesized to have arisen as a result of different reproductive costs incurred by male and female plants, with females typically investing more in reproduction than do males (Lloyd & Webb, 1977; Allen & Antos, 1988; Dawson & Ehleringer, 1993; Dawson & Geber, 1999). Growth and physiological differences between genders have also been observed before reproduction (Bourdeau, 1958). If carbon assimilation of male and female individuals of dioecious species is differentially affected by CO2 enrichment, their productivity, distribution and population structure might be altered as atmospheric CO2 concentration rises. However, little is known about the gender-specific physiological responses to elevated CO2 by P. tremuloides or any other species. Because of their prominence in terrestrial ecosystems, not accounting for gender-specific variation in carbon assimilation in dioecious species could lead to incorrect estimates of the potential responses of plants to global environmental changes (Jones et al., 1999).

While it has been well documented that atmospheric CO2 enrichment can substantially increase photosynthesis and plant growth (Strain & Cure, 1986, 1994; Curtis & Wang, 1998), our understanding of the effects of rising CO2 on plant dark respiration (Rd) is much less certain. The mechanisms controlling photosynthetic and respiratory responses to CO2 are different (Bunce & Ziska, 1996) and what we have learned about photosynthetic responses to elevated CO2 cannot be readily extrapolated to respiratory responses. Indeed, Rd at elevated CO2 has been found to increase significantly in some studies, but to decrease significantly in others (Amthor, 1991; Poorter et al., 1992; Drake et al., 1999; Amthor, 2000). The importance of Rd as a component of the plant and ecosystem carbon budget, however, must not be overlooked, since up to 50% of carbon assimilated by photosynthesis can be lost through respiration (Kira, 1975; Amthor, 1989; Bunce, 1994). Despite their different responses to elevated CO2, photosynthesis and respiration are two interdependent processes, and both provide reductants and ATP to meet energy demands for growth and maintenance (Kromer, 1995; Foyer & Noctor, 2000). It is therefore reasonable to expect a possible differential effect of CO2 enrichment on the respiration of male and female P. tremuloides plants, if their photosynthesis differs in responding to elevated CO2. To our knowledge, no studies have been conducted to investigate the effects of elevated CO2 on Rd of male and female individuals of any dioecious species.

The primary objective of our study was to examine the carbon assimilation physiology and growth of male and female P. tremuloides trees grown at ambient or elevated CO2. We chose P. tremuloides, a dioecious woody species, because it is the most widespread tree species in North America (Fowells, 1965) and is important both ecologically and economically (Farmer, 1996). Although the trees in our study did not reach reproductive size, gender-related differences in physiology and growth in dioecious species often occur before the onset of flowering (Bourdeau, 1958). We hypothesized that female trees would have higher photosynthetic and respiratory rates than male trees, and that soil nitrogen (N) availability would interact with CO2 levels in affecting physiological and growth responses to CO2 enrichment. Our secondary objective was to gain insight into the mechanisms of CO2 effects on leaf Rd by investigating the relationship between leaf Rd and leaf chemistry. We hypothesized that leaf Rd would be positively related to the supply of respiratory substrates, or leaf carbohydrates, as well as to the demand for energy from respiration, indicated by leaf N level.

Materials and Methods

Plant growth

Our experiment was conducted at the University of Michigan Biological Station (UMBS), Michigan, USA (45°34′ N, 84°40′ W). We chose two previously identified and marked female clones and two male clones of Populus tremuloides Michx (trembling aspen) from clones growing naturally in the Pellston Plain, approx. 5 km from the experimental site (female clones 29 and 44, and male clones 42 and 61, from Barnes (1969)). Saplings were propagated from root segments of approx. 2-cm diameter at the Ohio State University. One sapling of each clone was transplanted on 24 May 1997 into each of 16 1.2 m × 1.2 m × 0.4 m open-bottom wooden boxes at UMBS, hence there were four saplings in each box. Eight boxes were filled with the A horizon of a Kalkaska series topsoil (high-N soil treatment), a common soil type in Michigan, while the other eight were filled with a mixture of 20% Kalkaska topsoil and 80% Rubicon sand taken from the C horizon at the experimental site (low-N soil treatment). Net N mineralization was significantly higher in high-N soil (318 ng N g−1 d−1) than in low-N soil (62 ng N g−1 d−1) (Zak et al., 2000) (see Curtis et al., 2000 for other physical and chemical properties of the soils).

Open-top chambers, of dimensions 0.85 m × 0.85 m × 1.8 m, were used to manipulate atmospheric CO2 concentration (Curtis & Teeri, 1992). One chamber was put on top of each wooden box. Pure CO2 was dispensed via manual flow meters into input blowers and then into eight elevated CO2 chambers to increase CO2 concentration, which was monitored with an infrared gas analyser (LI-6262, Li-Cor Inc., Lincoln, NB, USA). Daytime (07:00–19:00 h) and night-time (19:00–07:00 h) CO2 concentrations were 728 ± 16.5 and 799 ± 43.3 µmol mol−1 (mean ± SE), respectively, for the elevated CO2 treatment, and 372 ± 6.7 and 391 ± 4.7 µmol mol−1 (mean ± SE), respectively, for the ambient CO2 treatment. Elevated CO2 treatment was maintained from 27 May to 16 October 1997. The temperature inside the chambers was 1.34 ± 0.24°C (mean ± SE) higher than outside the chambers. The plastic chamber covering (0.02-cm polyvinyl chloride film with UV inhibitors) transmitted approximately 80% of photosynthetically active radiation (PAR). The trees were kept well watered throughout the experiment. In mid-June, ammonium nitrate was applied to all chambers at a rate of 2 g N m−2, which was equivalent to the amount mineralized annually from the local xeric oak ecosystems (Zak & Pregitzer, 1990). The chambers were vertically extended by 0.8 m on 17 July to accommodate rapid tree growth. Plants were destructively harvested beginning on 16 October 1997. All leaves were measured for total leaf area with an LI-3000 leaf area meter (Li-Cor Inc., Lincoln, NB, USA). Leaves, stems, and coarse and fine roots from each tree were harvested, dried and weighed separately.

Gas exchange

Net CO2 assimilation rates at saturating PAR (A) and photosynthetic light responses were measured between 10:00 and 16:00 h. The CO2 concentration in the cuvette during Rd measurements was the growth CO2 concentration, i.e. approx. 370 µmol mol−1 for the ambient and 730 µmol mol−1 for the elevated CO2 treatment. Daytime leaf Rd was measured five times during the growing season by shading leaves during daytime. Night-time leaf Rd measurements were made after 23:00 h on 18 and 19 August. During Rd measurements, we took care to ensure that the cuvette was not leaking by blowing high-CO2 air on to the cuvette from time to time. If the cuvette gasket was damaged, the CO2 concentration in the sampling chamber would rise quickly, indicating a leak in the cuvette. To minimize possible human bias, three readings were logged at an interval of 1–2 min and the mean of the three readings was used as Rd for the leaf measured. All gas exchange measurements were made on the youngest mature leaves using an LI-6400 Portable Photosynthesis System (Li-Cor Inc., Lincoln, NB, USA). The leaf temperature inside the cuvette was maintained at 27.7 ± 0.2°C (mean ± SE), and the relative humidity in the cuvette was kept at approx. 60%. Leaves were allowed to equilibrate for 5 min before any measurement was taken. Stem Rd measurements were made from 21 to 27 July using a custom-made cuvette attached to the LI-6400 Portable Photosynthesis System (Li-Cor Inc.). The apparent photosynthetic quantum yield (Φ) and light compensation point (LCP) were derived by linear regression of A against PAR between 0 and 200 µmol m−2 s−1.

For A vs intercellular CO2 concentration (Ci) curves, leaves were first exposed to their growth CO2 concentration until steady-state gas exchange was observed. The intercellular CO2 concentration was then reduced to approx. 80 µmol mol−1 and increased stepwise to 1000 µmol mol−1. The maximum rate of CO2 fixation by Rubisco (Vcmax), maximum electron transport rate (Jmax) and rate of inorganic phosphate turnover (PiRC) were estimated from A vs Ci data (Farquhar et al., 1980; Harley et al., 1992). The relative effect of elevated CO2 on AA) or on stomatal conductance (Δgs) was calculated as the difference in A or gs at elevated compared with ambient CO2 divided by the rate at ambient CO2.

Leaf chemistry

Immediately after A and leaf Rd measurements, 2.6-cm2 leaf discs were excised from the same leaf, frozen on dry ice and stored at −80°C until lyophilized. Total leaf N was analysed using a Carlo Erba CHN analyser (Milan, Italy). For carbohydrate analysis, samples were first extracted with 80% ethanol at 80°C. The supernatant was evaporated to dryness and then re-dissolved in aqueous polyvinylpolypyrrolidone solution. Soluble carbohydrates were analysed enzymatically using a modification of the procedure developed by Jones et al. (1977). Starch concentration was determined by the degradation of starch by amyloglucosidase to glucose and assayed for glucose enzymatically. The ethanol-extracted tissue pellet was suspended in 0.2 N potassium hydroxide (KOH), boiled for 20 min, and brought to pH 7.0 with 1.0 N acetic acid (CH3COOH). Amyloglucosidase was added to the re-suspended pellet and the mixture was incubated at 55°C in a water bath for 1 h to degrade the starch. The starch concentration was determined using the same procedure as for soluble sugars and calculated as glucose equivalent.

Statistical analysis

The effects of CO2 and soil N availability were analysed by repeated-measures analysis of variance (ANOVA) for split plot randomized complete block design, where the chamber was the experimental unit. The main effects (CO2 and soil N availability) and CO2 × soil N availability interaction were tested over the CO2 × soil N nested within block mean square, i.e. block (CO2 × N). Since genotype identities were not the same across genders, we tested for genotype effects within males and females separately. There were no significant differences between male genotypes or between female genotypes for any variable. Hence the split plot effect of gender and all treatment interactions with gender were tested over the error mean square. Comparisons among CO2 and soil N treatment means were made by LSD for a priori comparisons (between CO2 treatments within a soil N treatment), and by minimum significant difference (MSD) for all a posteriori comparisons (between genders) (Sokal & Rohlf, 1981).


Carbon assimilation and stomatal conductance

Overall, elevated-CO2-grown trees had significantly higher A than did ambient-CO2-grown trees throughout the experiment (Fig. 1, Table 1). Males had significantly higher A than did females at both CO2 treatment levels, and were more responsive to CO2 enrichment than were females. This significant interaction between CO2 and gender resulted in greater differences in A at elevated than at ambient CO2 concentration. For example, on day of year (DOY) 236 (late August), male trees had 16% higher A than female trees at ambient CO2, but 31% higher A at elevated CO2. Soil N availability had no effect on A for either male or female trees (Table 1).

Figure 1.

Seasonal changes in net CO2 assimilation rates of male (squares) and female (circles) Populus tremuloides trees grown at ambient (open symbols) or elevated (filled symbols) CO2. Rates were averaged across high- and low-N soils because there was no N effect or N × CO2 interaction. The asterisks indicate a significant difference between genders within a CO2 treatment at P < 0.05. The vertical bars indicate 1 SE, n = 8.

Table 1.  Mean squares (MS) and F values (F) of repeated measures ANOVA for net CO2 assimilation rate (A), stomatal conductance (gs), and area-based (Rda) and mass-based (Rdm) leaf dark respiration during the day in Populus tremuloides. The main effects (CO2, soil N level and gender) were analysed for split plot randomized complete block design, where the chamber was the experimental unit
  • *

    , P < 0.05;

  • **

    , P < 0.01;

  • ***

    , P < 0.001.

Between subjects         
 CO212592115***0.089 3.9013.822.8*** 218.7 1.28
 N concentrations1  32.1  1.250.06 2.72 2.54 4.18 901.9 5.27*
 CO2 × N concentrations1  42.3  1.650.001 0.06 0.62 1.03  85.3 0.50
 Gender11107 60.7***0.6138.1*** 1.14 7.12*  39.0 0.33
 CO2 × Gender1 217.2 11.9***0.004 0.27 0.16 0.97  29.0 0.25
 N concentrations × gender1   0.87  0.050.002 0.13 0.15 0.95   7.48 0.06
 CO2 × N concentrations × Gender1  26.6  1.460.013 0.79 0.048 0.30   1.65 0.01
Within subjects         
 Time × CO24 429.2 20.0***0.06 4.95*** 1.26 1.96 427.7 3.11*
 Time × N concentrations4  37.9  1.770.046 3.65*** 0.81 1.26 183.2 1.33
 Time × CO2 × N concentrations4  56.6  2.63*0.023 1.86 0.31 0.49  94.6 0.69
 Time × Gender4  51.7  4.59***0.029 6.14*** 0.35 1.56  57.4 0.70
 Time × CO2 × Gender4  13.5  1.200.005 0.99 0.10 0.44  36.8 0.45
 Time × N concentrations × Gender4  28.3  2.51*0.017 3.64*** 0.57 2.54  58.1 0.71
 Time × CO2 × N concentrations × Gender4  31.3  2.78*0.011 2.29 0.02 0.07   9.72 0.12

Stomatal conductance varied significantly during the growing season (Table 1), but was always higher in male than in female trees (Table 2). There were no other treatment effects on gs. We found a strong positive correlation between ΔA and Δgs for male and female trees at ambient and elevated CO2 at both soil N levels (Fig. 2).

Table 2.  Stomatal conductance (gs) and photosynthetic light response parameters of male (M) and female (F) Populus tremuloides trees grown at ambient or elevated CO2 in low- or high-N soil. Photosynthetic and gs measurements were taken from 1 to 8 September 1997. Apparent quantum yield (Φ) and light compensation point (LCP) were derived by linear regression of photosynthetic rate against photosynthetically active radiation (PAR) of 0–200 µmol m−2 s−1. The maximum rate of CO2 fixation by Rubisco (Vcmax), the rate of electron transport (Jmax) and the rate of inorganic phosphate turnover (PiRC) were derived from A/Ci data. Mean (SE), n = 4
 Low NHigh N
 Ambient CO2Elevated CO2Ambient CO2Elevated CO2
  • a

    Similar superscripts within a row indicate no significant difference at P < 0.05.

gs (mol H2O m−2 s−1)  0.46 (0.06)a  0.40 (0.04)b  0.46 (0.06)a  0.38 (0.05)bc  0.46 (0.02)a  0.39 (0.03)bc  0.43 (0.02)ab  0.34 (0.04)c
Φ (mol C mol−1 photon)  0.054 (0.001)c  0.052 (0.001)c  0.065 (0.001)a  0.059 (0.004)b  0.053 (0.001)c  0.052 (0.002)c  0.065 (0.001)a  0.060 (0.002)b
LCP (µmol m−2 s−1) 47.6 (2.84)a 46.0 (4.51)abc 46.4 (2.18)ab 47.8 (3.86)a 43.3 (2.39)bcd 41.6 (3.09)d 42.0 (3.27)cd 44.7 (4.00)abcd
Vcmax (µmol m−2 s−1) 79.5 (1.44)a 69.7 (2.55)b 69.7 (3.51)b 55.3 (5.86)d 82.7 (1.69)a 73.7 (2.75)b 70.2 (1.13)b 64.7 (2.24)c
Jmax (µmol m−2 s−1)157.9 (3.36)a136.3 (4.15)c147.7 (11.6)b117.0 (11.6)d159.3 (6.66)a143.8 (5.67)bc151.7 (3.54)ab139.7 (5.36)c
PiRC (µmol m−2 s−1) 11.5 (0.38)a 10.0 (0.43)b 11.2 (1.15)a  9.0 (0.90)c 11.3 (0.55)a 10.3 (0.55)b 11.2 (0.34)a 10.5 (0.37)b
Figure 2.

The relationship between the relative effects of CO2 enrichment on net CO2 assimilation (ΔA) and stomatal conductance (Δgs) of male (filled symbols) and female (open symbols) Populus tremuloides trees in high-N (triangles) or low-N (circles) soils. ΔA(%) = [1 − ((Ae − Aa)/Aa)] × 100 and Δgs(%) = [1 − ((gse − gsa)/gsa)] × 100, where Aa and Ae denote A, and gsa and gse denote gs, at ambient and elevated CO2, respectively. A and gs were measured at growth CO2 concentrations throughout the growing season.

Male trees had significantly higher Φ than female trees at elevated CO2, but not at ambient CO2 (Table 2). Elevated CO2 increased Φ in both male and female trees, while soil N availability had no effect on Φ. The light compensation point was largely unaffected by gender or CO2, but was higher in low-N- than in high-N-soil-grown plants, except for females grown at elevated CO2 (Table 2).

Male trees had higher Vcmax, Jmax and PiRC than female trees when compared at the same CO2 and soil N levels (Table 2). Elevated CO2 significantly reduced Vcmax of male and female trees in both low- and high-N soils, while Jmax was reduced by CO2 enrichment only in low-N soil. When measured at the same Ci, ambient-CO2-grown trees had higher A than elevated-CO2-grown trees, indicating a significant negative photosynthetic adjustment to elevated CO2 in both males and females (Fig. 3). Since there was no effect of soil N availability on the magnitude of photosynthetic adjustment, A was averaged across soil N concentrations. The adjustment in photosynthetic capacity, calculated as the reduction of A in elevated-compared with ambient-CO2-grown plants both measured at elevated CO2, was 16% for males and 14% for females.

Figure 3.

Net CO2 assimilation rate of female (a) and male (b) Populus tremuloides grown at ambient (open bars) or elevated (hatched bars) CO2 and measured at ambient or elevated CO2. Mean ± 1 SE, n = 8. †, P < 0.10; *, P < 0.05; **, P < 0.01.

Leaf and stem Rd

Averaged across five measurements over the growing season, males had a significantly greater daytime leaf Rda than females grown at elevated CO2 and in low-N soil, but not in other treatments (Table 3). Daytime Rda was significantly higher at elevated than at ambient CO2, but daytime Rdm was unaffected by gender or CO2 treatment (Tables 1, 3). In high-N soil, male trees had a significantly higher night-time leaf Rda than female trees at ambient CO2, but this was reversed at elevated CO2. In low-N soil, night-time leaf Rda did not differ between genders at either ambient or elevated CO2 (Table 3). Like daytime Rda, night-time leaf Rda was significantly higher in elevated- than in ambient-CO2-grown trees. Stem Rda was significantly higher in male than in female trees and in high- than in low-N soils (Table 3). Elevated CO2 increased the stem Rda of both male and female trees in high-N soil, but only in male trees in low-N soil.

Table 3.  Daytime leaf dark respiration expressed on an area (Day Rda) and on a mass (Day Rdm) basis, night-time leaf dark respiration (Night Rda), and stem dark respiration (Stem Rda) of male (M) and female (F) Populus tremuloides grown under conditions of ambient or elevated CO2 and in low- or high-N soils. Daytime Rda and Rdm were the average of five measurements during the growing season. n = 8 for night-time Rda and n = 4 for stem Rda. Mean (1 SE)
 Low NHigh N
 Ambient CO2Elevated CO2Ambient CO2Elevated CO2
  • a

    Similar superscripts within a row indicate no significant difference at P < 0.05.

Day Rda (µmol m−2 s−1) 3.12 (0.12)de 2.99 (0.12)e 3.84 (0.12)a 3.57 (0.12)b 2.99 (0.12)e 2.91 (0.11)e 3.42 (0.12)bc 3.28 (0.12)cd
Day Rdm (µmol kg−2 s−1)46.4 (2.0)abc45.8 (1.98)bc50.5 (1.7)a48.4 (2.0)ab43.1 (1.8)c43.5 (2.0)c44.8 (1.4)bc42.8 (1.8)c
Night Rda (µmol m−2 s−1) 2.27 (0.12)d 2.15 (0.13)de 2.73 (0.12)ab 2.69 (0.15)ab 2.49 (0.16)c 2.11 (0.11)e 2.66 (0.12)b 2.82 (0.11)a
Stem Rda (µmol m−2 s−1) 1.67 (0.19)d 1.56 (0.04)e 1.80 (0.23)c 1.63 (0.21)de 2.41 (0.24)b 1.81 (0.13)c 3.09 (0.26)a 2.47 (0.32)b

Leaf chemistry

Leaf N concentration was significantly higher in ambient- than in elevated-CO2-grown trees and in high- than in low-N soils (Table 4). In the middle of the growing season, the daytime soluble sugar content was unaffected by CO2 treatment, but was higher in high- than in low-N-soil-grown trees. The night-time sugar content was significantly higher at elevated CO2 compared with ambient CO2 in low-N soil, but not in high-N soil. Trees grown at elevated CO2 had significantly higher daytime and night-time starch content compared with those grown at ambient CO2. Specific leaf area (SLA) was significantly greater in ambient- than in elevated-CO2-grown trees in both high- and low-N soils, and greater in low- than in high-N soils at both CO2 levels (Table 4). Leaf chemistry and SLA were largely unaffected by gender.

Table 4.  Leaf nitrogen concentration, daytime and night-time soluble sugar and starch contents, and specific leaf area (SLA) of Populus tremuloides trees grown at ambient or elevated CO2 and in low- or high-N soils in 1997. The data were averaged across male and female trees since the only gender difference was a higher night-time soluble sugar and starch content in female than in male trees grown at elevated CO2 and in high-N soil. Leaves for daytime soluble sugar and starch contents were taken on 29 July, and those for night-time soluble sugar and starch contents on 18 and 19 August, after night-time dark respiration (Rd) measurements. Leaf N content was the seasonal average measured on six different dates. Mean (1 SE), n = 8
 Low-N soilHigh-N soil
 Ambient CO2Elevated CO2Ambient CO2Elevated CO2
  • a

    Similar superscripts within a row indicate no significant difference at P < 0.05.

Leaf N (mg g−1) 27.8 (0.4)b 21.6 (0.4)d 30.1 (0.5)a 25.3 (0.3)c
Soluble sugar (mg g−1)    
 Daytime 95.8 (6.4)b100.5 (3.2)b114.9 (3.7)a119.8 (2.7)a
 Night-time 88.7 (5.8)c102.2 (1.5)a 95.3 (2.9)b 98.2 (6.9)ab
Starch (mg g−1)    
 Daytime 40.1 (4.2)c 66.7 (10.9)b 30.5 (5.5)c 80.4 (10.8)a
 Night-time 26.6 (3.3)c 73.5 (9.9)a 26.2 (2.4)c 47.8 (7.0)b
SLA (cm2 g−1)142.1 (4.0)a129.5 (2.8)b132.4 (3.2)b119.0 (3.0)c

There was a positive correlation between daytime leaf Rda and starch content, regardless of gender and CO2 concentration (Fig. 4a). Averaged across genders and soil N levels, the leaf starch content of elevated-CO2-grown trees was 0.65 ± 0.03 mg cm−2, significantly higher than that of ambient-CO2-grown trees (0.28 ± 0.01 mg cm−2). It is also obvious that elevated-CO2-grown trees had higher daytime leaf Rda than ambient-CO2-grown trees (Fig. 4a). However, there was no correlation between daytime leaf Rda and leaf N content, regardless of genders, CO2 or soil N availability (Fig. 4b).

Figure 4.

Relationships between leaf dark respiration (Rda) and starch content (a), and between leaf Rda and N content (b) for male (squares) and female (circles) Populus tremuloides at ambient (open symbols) or elevated (filled symbols) CO2 and in high-N (symbols with pluses) or low-N (symbols without pluses) soils. Measurements were taken on leaves of different positions at different times of the growing season. Error bars represent 1 SE (n = 40 for leaf starch, n = 24 for leaf N). For regressions between leaf Rda and starch content, P < 0.001 for all trees (n = 320), P = 0.007 for ambient-CO2-grown trees (n = 160), and P = 0.187 for elevated-CO2-grown trees (n = 160). For regression between leaf Rda and N content, P = 0.798 for all trees (n = 192), P = 0.755 for ambient-CO2-grown trees (n = 96), and P = 0.347 for elevated-CO2-grown trees (n = 96).


Gender effects on biomass were largely restricted to trees grown in low-N soils (Table 5). There, females had significantly greater tree height, leaf area, leaf mass, stem mass, coarse root mass and total biomass than males at ambient CO2, while females had greater tree height, leaf area and fine root mass than males at elevated CO2 (Table 5). In high-N soils, the only gender effect was a greater fine root mass in females than in males at elevated CO2.

Table 5.  Tree height, leaf area, leaf mass, stem mass, root mass and total mass of male (M) and female (F) Populus tremuloides trees grown at ambient or elevated CO2 and in low- or high-N soils. Trees were harvested in early October 1997. Mean (SE), n = 8
 Low NHigh N
 Ambient CO2Elevated CO2Ambient CO2Elevated CO2
  • a

    Similar superscripts within a row indicate no significant difference at P < 0.05.

Tree height (cm)115 (6.8)d132 (9.8)b116 (11.9)cd131 (14.3)b138 (4.8)b122 (13.6)bcd165 (7.9)a156 (15.9)a
Leaf area (m2)  0.14 (0.01)c  0.21 (0.02)b  0.15 (0.02)c  0.21 (0.03)b  0.19 (0.01)b  0.20 (0.03)b  0.26 (0.02)a  0.28 (0.05)a
Leaf mass (g) 12.1 (0.58)e 19.1 (2.19)cd 18.7 (2.33)cd 22.9 (3.85)bc 17.1 (1.39)d 17.4 (2.78)d 25.3 (2.55)ab 28.0 (5.37)a
Stem mass (g) 15.8 (1.15)c 25.8 (3.53)b 26.0 (3.97)b 28.3 (5.79)b 23.7 (2.16)b 22.2 (3.32)b 36.1 (3.00)a 39.2 (7.97)a
Coarse root mass (g) 26.4 (1.62)d 36.5 (2.70)c 45.8 (5.80)b 45.5 (7.85)b 36.0 (3.56)c 32.4 (3.57)c 61.0 (3.63)a 56.2 (9.09)a
Fine root mass (g)  4.3 (0.45)e  5.1 (0.65)de  6.5 (0.71)c  8.6 (1.34)ab  5.6 (0.85)cd  6.2 (1.08)c  8.0 (0.72)b  9.4 (0.76)a
Total mass (g) 58.6 (3.01)d 86.5 (8.33)c 97.0 (12.5)bc105.3 (18.4)b 82.4 (7.49)c 72.8 (9.85)c130.4 (8.01)a132.7 (22.4)a

The effects of CO2 enrichment, however, were more profound in high-N soils, where every biomass component of both genders was significantly greater at elevated compared with ambient CO2 (Table 5). In low-N soils, elevated CO2 had no effect on the tree height or leaf area of male and female trees, but significantly increased the leaf and stem mass of male trees. Elevated CO2 significantly increased the total biomass of both males and females, regardless of soil N levels. The high-N treatment significantly increased the biomass components of male trees when compared at the same CO2 concentration (Table 5). While female trees generally had greater biomass in high- than in low-N soils at elevated CO2, they did not respond to soil N availability at all at ambient CO2.


We found significant differences in photosynthetic rates between male and female P. tremuloides at both ambient and elevated CO2; males were more responsive to elevated CO2 than were females. Our results lead us to reject our initial hypothesis that females would have higher A than males. Since gender difference in P. tremuloides occurs before the onset of flowering, the different reproductive costs of males and females (Lloyd & Webb, 1977; Allen & Antos, 1988) may not be the only reason for the physiological and ecological differences between genders. The male P. tremuloides trees were able to sustain a higher rate of carbon assimilation in part because they had higher stomatal conductance throughout the season. The males also had some biochemical characteristics that enabled them to fix CO2 at a higher rate than females. For instance, Rubisco in the leaves of male trees had a higher maximum rate for CO2 fixation as shown by a higher Vcmax. At higher Ci, male trees also were less limited by rates of ribulose-1.5-biphosphate (RuBP) regeneration and triose phosphate utilization because they had higher Jmax and PiRC. In addition, since soil N availability did not affect these responses, it is likely that gender-specific differences in carbon assimilation at high CO2 will persist, even when soil nutrient levels are low. This is important because low soil N availability is a major factor limiting carbon assimilation at elevated CO2 (Gunderson & Wullschleger, 1994; Curtis et al., 2000). However, other environmental factors such as temperature might also affect the responses of dioecious species to elevated CO2. Jones et al. (1999) found that elevated-CO2-grown female S. arctica had lower A than males at 12°C, while the opposite was true at 5°C.

Although male P. tremuloides had higher A compared with females throughout the growing season, females had significantly greater biomass components than males in low-N soil and at ambient CO2. This discrepancy in the A and biomass responses to elevated CO2 was most likely due to the greater leaf area and therefore greater carbon assimilation in females in low-N soil. Another possible explanation for this discrepancy was the lower stem Rda in females than in males. Stem Rda can comprise a sizeable portion of the whole plant carbon budget because of the large stem surface area. In young Betula pendula trees, for example, stem respiration accounted for as much as 23% of total plant respiration (Wang et al., 1998). Higher stem Rda in male trees could therefore have offset the higher A and resulted in lower biomass accumulation in male trees. Our results are consistent with those of Laporte & Delph (1996), who found that male Silene latifolia had consistently higher A, but failed to grow larger than females. Sakai & Burris (1985) found that female P. tremuloides clones growing in nutrient-poor soils near our research site had larger numbers of ramets and greater basal area than did male clones, indicating more vegetative growth by females in this habitat. In contrast, Dawson & Ehleringer (1993) observed greater vegetative growth in male than female A. negundo in xeric habitats, even when males showed lower A than females.

We observed significant negative adjustment of photosynthesis at high CO2 in both males and females, but there was little difference in adjustment between genders, notwithstanding that males had a smaller reduction in Vcmax, Jmax and PiRC than females at elevated CO2. The magnitude of photosynthetic adjustment found in our study (14–16%) was smaller than the overall 21% negative adjustment across 39 species in 20 studies reported by Gunderson & Wullschleger (1994). In a study with hybrid poplar (Populus × euramericana) on the same research site, Curtis et al. (1995) found that the level of adjustment ranged from 9% stimulation to 52% reduction, with the average being an 18% reduction in photosynthetic capacity. Negative adjustment of A was also more profound in P. × euramericana grown in low- than in high-N soil. In a meta-analysis of 24 studies, we found significant negative adjustment in plants grown in small pots (< 0.5 l), but no consistent evidence for overall negative adjustment in plants grown in large pots (Curtis & Wang, 1998). It seems that photosynthetic adjustment is a common but not universal response by plants to growth under high concentrations of atmospheric CO2.

We found no CO2 effect on gs in either male or female trees, although males had significantly higher gs than females, with the difference being greater at elevated than at ambient CO2. This is different from what we observed in well-watered male P. tremuloides genotypes, where elevated CO2 significantly reduced the gs of plants in both low- and high-N soil (Wang et al., 2000). In both studies, however, we found that plants with less sensitive stomatal responses to CO2 enrichment were more responsive photosynthetically to elevated CO2; in other words, the less the reduction of gs at high CO2, the greater the photosynthetic responses. The variation in gs response to CO2 enrichment in P. tremuloides is typical of the substantial variation in gs responses to CO2 observed among woody plants. We also found that females had greater fine root mass than males at elevated CO2. Since some areas are likely to become drier with rising atmospheric CO2 and increased evaporation (Rind et al., 1990), forest ecosystems in these regions will probably be subject to more frequent and/or severe drought stress. Greater fine root mass and lower stomatal conductance could allow female trees to develop a competitive edge over male trees in chronically dry regions or during drought.

Elevated CO2 can have direct and indirect effects on Rd (Amthor, 1991; Thomas & Griffin, 1994). What was observed in our study was primarily the indirect effect of elevated CO2 because leaf Rd was measured at growth CO2 concentrations. The increased daytime leaf Rda at elevated CO2 that we observed can be explained in part by lower SLA, hence greater leaf biomass per unit leaf area, and greater total nonstructural carbohydrate (TNC) in higher-CO2-grown plants. Since enzymes catalysing respiratory reactions are generally present in amounts that exceed that required to explain the observed rates of respiration in mature leaves (Amthor, 1991), higher leaf carbohydrate content at high CO2 commonly leads to higher leaf Rda (Azcon-Bieto & Osmond, 1983; Farrar, 1985; Hrubec et al., 1985; Amthor, 1989; Lambers et al., 1989; Thomas et al., 1993). Higher carbohydrate content might also enhance leaf Rd through increased phloem loading and translocation (Amthor, 2000), which require a higher rate of respiration. The lower leaf Rda observed in elevated-CO2-grown plants in some studies (Lambers et al., 1989; Baker et al., 1992; Wullschleger & Norby, 1992) has been attributed to lower leaf N or protein content compared with ambient-CO2-grown plants. We found no correlation between daytime leaf Rda and N content, which was significantly lower in elevated-CO2-grown plants, but we did observe a positive correlation between leaf Rda and starch content, which was significantly higher in elevated-CO2-grown plants. We initially hypothesized that leaf Rda would be positively correlated to both starch and N contents, but our results suggest that the respiratory substrate level is more important than the total leaf N content in determining leaf Rda in fast-growing P. tremuloides. This suggestion is supported by the results of Azcon-Bieto & Osmond (1983), who found a positive correlation between Rda and carbohydrate level and between Rda and A, which has shown a consistent stimulation by elevated CO2 in a variety of C3 species.

In summary, we found differential effects of elevated CO2 on carbon assimilation in the male and female P. tremuloides trees that we studied. As a result, the productivity, distribution and population structure of P. tremuloides may be altered as atmospheric CO2 concentration rises if the gender difference is widespread. Because of the prominence of dioecious species in terrestrial ecosystems, gender-specific physiological responses provide a new mechanism by which community structure and functioning might be affected by atmospheric CO2 enrichment.


This research was supported in part ($5000, or 40% of the total cost) by the National Institute for Global Environmental Change (NIGEC) through the US Department of Energy (Cooperative Agreement No. DE-FC03–90ER61010). Any opinions, findings and conclusions or recommendations expressed herein are those of the authors and do not necessarily reflect the views of the DOE. This research was also supported by an Ohio State University Graduate Student Alumni Research Award to XZW and the University of Michigan Biological Station. We gratefully acknowledge the constructive comments and suggestions by Dr Richard Norby and two anonymous reviewers. We thank Dr James Teeri and the staff of the University of Michigan Biological Station for logistical support. Our special thanks also go to Drs Donald Zak and David Karowe for their help in setting up this experiment, and Dr Chris Vogel for diverse technical support.