Journal of Experimental Botany, Vol. 47, No. 294, pp. 61-69, January 1996
Journal of
Experimental
Botany
Long-term effects of a doubled atmospheric C0 2
concentration on the CAM species Agave deserti
Eric A. Graham and Park S. Nobel1
UCLA-DOE Laboratory and Department of Biology, University of California, 900 Veteran Avenue,
Los Angeles, CA 90024-1786, USA
Received 9 March 1995; Accepted 25 September 1995
Abstract
Introduction
To examine the effects of a doubled atmospheric CO2
concentration and other aspects of global climate
change on a common CAM species native to the
Sonoran Desert, Agave deserti was grown under 370
and 750//mol CO2 mol 1 air and gas exchange was
measured under various environmental conditions.
Doubling the CO2 concentration increased daily net
CO2 uptake by 49% throughout the 17 months and
decreased daily transpiration by 24%, leading to a
110% increase in water-use efficiency. Under the
doubled CO2 concentration, the activity of ribulose1,5-bisphosphate carboxylase/oxygenase (Rubisco)
was 11 % lower, phosphoenolpyruvate carboxylase was
34% lower, and the activated:total ratio for Rubisco
was 25% greater than under the current CO2 concentration. Less leaf epicuticular wax occurred on
plants under the doubled CO2 concentration, which
decreased the reflectance of photosynthetic photon
flux [PPF); the chlorophyll content per unit leaf area
was also less. The enhancement of daily net CO2
uptake by doubling the C0 2 concentration increased
when the PPF was decreased below 25 mol m~2 d ~ \
when water was withheld, and when day/night temperatures were below 17/12 °C. More leaves, each with a
greater surface area, were produced per plant under
the doubled CO2 concentration. The combination of
increased total leaf surface area and increased daily
net CO2 uptake led to an 88% stimulation of dry mass
accumulation under the doubled CO2 concentration. A
rising atmospheric C0 2 concentration, together with
accompanying changes in temperature, precipitation,
and PPF, should increase growth and productivity of
native populations of A. deserti.
The global atmospheric CO2 concentration is steadily
increasing and may double by the second half of the
twenty-first century (King et al, 1992; Rogers and
Dahlman, 1993). An increased CO2 concentration has
various effects on plants, depending on the carbon fixation
pathway, the duration of exposure, and the accompanying
environmental conditions. For instance, doubling the CO2
concentration can increase net CO2 uptake and growth
for C3 plants, reflecting the increased efficiency of ribulose-l,5-bisphosphate carboxylase/oxygenase (Rubisco)
forfixingCO2 at higher COJOj ratios (Bowes, 1991; Idso
and Kimball, 1992). Exposure to a doubled CO2 concentration increased shoot and whole plant dry mass for 19
out of 25 native herbaceous C3 species (Hunt et al, 1991).
Acclimation to increased CO2 concentration after a shortterm exposure of 4-6 weeks can occur, returning net CO2
uptake and growth to previous lower rates (Sage et al,
1989; Yelle et al., 1989; Stitt, 1991). Little effect on CO2
uptake generally occurs for C4 plants under increased CO2
concentrations, reflecting the relative insensitivity of phosphoenolpyruvate carboxylase (PEPCase) to COz/Oj ratios
(Lawlor and Mitchell, 1991; Stitt, 1991). In addition,
alterations in temperature, radiation, and rainfall patterns
accompanying global climate change can alter the productivity and competitive ability of crop and native plants
under increased CO2 concentrations (Szarek et al., 1987;
Huerta and Ting, 1988; Nobel and Israel, 1994).
Contradictions exist among the relatively few studies
on increased CO2 concentration for Crassulacean acid
metabolism (CAM) plants. For instance, doubling the
CO2 concentration has little effect on Kalanchoe daigremontiana (Holtum et al., 1983) and Ananas comosus
(Ziska et al, 1991), increases nocturnal CO2 uptake for
Opuntiaficus-indica(Cui et al, 1993) and well-watered
Agave vilmoriniana (Szarek et al, 1987), but reduces
Key words: Crassulacean acid metabolism, gas exchange,
global climate change, Sonoran Desert.
1
To wtiom correspondence should be addressed. Fax: + 1 310 825 9433.
© Oxford University Press 1996
62
Graham and Nobel
nocturnal CO2 uptake for Portulacaria afra (Huerta and
Ting, 1988). Carbon fixation by CAM plants combines
aspects of both C3 and C4 plants, possibly accounting for
some of the variation observed. In particular, many CAM
plants, under favourable environmental conditions, will
take up substantial amounts of CO2 both during the
daytime, using Rubisco for the initial fixation of CO2
into stable products, and at night using the CAM pathway
and PEPCase (Nobel, 1988). Of the studies on the effects
of increased CO2 concentration on CAM plants, few have
examined the long-term effects on growth or the interactive effects of CO2 concentration with other environmental
conditions on net CO2 uptake.
Agave deserti is a relatively slow-growing succulent
CAM species with a basal rosette of leaves that unfold
from a central spike. It is native to the Sonoran Desert
where locally it can be the dominant species and a major
contributor to biomass production of the community
(Nobel, 1976; Gentry, 1982). The effects of environmental
variables such as light, temperature, and water availability
on gas exchange of A. deserti have been studied in the
laboratory and in the field (Nobel, 1976, 1984, 1988). Of
the other CAM plants that have been as extensively
studied, only a few species, including the cultivated and
highly productive O. ficus-indica, have been the subject
of further studies on the effects of increased CO2 concentrations (Cui and Nobel, 1994; Nobel and Israel, 1994).
Agave deserti should be more representative of less productive native CAM plants and, as a species whose
growth and gas exchange have been studied extensively,
it is well suited for the investigation of long-term effects
on growth, morphology and gas exchange of a CAM
plant exposed to various environmental conditions under
a doubled CO2 concentration.
Materials and methods
Plant material
Twenty-four plants of Agave deserti Engelm. (Agavaceae) with
an average of 12 leaves, 1.5 kg fresh mass, and 275 g dry mass
per plant were collected from the University of California Philip
L. Boyd Deep Canyon Desert Research Center at Agave Hill
(33-38^, 116°24'W, 850 m elevation). They were planted in
0.013 m 3 pots in soil collected from Agave Hill and divided
randomly into two groups; each group was transferred to a
Conviron E-15 environmental chamber (Controlled Environments, Pembina, North Dakota). Plants were acclimated to the
chambers for 1 month before the start of the experiment with
a photoperiod of 12 h at a photosynthetic photon flux (PPF,
wavelengths from 400-700 nm) of 22 mol m ~ 2 d ~ ' on a
horizontal plane at the top of the canopy (15 mol m ~ 2 d ~ ' on
the adaxial surfaces of the leaves used for gas exchange
measurements), measured with a LI-191SB Line Quantum
Sensor (Li-Cor, Lincoln, Nebraska). Day/night air temperatures
were 25/20°C (day/night leaf temperatures averaged 28/19 °C).
The plants were watered weekly with 0.1 strength Hoagland's
solution No. 1 supplemented with micronutrients (Hoagland
and Arnon, 1950) so that the soil water potential in the root
zone was always above —0.4 MPa, as measured with PT51-O5
thermocouple psychrometers (Wescor, Logan, Utah).
For the doubled CO 2 treatment, the CO 2 concentration was
controlled with the continual release of 100% CO 2 into one of
the environmental chambers, regulated by a mass-flow meter
attached to a needle valve. The CO 2 concentration averaged
750fimol CO 2 mol" 1 air for the doubled CO 2 treatment and
370 ^mol m o P 1 for the current CO 2 treatment, monitored daily
using a Li-Cor LI-6262 infra-red gas analyser. A bank of
fluorescent and incandescent lamps with adjustable height was
used to vary the instantaneous PPF in the plane of the leaves,
as measured with a Li-Cor LI-190S Quantum Sensor. Plants
were allowed to acclimate for 10-14 d to changes in light or
temperature before gas exchange measurements were made.
Drought commenced with the withholding of the weekly
watering. Plants and CO 2 concentrations were switched between
chambers every 14 d, and plants were positioned randomly in
the chambers.
Gas exchange measurements
Net CO 2 uptake and water loss for plants in both CO 2
treatments were simultaneously measured over 24 h periods
with the LI-6262 infra-red gas analyser in a differential mode;
CO 2 measurements were corrected for dilution effects caused
by the flux of water vapour. One leaf from the mid-canopy of
each of three plants in both CO 2 treatments was inserted into
a cylindrical transparent acrylic cuvette 300 mm long and
50 mm in diameter, which was lined with transparent teflon to
reduce water vapour adsorption; all six cuvettes were sealed to
the leaf bases with plastic putty. Air from each environmental
chamber was pumped separately to two 0.020 m 3 mixing vessels
and then to the three cuvettes in the respective chambers, with
flow rates averaging 50 x 10~6 m 3 s ~ ' measured using LFC-3
mass-flow meters (Technology Incorporated, Dayton, Ohio)
and the air dewpoint with a 1211H remote optical dewpoint sensor (General Eastern Instruments, Watertown,
Massachusetts). Time-controlled solenoid valves allowed the
infra-red gas analyser to sample air exiting each cuvette.
Copper-constantan thermocouples (0.2 mm in diameter) were
attached to the adaxial and abaxial surfaces of the leaves to
determine leaf temperature; air in each cuvette was stirred with
a Li-Cor LI-6200 cuvette fan.
Data from the infra-red gas analyser, mass-flow meters,
dewpoint sensor, and thermocouples were collected every 10 min
by a Polycorder 516-24 data logger (Omnidata International,
Logan, Utah) and were transferred daily to a microcomputer.
Gas exchange measurements for responses to PPF, temperature,
and drought were made after 12 months exposure to the current
or the doubled CO 2 concentration and are expressed on the
basis of the total leaf area in the cuvette. Leaf area was
separately measured for the adaxial and abaxial leaf surfaces
using contact paper placed on the leaf; the adaxial surfaces
averaged 26% less in area than the abaxial surfaces. Daily net
CO 2 uptake refers to values integrated over a 24 h period.
Rubisco and PEPCase activities
Samples for enzyme assays were removed using a cork borer
7.7 mm in diameter from the middle of leaves approximately
the same age as those used for gas exchange measurements
after 12 months under the current and the doubled CO 2
concentrations. The samples for the assay of Rubisco activity
were removed 2 b after daytime began and those for PEPCase
activity 2 h after night-time began. The chlorenchyma of the
adaxial side of the cores was separated from the water-storage
parenchyma with a razor blade, wrapped in aluminium foil.
Doubled CO2 and Agave
and immediately frozen in liquid nitrogen. Rubisco and
PEPCase activities were determined using 14CO2 fixed into
stable products (Israel and Nobel, 1994).
Frozen samples for total Rubisco activity were ground in
2.0 cm3 of 200 mM TRIS-HC1 (pH 8.0), 10 mM MgCl2, 2%
(v/v) polyvinylpyrrolidone, 10 mM L-ascorbic acid, 5 mM
dithiothreitol (DTT), and 10 mM NaHCO3. Subsamples of
0.1 cm3 were pre-incubated for 10 min at 30 °C in 0.4 cm3 of
50 mM TRIS-HC1 (pH 8.0), 10 mM MgCl2, and 5 mM DTT.
The reaction was initiated by adding 0.025 cm3 of 11 mM
ribulose-l,5-bisphosphate and 0.025 cm3 of 450 mM NaH14CO3
and was terminated after 2 min by adding 0.1 cm3 of 6 N HC1.
The activity of activated Rubisco, which indicates the amount
of enzyme in the active state in vivo (Vu et al., 1986), was
determined by a similar procedure, except that MgCl2 and
NaHCO3 were not added to either the grinding or the incubation
solutions and the pre-incubation period was eliminated.
PEPCase activity was determined by a procedure similar to
that for total Rubisco activity except that 200 mM HEPES (N[2-hydroxyethyl ] piperazine-JV'-[2-ethanesulphonic acid])-KOH
(pH 7.2) replaced the TRIS-HC1 and 0.2% (v/v) bovine serum
albumin was added to the grinding solution. The incubation
solution contained 100 mM HEPES (pH 8.0), 10 mM MgCl2,
6 units of malate dehydrogenase, 25 ^M NADH, and 10 mM
NaH14CO3. The reaction at 25 °C was initiated by adding 1 mM
phosphoenolpyruvate and was terminated after 2 min by adding
0.1 cm3 of 6 N HC1 (Israel and Nobel, 1994).
Morphological measurements
Tissue samples from mid-leaf of leaves at mid-canopy were
removed with the cork borer, and the thickness of the adaxial
chlorenchyma was determined with an ocular micrometer
attached to a dissecting microscope capable of resolving
0.04 mm. The amount of epicuticular wax was determined from
15 discs of epidermis per plant from newly unfolded leaves
(removed with the cork borer), which were immersed for 15 s
in 15 cm3 of chloroform (Jordan et al., 1984) in pre-weighed
aluminium dishes. Chloroform was evaporated under a partial
vacuum, and the mass of the desiccated sample minus the
mass of the residue left by 15 cm3 of evaporated chloroform
was defined as the mass of the wax (Chiu et al., 1992). Leaf
thickness was determined with a fine-gauge calibrated wire
inserted through two leaves per plant mid-way along the leaf
length at one-third leaf width in from both sides. After 17
months under the current and the doubled CO2 concentrations,
dry mass of the entire root systems, leaf cores, and stem cores
was determined after drying samples at 80 °C in a forceddraught oven until no further mass change occurred (approximately 3d). Data were analysed using Student's f-test.
63
and transmittance were measured from 350-1100 nm in 2 nm
increments.
Results
Gas exchange and enzyme activities
Net CO2 uptake and transpiration for Agave deserti
occurred mainly at night (Fig. 1). For plants under the
doubled CO2 concentration, daily net CO2 uptake was
about 50% greater and the maximal instantaneous net
CO2 uptake rate was about 38% greater than for plants
under the current CO2 concentration (P<0.05 in both
cases). For plants under the current CO2 concentration,
17% of the daily net CO2 uptake occurred during the
daytime compared to 24% for plants under the doubled
CO2 concentration (P<0.05; Fig. 1A). Daily transpiration was 24% less for plants under the doubled compared
with the current CO2 concentration, with nearly all water
loss occurring at night (P<0.05; Fig. IB). The daily
water-use efficiency averaged 0.020 + 0.005 (mean + s.e.
for n = 6 plants) mol CO2 mol"1 H2O for plants under
the current CO2 concentration and 0.042 + 0.003 mol CO2
mol ~ * H2O for plants under the doubled CO2 concentration (P< 0.05).
For plants in both CO2 treatments, daily net CO2
uptake decreased steadily for 17 months (the slopes of
3 '3 I
e, S
O
_
U
o
-
B
a
o
u
3
•o
Chlorophyll content, reflectance, and transmittance measurements
Using samples from mid-leaf obtained with the cork borer 4 h
after daytime began, chlorophyll was extracted from the
chlorenchyma of both leaf sides with 5 cm3 of N, jV-dimethylformamide at 4°C for 48 h (Moran, 1982); absorptances at 647 nm
and 664 nm were measured using a DU-64 spectrophotometer
(Beckman Instruments, Fullerton, California). For reflectance
and transmittance measurements, a leaf that had unfolded
during the course of the experiment was excised from each
plant and a cuboid section approximately 20 mm each side was
removed from mid-leaf. The cuticle, epidermis, and accompanying hypodermis were peeled from the adaxial surface of the
section and placed in a Li-Cor LI-1800-12 integrating sphere
connected to a Li-Cor LI-1800 spectroradiometer. Reflectance
c
o
o
o
«
X
Fig. 1. Daily time-course of the net CO2 uptake rate (A) and the leaf
water vapour conductance (B) for leaves of Agave deserti under the
current (370 ^mol mol"1; O) and the doubled (750^mol mol"1; A)
CO2 concentrations. Gas exchange was measured after three months of
exposure. Data are means ±s.e. (n = 6 plants).
64
Graham and Nobel
the regression lines were negative, P<0.05; Fig. 2). At
the beginning of the experiment, daily net CO2 uptake
was 146+13 (mean + s.e. for n = \2 plants) mmol CO2
m~2 d" 1 for plants under the current CO2 concentration,
decreasing 38+4 mmol CO2 m ~ 2 d ~' (n = 6 plants) by 17
months; for plants under the doubled CO2 concentration,
daily net CO2 uptake was 206+ 10 mmol m " 2 d " ' at 3
months, decreasing 30+ 4mmol CO2 m~ 2 d~ 1 by 17
months. Daily net CO2 uptake for plants under the
doubled CO2 concentration averaged 49% greater than
for plants under the current CO2 concentration for the
17 month period (P<0.01; Fig. 2).
PEPCase activity was 34% lower and total Rubisco
activity was 11% lower for plants under the doubled
compared with the current CO2 concentration (P<0.05
in both cases; Table 1). The fraction of Rubisco in the
activated state in vivo tended to be greater for plants
under the doubled CO2 concentration. The ratio of
Rubisco in the activated state in vivo to total Rubisco
was 25% greater for plants under the doubled compared
with the current CO2 concentration (Table 1).
concentrations (Fig. 3). As PPF was increased from 5 to
25 mol m " 2 d - 1 , daily net CO2 uptake increased by
95 + 12 (mean+_s.e. for « = 6 plants) mmol m~ 2 d"' for
plants under the current CO2 concentration and by
144 + 21 mmol m""2 d ~l for plants under the doubled CO2
concentration. Plants under the doubled CO2 concentration had 44% greater uptake at 10 mol m" 2 d" 1 and 38%
greater uptake at 25 mol m~2 d"1 than plants under the
current CO2 concentration (P<0.05 in both cases; Fig. 3).
Daily net CO2 uptake was maximal for plants under
both the current and the doubled CO2 concentrations
near day/night temperatures of 17/12°C (Fig. 4). Plants
under the doubled CO2 concentration had 53% greater
CO2 uptake at 17/12 °C than plants under the current
CO2 concentration (P<0.05). As day/night temperatures
i
250
"
200
A"
o
B 150
Net CO2 uptake under various conditions
Daily net CO2 uptake increased with increasing PPF for
plants under both the current and the doubled CO2
O 370
^ 750
ID
M
B
..A-"" ^_s,
100
-tJ
a,
a
IS
2
-
50
0
jY
10
15
20
25
PPF (mol m" 2 d
Fig. 3. Influence of total daily PPF on daily net CO2 uptake for plants
under the current (370fimol mol"1, O) and the doubled (750^mol
mol"1; A) CO2 concentrations. The indicated PPF was for the adaxial
leaf surface, the abaxial surface receiving an average of 61% as much.
Data are means±s.e. (n = 6 plants).
5
10
Time (months)
15
Fig. 2. Monthly time-course of daily net CO2 uptake for A deserti
under the current (370/tmol mol"1; O) and the doubled (750^mol
mol"1: A) CO2 concentrations. Data are means±s.e. (n = 6 plants).
Table 1. Carboxylation enzyme activites of Agave deserti after
12 months under the current CO2 and doubled CO2 concentrations
Data are means±s.e. (n = 6 plants), where * indicates P<0.05.
Carboxylation enzyme activity
r's-)
PEPCase
RuBisCO-total
RuBisCO-activated
Activated:total for RuBisCO
CO, concentration (/xmol mol"1)
—
370
750
55.2±7.1
39.5 ±1.3
22.9±2.1
0.59 ±0.05
36 6±2.2*
35.2± 1.2*
26.3 ±1.0
0.74±0.04*
20/15
30/25
Day/night temperatures (°C)
40/35
Fig. 4. Influence of day/night air temperatures on daily net CO2 uptake
for plants under the current (370 ^mol mol"1; O) and the doubled
(750fimol mol"1: A) CO2 concentrations Leaf temperatures averaged
3 °C above air temperatures during the daytime and 1 X below at
night. Data are means±s.e. (n = 6 plants).
Doubled C02 and Agave
were increased from 17/12 °C to 40/35 °C, daily net
CO2 uptake decreased by 61±9mmol m " 2 d ' 1 for
plants under the current CO2 concentration and by
114+18mmol m~ 2 d" 1 for plants under the doubled
CO2 concentration (Fig. 4). Daytime CO2 uptake also
decreased at higher temperatures; when plants were
shifted from day/night temperatures of 17/12 °C to
40/35 °C, daytime CO2 uptake decreased from 17% to 7%
of daily net CO2 uptake for plants under the current CO2
concentration and from 24% to 17% under the doubled
CO2 concentration.
At 14 d of drought, daily net CO2 uptake for plants
under both the current and the doubled CO2 concentrations decreased from the well-watered conditions
(i><0.05; Fig. 5); plants under the doubled CO2 concentration then had 56% greater daily net CO2 uptake than
plants under the current CO2 concentration (P<0.05).
At 28 d of drought, daily net CO2 uptake had decreased
84% for plants under the current CO2 concentration and
64% for plants under the doubled CO2 concentration
(Fig. 5), with daytime CO2 uptake averaging 9% of daily
net CO2 uptake in both cases. At 28 d of drought, daily
net CO2 uptake was 3-fold greater for plants under the
doubled compared with the current CO2 concentration
CP<0.05). A 50% reduction in daily net CO2 uptake from
the well-watered conditions occurred at about 19 d for
plants under the current CO2 concentration and 23 d for
plants under the doubled CO2 concentration (Fig. 5).
Growth and cellular properties
The number of newly unfolded leaves per plant increased
steadily over 17 months for both CO2 concentrations
(Fig. 6). At 12 months, 27% more leaves had unfolded
for plants under the doubled compared with the current
CO2 concentration (P<0.05). Also at 17 months, 27%
65
15
10
Time (months)
Fig. 6. Monthly time-course for the cumulative total of new leaves
unfolding under the current (370/xmol mol"1; O) and the doubled
(750 ^mol mol"1; A) CO2 concentrations. Data are means±s.e. (n =
12 plants).
more had unfolded, leading to 7.1 new leaves per plant
under the current CO2 concentration and 9.0 under the
doubled CO2 concentration (P<0.01; Fig. 6).
At 17 months, plants had accumulated 88% more dry
mass under the doubled (311 ±29 g) compared with
the current (166±24g) CO2 concentration (P<0.05;
Table 2). The root-to-shoot ratio was about 0.12 in both
cases. Individual leaves that had unfolded on plants under
the doubled CO2 concentration had 24% greater dry
mass, were 17% longer, had similar widths, and were 11%
thicker than for those on plants under the current CO2
concentration. At 17 months, total leaf area per plant
had increased by 0.165 + 0.011 m2 for plants under the
current CO2 concentration and 0.288 + 0.016 m2 for plants
under the doubled CO2 concentration (P<0.0l; Table 2).
The chlorenchyma was 19% thicker for plants under
the doubled compared with the current CO2 concentration
(Table 3). Total chlorophyll per unit leaf area was 20%
less for the higher CO2 concentration. The chlorophyll
Table 2. Dry mass and leaf morphological properties of A. deserti
after 17 months under current and doubled CO2 concentrations
The estimated initial dry mass per plant was 275±20 g (based on leaf
and stem dry masses) and the measured initial total leaf area (adaxial
plus abaxial surfaces) per plant was 0.249 ±0.021 m2. Data are
means±s.e. (« = 6 plants), where * and ** indicate /J<0.05 and
/><0.01, respectively.
Property
10
15
20
Drought (d)
25
30
Fig. 5. Influence of drought duration on daily net CO2 uptake for
plants under the current (370^mol mol"1; O) and the doubled
(750jimol mol"1; A) CO2 concentrations. The soil water potential in
the root zone was —1.1 MPa after 1 week of drought and below
— 6 MPa after 2 weeks. Data are means±s.e. (n = 6 plants).
Shoot mass (g)
Root mass (g)
Mass per leaf (g)
Leaf length (mm)
Leaf width (mm)
Leaf thickness (mm)
Plant leaf area (m2)
CO2 concentration (^mol mol ')
370
750
395 ±30
52 ±14
22.3 + 2.0
241 ±12
39.2±2.5
6.6±0.2
0.408 ±0.051
523 + 49*
61±11
27.7±1.2*
281 ±10*
39.2 ±2.1
7.3 ±0.3*
0.543 ±0.026**
66
Graham and Nobel
Table 3. Chlorenchyma thickness, chlorophyll, and epicuticular
wax of A. derserti at 12 months under current and doubled CO2
concentrations
Data are means±s.e. (n = 6 plants), where * indicates P<0.05.
CO2 concentration (/imol mol"1)
Property
Chlorenchyma thickness (mm)
Total chlorophyll (gm~ 2 )
Chlorophyll ajb
Epicuticular wax (gm~ 2 )
370
750
1.39 ±0.08
0.80 + 0.03
2.33±0.07
1.36±0.14
1.65 + 0.08*
0.64 ±0.02*
2.55 ±0.05*
0 83 + 0.10*
ajb ratio was 9% greater for plants under the doubled
compared with the current CO2 concentration (Table 3).
Epicuticular wax averaged 64% higher for plants under
the current compared with the doubled CO2 concentration
(Table 3).
The transmittance and reflectance of the cuticle, with
accompanying epidermis and hypodermis, was lowest
from 350 to 500 run; above 500 nm, transmittance
increased and reflectance decreased with increasing wavelength (Fig. 7). From 400 to 700 nm, transmittance
through the cuticle and epidermis was 6% greater and
reflectance was 14% less for plants under the doubled
compared with the current CO2 concentration (P<0.05
in both cases), resulting in a similar absorptance
(1 — transmittance — reflectance) of 0.099 for plants under
both CO2 concentrations (Fig. 7).
Discussion
Doubling the atmospheric CO2 concentration increased
daily net CO2 uptake for Agave deserti by about 50%
throughout 17 months of exposure. Concurrent with the
increased daily net CO2 uptake was a decrease in noc-
- 370
- 750
0.8 -
Transmittance
0.6 •a
a
o
0.4
o
o
a
o
0.2
-
/
Reflectance
—S
gg
-
o n
400
600
800
Wavelength (nm)
1000
Fig. 7. Reflectance and transmittance of the epidermis with attached
cuticle and hypodermis from plants under the current (370ftmol
mol"1;
) and the doubled (750 ^mol mol"1;
) CO2 concentrations. Data are means±s.e. at 100 nm intervals (n = 6 plants).
turnal water vapour conductance, presumably by partial
stomatal closure, leading to a doubled water-use efficiency. A higher water-use efficiency can lead to a greater
plant water content, allowing a greater daytime net CO2
uptake using the C3 pathway for A. deserti under the
doubled CO2 concentration. Increased CO2 concentrations have also increased the water-use efficiency for C3
plants (Eamus, 1991; Ryle et al., 1992, Tschaplinski et al.,
1993) and the CAM plant Opuntiaficus-indica(Cui et al.,
1993). In this regard, increases in water-use efficiency can
be particularly important for plants, such as A. deserti,
that are native to areas where water availability frequently
limits assimilation. The small decrease in daily net CO2
uptake over the 17 month period for A. deserti under
both CO2 concentrations possibly reflected the limited
soil volume in which the plants were grown, as also
occurs for other species (Ceulemans and Mousseau, 1994;
Nobel et al., 19946).
Long-term enhancement of net CO2 uptake by doubling the atmospheric CO2 concentration also occurs for
other CAM species (Nobel and Hartsock, 1986; Nobel
and Israel, 1994), but is often not the case for C3
species (DeLucia et al., 1985; Ehret and Jolliffe, 1985).
Acclimation of C3 and CAM species to increased CO2
concentration can occur by the down-regulation of
Rubisco in response to the increased efficiency of carboxylation by Rubisco at higher CCVOj ratios (Campbell
et al., 1988; Bowes, 1993; Israel and Nobel, 1994). When
the CO2 concentration was doubled for A. deserti, the
total activity of Rubisco decreased, but daily net CO2
uptake increased, as a larger fraction of Rubisco was
then in the activated state in vivo. CO2 is an activator as
well as a substrate for Rubisco (Lorimer et al., 1976), so
increased internal CO2 concentrations could lead to more
active Rubisco in vivo. In this regard, after 3 months
exposure the internal CO2 concentration based on leaf
water vapour conductance and net CO2 uptake measured
in the middle of the dark period was 180/nmol moP 1
under the current CO2 concentration and 410 ^mol moP 1
under the doubled CO2 concentration. Doubling the CO2
concentration also reduced the activity of PEPCase
for A. deserti, consistent with the increased internal
CO2 concentration reducing the required amount of this
enzyme.
Chlorophyll per unit leaf area decreased for A. deserti
under the doubled CO2 concentration, as for other species,
which suggests an increased photosynthetic efficiency
(Szarek et al., 1987; Wullschleger et al., 1992). Indeed,
the chlorophyll a/b ratio increased for A. deserti under
the doubled CO2 concentration, indicating a relative
decrease of chlorophyll in the light-harvesting complexes.
Chlorenchyma thickness increased when the atmospheric
CO2 concentration was doubled, which may be related to
a higher CO2 concentration deeper in the leaf (Powles
et al., 1980) or to greater PPF penetration because of the
Doubled CO2 and Agave
reduction in chlorophyll. In addition, less epicuticular
wax accumulated on newly unfolded leaves of A. deserti
under the doubled CO2 concentration, consistent with the
reduced reflectance of PPF by their cuticles and possibly
correlated with an increased leaf water content (Jordan
et al., 1983). The accompanying increase in transmittance
allows a greater PPF to reach the photosynthetic cells
under the doubled CO2 concentration, influencing the
tissue depth at which chlorophyll occurs. Similar reductions in chlorophyll content and increases in chlorenchyma thickness occur after exposure of O. ficus-indica
to a doubled CO2 concentration (Nobel et al., 1994a).
More leaves of A. deserti unfolded per plant, and
unfolded leaves were longer and thicker under the doubled
CO2 concentration. The resulting greater plant leaf surface area coupled with the 50% increase in daily net CO2
uptake per unit leaf area both contributed to the greater
CO2 uptake per plant under the doubled CO2 concentration. Because the leaf water vapour conductance was
lower under the doubled CO2 concentration, plant water
loss was relatively unaffected. At 17 months, plants under
the doubled CO2 concentration had accumulated 88%
more dry mass than those under the current CO2 concentration. Developing photosynthetic stems of O. ficusindica also thicken after exposure to a doubled CO2
concentration and show a 37—40% increase in aboveground biomass after 12 months (Nobel and Israel, 1994);
a 30—40% increase in growth likewise occurs for various
C3 crops exposed to a doubled CO2 concentration (Cure
and Acock, 1986; Rogers et al., 1986). The combined
effects by the doubled atmospheric CO2 concentration of
increased daily net CO2 uptake and increased leaf surface
area per plant resulted in the relatively large stimulation
of biomass production for A. deserti.
Daily net CO2 uptake under controlled conditions in
the laboratory can be used to calculate an Environmental
Productivity Index for predicting biomass production in
the field (Nobel, 1988). Component indices relating CO2
uptake to PPF, temperature, and water availability have
been determined for A. deserti under the current CO2
concentration (Nobel, 1984). The responses of this species
to these environmental variables were slightly different
under the doubled CO2 concentration. Specifically, the
increase in daily net CO2 uptake then occurred only
above a PPF of 5 mol m~2 d"1, below which PPF was
most likely the limiting factor for CO2 uptake. Daily net
CO2 uptake was maximal near day/night temperatures of
17/12 CC for A. deserti under both the current and the
doubled CO2 concentrations, reflecting the optimal nighttime temperature for carboxylation by PEPCase (Winter,
1985). As drought length increased for A. deserti, the per
cent enhancement of daily net CO2 uptake by doubling
the CO2 concentration increased; daily net CO2 uptake
decreased to half-maximal for plants under the doubled
CO2 concentration 4 d after those under the current CO2
67
concentration, possibly because of the increased wateruse efficiency and thus higher plant water content under
the doubled CO2 concentration. The effects of PPF,
temperature, and drought length on daily net CO2 uptake
by O.ficus-indicaunder a doubled CO2 concentration are
similar, with proportionally lower reductions of daily net
CO2 uptake occurring under the doubled CO2 concentration at the extremes of PPF and temperature and
increased drought length (Nobel and Israel, 1994).
General circulation models predict that global temperatures will rise from 2.8 °C to 6.3 °C and precipitation
will increase from 8% to 15% with a doubling of the
atmospheric CO2 concentration (Smith et al., 1992; Cao
et al., 1992). Although the resolution for such predicted
climate change is coarse, and local topography will influence climate effects, the Sonoran Desert may receive an
increase in precipitation, changing the climate zone from
desert to subtropical savanna (Smith et al., 1992). Such
an increase in precipitation will reduce the amount of
time that daily net CO2 uptake is limited by water
availability, although increased cloud cover will reduce
PPF. An increase in average air temperature of 4 °C in
the Sonoran Desert will raise the average minimum air
temperature to about 24 °C in the summer, 20 °C in the
spring and autumn, and 13°C in the winter (Nobel,
1988); this will result in about a 15% decrease in daily
net CO2 uptake per leaf area of A. deserti during the
summer, less than a 10% decrease in the spring and
autumn, and very little change during the winter.
Assuming that the increase in precipitation more than
offsets the associated decrease in PPF, and taking the
seasonal temperature effects into consideration, average
annual net CO2 uptake per leaf area should then be about
40% gTeater under a doubled compared with the current
CO2 concentration. This enhanced net CO2 uptake will
be compounded by an increase in leaf area, further
increasing growth. Carbon gain under a doubled atmospheric CO2 concentration and higher temperatures would
then considerably exceed that for lower temperatures
under the current CO2 concentration, resulting in
increased productivity and a possible competitive advantage for native populations of A. deserti.
Acknowledgements
We thank Dr Alvaro A. Israel for his assistance with enzyme
assays. This research was supported by the Environmental
Sciences Division, Office of Health and Environmental Research,
US Department of Energy, Carbon Dioxide Research Program
grant DE-FG03-ER61252 and Program for Ecosystem Research
grant DE-FGO3-93ER61686.
References
Bowes G. 1991. Growth at elevated CO 2 : photosynthetic
responses mediated through Rubisco. Plant, Cell and
Environment 14, 795-806.
68
Graham and Nobel
Bowes G. 1993. Facing the inevitable: plants and increasing
atmospheric CO 2 . Annual Review of Plant Physiology and
Plant Molecular Biology 44, 309-32.
Campbell WJ, Allen Jr LH, Bowes G. 1988. Effects of CO 2
concentration on Rubisco activity, amount, and photosynthesis in soybean leaves. Plant Physiology 88, 1310-16.
Cao HX, Mitchell JFB, Lavery JR. 1992. Simulated diurnal
range and variability of surface temperature in a global
climate model for present and doubled CO 2 climates. Journal
of Climate 5, 920-43.
Ceulemans R, Mousseau M. 1994. Tansley Review No. 71.
Effects of elevated atmospheric CO 2 on woody plants. jVeii'
Phytologist 127, 425-46.
Chiu ST, Anton LH, Ewers FW, Hammerschmidt R, Pregitzer
KS. 1992. Effects of fertilization on epicuticular wax morphology of needle leaves of Douglas fir, Pseudotsuga menziesii
(Pinaceae). American Journal of Botany 79, 149-54.
Cui M, Miller PM, Nobel PS. 1993. CO 2 exchange and growth
of the Crassulacean acid metabolism plant Opuntia ficusindica under elevated CO 2 in open-top chambers. Plant
Physiology 103, 519-24.
Cui M, Nobel PS. 1994. Gas exchange and growth responses
to elevated CO 2 and light levels in the CAM species Opuntia
ficus-indica. Plant, Cell and Environment 17, 935-44.
Cure JD, Acock B. 1986. Crop responses to carbon dioxide
doubling: a literature survey. Agricultural and Forest
Meteorology 38, 127-45.
DcLucia EH, Sasek TW, Strain BR. 1985. Photosynthetic
inhibition after long-term exposure to elevated levels of
atmospheric carbon dioxide. Photosynthesis Research 7,
175-84.
Eamus D. 1991. The interaction of rising CO 2 and temperatures
with water use efficiency. Plant, Cell and Environment
14, 843-52.
Ehret DL, Jolliffe PA. 1985. Photosynthetic carbon dioxide
exchange of bean plants grown at elevated carbon dioxide
concentrations. Canadian Journal of Botany 63, 2026-30.
Gentry HS. 1982. Agaves of continental North America. Tucson:
The University of Arizona Press.
Hoagland DR, Amon DI. 1950. The water-culture method for
growing plants without soil. California Agricultural
Experimental Station Circular 347, 1-32.
Holrum JAM, O'Leary MH, Osmond CB. 1983. Effect of
varying CO 2 partial pressure on photosynthesis and on
carbon isotope composition of carbon-4 of malate from the
Crassulacean acid metabolism plant KalanchoS daigremontiana Hamet et Perr. Plant Physiology 71, 602-9.
Huerta AJ, Ting IP. 1988. Effects of various levels of CO 2 on
the induction of Crassulacean acid metabolism in Portulacaria
afra (L.) Jacq. Plant Physiology 88, 183-8.
Hunt R, Hand DW, Hannah MA, Neal AM. 1991. Response to
CO 2 enrichment in 27 herbaceous species. Functional Ecology
5,410-21.
Idso SB, Kimball BA. 1992. Above-ground inventory of sour
orange trees exposed to different atmospheric CO 2 concentrations for three full years. Agricultural and Forest Meteorology
60, 145-51.
Israel AA, Nobel PS. 1994. Activities of carboxylating enzymes
in the CAM species Opuntia ficus-indica grown under current
and elevated CO 2 concentrations. Photosynthesis Research
40, 223-9.
Jordan WR, Monk RL, Miller FR, Roseoow DT, Clark LE,
Shouse PJ. 1983. Environmental physiology of sorghum. I.
Environmental and genetic control of epicuticular wax load.
Crop Science 23, 552-8.
Jordan WR, Shouse PJ, Blum A, Miller FR, Monk RL, 1984.
Environmental physiology of sorghum. II. Epicuticular wax
load and cuticular transpiration. Crop Science 24, 1168-73.
King AW, Emanuel WR, Post WM. 1992. Projecting future
concentrations of atmospheric CO 2 with global carbon cycle
models: the importance of simulating historical changes.
Environmental Management 16, 91-108.
Lawlor DW, Mitchell RAC. 1991. The effects of increasing CO 2
on crop photosynthesis and productivity: a review of field
studies. Plant, Cell and Environment 14, 807-18.
Lorimer GH, Badger MR, Andrews TJ. 1976. The activation of
ribulose-l,5-bisphosphate carboxylase by carbon dioxide and
magnesium ions. Equilibria, kinetics, a suggested mechanism,
and physiological implications. Biochemistry 15, 529-36.
Moran R. 1982. Formulae for determination of chlorophyllous
pigments extracted with jV,./V-dimethylfbrmamide. Plant
Physiology 69, 1376-81.
Nobel PS. 1976. Water relations and photosynthesis of a desert
CAM plant, Agave deserti. Plant Physiology 58, 576-82.
Nobel PS. 1984. Productivity of Agave deserti: measurement by
dry weight and monthly predictions using physiological
responses to environmental parameters. Oecologia 64, 1-7.
Nobel PS. 1988. Environmental biology of agaves and cacti. New
York: Cambridge University Press.
Nobel PS, Cui M, Israel AA. 1994a. Light, chlorophyll,
carboxylase activity, and CO 2 fixation at various depths in
the chlorenchyma of Opuntia ficus-indica (L.) Miller under
current and elevated CO 2 . New Phytologist 128, 315-22.
Nobel PS, Cui M, Miller PM, Luo Y. 19946. Influences of soil
volume and an elevated CO 2 level on growth and CO 2
exchange for the Crassulacean acid metabolism plant Opuntia
ficus-indica. Physiologia Plantarum 90, 173-80.
Nobel PS, Hartsock TL. 1986. Short-term and long-term
responses of Crassulacean acid metabolism plants to elevated
CO2. Plant Physiology 82, 604-6.
Nobel PS, Israel AA. 1994. Cladode development, environmental
responses of CO 2 uptake, and productivity for Opuntia ficusindica under elevated CO 2 . Journal of Experimental Botany
45, 295-303.
Powles SB, Chapman KSR, Osmond CB. 1980. Photoinhibition
of intact attached leaves of C 4 plants: dependence on CO 2
and O 2 partial pressures. Australian Journal of Plant
Physiology 7, 737-47.
Rogers HH, Cure JD, Smith JM. 1986. Soybean growth and
yield response to elevated carbon dioxide. Agriculture
Ecosystems and Environment 16, 113-28.
Rogers HH, Dahlman RC. 1993. Crop responses to CO 2
enrichment. Vegetatio 104, 117-31.
Ryle GJA, Woledge J, Tewson V, Powell CE. 1992. Influence of
elevated CO 2 and temperature on the photosynthesis and
respiration of white clover dependent on N 2 fixation. Annals
of Botany 70, 213-20.
Sage RF, Sharkey TD, Seemann JR. 1989. Acclimation of
photosynthesis to elevated CO 2 in five C 3 species. Plant
Physiology 89, 590-6.
Smith TM, Leeraans R, Shugart HH. 1992. Sensitivity of
terrestrial carbon storage to CO2-induced climate change:
comparison of four scenarios based on general circulation
models. Climatic Change 21, 367-84.
Stitt M. 1991. Rising CO 2 levels and their potential significance
for carbon flow in photosynthetic cells. Plant, Cell and
Environment 14, 741-62.
Szarek SR, Holthe PA, Ting IP. 1987. Minor physiological
response to elevated CO 2 by the CAM plant Agave vilmoriniana. Plant Physiology 83, 938-40.
Tschaplinski TJ, Norby RJ, Wulbchleger SD. 1993. Responses
Doubled CO2 and Agave
of loblolly pine seedlings to elevated CO 2 and fluctuating
water supply. Tree Physiology 13, 283-96.
Vu JCV, Bowes G, Allen Jr JH. 1986. Properties of
ribulose-l,5-bisphosphate carboxylase from dark- and lightexposed soybean leaves. Plant Science 44, 119-23.
Winter K. 1985. Crassulacean acid metabolism. In: Barber J,
Baker NR, eds. Photosynthetic mechanisms and the environment, Vol. 6. Amsterdam: Elsevier, 329-87.
Wullschleger SD, Norby RJ, Hendrix DL. 1992. Carbon
exchange rates, chlorophyll content, and carbohydrate status
69
of two forest tree species exposed to carbon dioxide
enrichment. Tree Phvsiologv 10, 21-31.
Yelle S, Beeson Jr RC, Tnidel MJ, Gosselin A. 1989. Acclimation
of two tomato species to high atmospheric CO 2 . II.
Ribulose-l,5-bisphosphate carboxylase/oxygenase and phosphoenolpyruvate carboxylase. Plant Phvsiologv 90, 1473-7.
Ziska LH, Hogan KP, Smith AP, Drake BG. 1991. Growth and
photosynthetic response of nine tropical species with longterm exposure to elevated carbon dioxide. Oecologia 86,
383-9.