Tree Physiology 28, 1831–1840
© 2008 Heron Publishing—Victoria, Canada
Close coupling of whole-plant respiration to net photosynthesis and
carbohydrates
TIMOTHY M. WERTIN1,2 and ROBERT O. TESKEY1
1
Warnell School of Forestry and Natural Resources, University of Georgia, Athens, GA 30602, USA
2
Corresponding author ([email protected])
Received May 20, 2008; accepted August 22, 2008; published online October 1, 2008
Summary We studied the effect of changes in net photosynthesis (Anet ) on respiration, soluble sugars and carbohydrates in
Populus deltoides Bartr. ex Marsh. saplings under controlled
environmental conditions by making daily measurements of
leaf respiration (Rd ), stem CO2 efflux and root CO2 efflux at a
constant temperature in growth chambers. After a pretreatment
period, one of three treatments was applied for 5 to 7 days: (1)
increased atmospheric CO2 concentration; (2) decreased
photoperiod and photosynthetic photon flux (PPF); or (3) continuous darkness. Increased Anet in response to elevated CO2
concentration resulted in a sustained increase in whole-plant
respiration, with Rd increasing 46%, stem CO2 efflux increasing 130% and root CO2 efflux increasing 16%. Elevated CO2
concentration also caused a significant increase in leaf soluble
sugars. Decreasing photoperiod and PPF or complete darkness
caused a rapid decrease in respiration throughout the saplings.
In the low light treatment, Rd decreased 40%, stem CO2 efflux
decreased 78%, root CO2 efflux decreased 74% and significant
decreases in leaf and root soluble sugar and leaf nonstructural
carbohydrate concentrations were observed. Continuous darkness resulted in a 70% decrease in Rd, a 65% decrease in stem
CO2 efflux, a 73% decrease in root CO2 efflux and significant
decreases in leaf and root soluble sugar and root carbohydrate
concentrations. In all treatments, changes in respiration rates in
all tissues occurred within hours of treatment application. In
addition, a diurnal pattern in root CO2 efflux was observed
throughout the experiment under constant environmental conditions. The observed rapid changes in whole-plant respiration
following treatment application and the diurnal patterns in root
CO2 efflux suggest that growth and maintenance respiration in
the saplings was strongly dependent on newly acquired carbohydrates.
Keywords: diurnal pattern, leaf respiration, Populus deltoides,
root CO2 efflux, stem CO2 efflux.
Introduction
The influence that environmental factors such as temperature
have on plant respiration is well documented (Amthor 1989).
However, the relationship between net photosynthesis (Anet )
and whole-plant respiration is less understood. A positive cor-
relation between Anet and leaf respiration (Rd ) has been observed (Amthor 1989, Dewar et al. 1999, Loveys et al. 2003,
Whitehead et al. 2004). It is presumed that increases in Anet
lead to increases in leaf carbohydrate concentration, which in
turn drive increases in Rd. However, some studies have observed increases in Anet without corresponding increases in Rd
(Poorter et al. 1992, Wullschleger et al. 1994, Bunce 1995,
2005).
The relationship between Anet and stem CO2 efflux to the atmosphere has received less attention than the relationship between Anet and Rd. Stem CO2 efflux, which is a combination of
CO2 released directly to the atmosphere by respiring cells in
the stem and transported CO2 diffusing from xylem sap
(Teskey et al. 2008) may also be linked to Anet. Zha et al.
(2004) found that stem CO2 efflux in 50-year-old Pinus
sylvestris L. trees was positively related to both photosynthetically active radiation and ecosystem gross primary production,
suggesting that photosynthesis may partially regulate stem
CO2 efflux. Cernusak et al. (2006) found that decreased stem
CO2 efflux was associated with decreased canopy photosynthesis, presumably due to decreased leaf carbohydrate
synthesis and subsequent translocation.
Several tree girdling studies, designed to disrupt the phloem
tissue in the stem and thereby inhibit carbohydrate transport to
the roots, have shown that root CO2 efflux partly depends on
the supply of newly available carbohydrates. For example,
within 5 days after girdling Pinus sylvestris trees, Högberg et
al. (2001) observed a 37% reduction in soil CO2 efflux. (Soil
CO2 efflux is a combination of CO2 released from root respiration, microbial respiration, which is typically strongly dependent on root exudates, and organic matter decomposition.) Similarly, a 22 to 36% reduction in soil CO2 efflux occurred 9 days
after girdling Castanea sativa Mill. trees (Frey et al. 2006), but
a smaller 16 to 24% reduction was seen in girdled Eucalyptus
grandis Hill Ex Maiden × urophylla S.T. Blake trees (Binkley
et al. 2006). Stem girdling caused a reduction in soil CO2
efflux within 1 to 2 days in Fagus salvatica L. trees, yet it took
6 weeks to produce a response in Picea abies (L.) Karst. trees
in the same study (Andersen et al. 2005). Binkley et al. (2006)
suggested that, in some species, decreases in root respiration
rates may take months if there are large amounts of stored carbohydrate in the roots. However, few studies have specifically
1832
WERTIN AND TESKEY
examined the relationship between Anet and root CO2 efflux.
The only such study in trees of which we are aware found that
root CO2 efflux in Tsuga canadensis (L.) Carr. seedlings was
34% higher when Anet was at light saturation than when the
plants were in darkness (Szaniawski and Adams 1974).
Hansen (1977) reported a similar response in Italian ryegrass
(Lolium multiflorum Lam.).
Our objective was to investigate changes in Rd, stem CO2
efflux, root CO2 efflux and leaf and root sucrose and carbohydrate concentrations with changes in Anet in Populus deltoides
Bartr. ex Marsh. saplings under constant environmental conditions. Specifically, we assessed how quickly changes in tissue
respiration occur with changes in Anet. We also examined diurnal variation in tissue respiration and determined whether
changes in Anet affect these diurnal patterns.
Materials and methods
Plant material
Populus deltoides (OP-367; Hybrid Poplar Trees, Glenmoore,
PA) cuttings were propagated in 8-l pots filled with a 3:1
(w/w) mix of sand:Turface ProMix (Profile Produces LLC,
Buffalo Grove, IL). A rooting hormone (Hormoding 1, Hybrid
Poplar Trees, Glenmoore, PA) was applied to the cuttings to
promote rooting. Plants were watered daily and fertilized
twice weekly with Peters Excel Cal Mag (The Scotts Company, Marysville, OH). Cuttings were grown in a greenhouse
for about 6 weeks, or until they were at least 1 m in height. Before treatment initiation, saplings were transferred to growth
chambers (GC 36; Environmental Growth Chambers, Chargin
Falls, OH) and acclimated for 1 week.
Experimental design
Initial chamber conditions were 25 °C, 65% relative humidity
(RH), 375 µmol CO2 mol –1, and a 14-h photoperiod with a
photosynthetic photon flux (PPF) of 500 µmol m –2 s –1. Air
temperature and RH were kept constant throughout the experiment. After the 1-week acclimation, Anet , Rd and stem and root
CO2 efflux were measured for 3 days (pretreatment period) to
establish the pretreatment base line, and then one of three
treatments was applied for 5 to 7 days: elevated CO2 concentration (EC), low light (LL), or no light (NL). Treatment conditions were the same as the pretreatment acclimation conditions except that, in the EC treatment chamber, CO2 concentration was increased to 1500 µmol mol –1, in the LL treatment,
the light regime was reduced to 8 h of 150 µmol m –2 s –1 PPF,
and in the NL treatment, light was reduced to 0 µmol m –2 s –1
PPF. Three saplings were assigned to each treatment.
Measurements of carbohydrates
In addition to the three saplings in each treatment, at the end of
the pretreatment period, three additional saplings of comparable age, height, leaf area and photosynthetic rate that had been
subjected to the same growth and pretreatment conditions
were used to estimate initial leaf and root soluble sugar and
carbohydrate concentrations. All destructive harvests were
made at the end of the dark period. Canopies were divided into
three vertical sections of equal size and dried for 48 h at 65 °C.
Roots were well washed and dried at 65 °C for 48 h. Dried
plant material was weighed, ground with a ball mill to a fine
powder, sealed in containers and stored with desiccant.
Concentrations of soluble sugars and nonstructural carbohydrates in the roots and leaves were determined as outlined
by Haissig and Dickson (1979) and Hansen and Møller (1975).
Briefly, soluble sugars from about 50 mg of ground plant material were extracted with 10 ml of 12:5:3 (v/v) methanol:
chloroform:water; the pellet was saved for carbohydrate determination. Following reaction of the extracted soluble sugars
with 0.01% (w/v) anthrone reagent, absorbance at 630 nm was
measured. Starch in the insoluble residue was converted to
glucose with amyloglucosidase. Glucose concentration was
determined by mixing the sample with peroxidase-glucose
oxidase-o-dihydrochloride and measuring the absorbance at
540 nm after a 30-min incubation at 37 °C. Soluble sugar and
nonstructural carbohydrate concentrations were calculated
from linear regressions based on glucose standards and
expressed on a percent dry mass basis.
At the end of the treatment period, the treated saplings were
destructively harvested. Leaf area was measured with an
LI-3100 area meter (Li-Cor, Lincoln, NE). Root and leaf materials were dried, weighed and ground, and the concentrations
of root and leaf soluble sugars and nonstructural carbohydrates were determined as described previously.
Measurements of net photosynthesis and respiration
We measured Anet, Rd, stem CO2 efflux and root CO2 efflux
daily on three plants per treatment for 3 days before treatment
application and for 5 to 7 days immediately following commencement of the treatments. Each sapling canopy was divided into three equal vertical sections and a representative
leaf from each section was selected for repetitive Anet and Rd
measurements. Measurements were made with a Li-Cor
LI-6400 portable photosynthesis system with a standard
red/blue LED broadleaf cuvette and a CO2 mixer. All Anet
measurements were made on fully expanded leaves that were
illuminated for a minimum of 1 h in the growth chamber. Measurements were made under current growth chamber conditions. Two photosynthetic measurements were made per day,
1 h after initiation of the light period and 1 h before the dark
period. Leaf respiration was measured on the same leaves.
Plants were dark acclimated for at least 30 min before measurements, and all measurements were allowed to stabilize for
5 min before recording. Three measurements of Rd were made
daily, at 30 min pre-illumination, at the mid-light period, and
1 h post-darkness.
Stem CO2 efflux was measured as described by Teskey and
McGuire (2005). Briefly, a 2-cm-diameter by 6-cm-long cylindrical white PVC chamber was clamped around the stem at
about 0.2 m above the base and closed at both ends with compressed foam gaskets. Air of known CO2 concentration was
passed through the chamber at 0.5 l s –1, and efflux from the
stem was measured with a Li-Cor LI-6252 IRGA. Respiration
measurements were made three times daily for 2 h.
TREE PHYSIOLOGY VOLUME 28, 2008
CLOSE COUPLING OF WHOLE-PLANT RESPIRATION TO NET PHOTOSYNTHESIS
Root respiration, the CO2 efflux from the soil substrate, being attributable primarily to root respiration and any microbial
or fungal respiration, was measured in a similar manner to
stem respiration. Briefly, the pots, which contained the soil
substrate and roots, were placed in a bucket, with the stem extending through a hole in the lid, and sealed with foam. Air of
known CO2 concentration was continually passed through the
root chamber at 1.5 l s –1 and CO2 efflux from the roots measured with a Li-Cor LI-6252. Saplings were watered daily with
deionized water which was acidified with sulfuric acid to
pH 5.4 to minimize CO2 absorption.
1833
low the first expanding leaf. We calculated CUE for each
sapling on a daily basis:
C UE =
C DG
A
LA
DL
∑( net,n n – Rlight,n LAnDL)
(2)
n
Leaf respiration in the light (Rlight ) was scaled from Rd measurements based on the results of Brooks and Farquhar (1985)
and Lloyd et al. (1995):
Rlight = (0.5 − 0.05 ln PPF ) Rd
(3)
Statistical analysis
We expressed Anet, Rd and stem CO2 efflux on a surface area
(m 2 ) basis. Root CO2 efflux was expressed on a per gram dry
mass basis because it was impossible to measure root surface
area or root volume accurately. Daily measurements of Anet, Rd
and stem CO2 efflux were averaged by sapling, and significant
differences both among days and between the 3 days of pretreatment and the last 3 days of treatment were tested by
one-way ANOVA. Measurements of Anet, Rd and stem CO2
efflux were averaged by measurement period to investigate
potential diurnal patterns. Root respiration measurements
were averaged by sapling per 24-h period and differences both
among days and between the 3 days of pretreatment and the
last 3 days of treatment were tested.
Correlations between Anet and tissue respiration and between tissue carbohydrate status and tissue respiration were
analyzed by linear regression. For each treatment, we compared Anet values averaged for the 3 days of pretreatment and
the last 3 days of treatment with mean Rd, stem CO2 efflux and
root CO2 efflux for the same time periods; respiration measurements for the last 3 days of the NL treatment were disregarded because of a lack of corresponding Anet measurements.
Leaf and root soluble sugar and starch concentrations measured at the end of the experiment were compared with corresponding mean tissue respiration rates for the last day of
treatment.
Estimates of daily carbon gain (CDG ) and carbon-use efficiency (CUE ) were calculated for saplings in the EC and LL
treatments. For each sapling, CDG was calculated as:
C DG =
∑n ( Anet,n LAnDL – Rd,n LA n NL)
(1)
− 86, 400 SE avg SA stem − 86, 400 REavg DM root
where n is the upper, middle or lower third of the canopy, LAn
is leaf area for the corresponding portion of the canopy, DL is
duration of the light period (s), Rd,n is Rd expressed as a positive
value for the corresponding portion of the canopy, NL is duration of the dark period (s), SEavg is mean stem CO2 efflux for
that day, SAstem is the surface area of the stem, REavg is mean
root CO2 efflux for the day, and DMroot is root dry mass. Stem
surface area estimates were based on measurements of stem
height and stem diameter made at the end of the experiment.
Stem diameter was measured above the soil interface and be-
Results
Net photosynthesis and respiration
As expected, the EC treatment increased Anet, decreasing light
flux decreased Anet, and the NL treatment resulted in a negative
Anet (Figure 1). The EC treatment increased Anet by about 45%
from a mean of 10.3 µmol m –2 s –1 for the 3 days of pretreatment to a mean of 14.9 µmol m –2 s –1 for the last 3 days of treatment (P < 0.001). During the first day of EC treatment, Anet increased about 73% to 19.2 µmol m –2 s –1 and then slowly decreased until values stabilized after Day 4 of treatment. The
LL treatment decreased Anet by 34% from a mean of 9.8 µmol
m –2 s –1 for the 3 days of pretreatment to a mean of 6.5 µmol
m –2 s –1 for the last 3 days of treatment (P < 0.001). On Day 1
of the LL treatment Anet decreased by over 56% to 5.0 µmol
m –2 s –1, but it increased to 6.6 µmol m –2 s –1 on Day 2 of treatment. The NL treatment caused a 103% decrease in Anet from a
mean of 9.3 µmol m –2 s –1 for the 3 days of pretreatment to a
mean of –0.31 µmol m –2 s –1 for the last 3 days of treatment (P
< 0.001). A 106% decrease in Anet occurred on Day 1 of the NL
treatment.
Changes in Anet significantly affected Rd (Figure 2). The EC
treatment significantly increased Rd (P < 0.001) from a mean
of 1.3 µmol m –2 s –1 for the 3 days of pretreatment to a mean of
1.9 µmol m –2 s –1 for the last 3 treatment days (46% increase),
with Rd increasing 16% within the first treatment day. The LL
treatment significantly decreased Rd (P < 0.001). Comparing
the mean values of the 3 days of pretreatment with those of the
last 3 days of treatment, Rd decreased 40%, from 1.5 to
0.9 µmol m –2 s –1. A decrease of 17% was observed on Day 1 of
the LL treatment, and Rd continued to decline throughout the
treatment. The NL treatment significantly decreased Rd (P <
0.001) with means in the 3 days of pretreatment and the last
3 days of treatment declining 70% from 1.0 to 0.3 µmol m –2
s –1, with a decrease of 37% on Day 1 of the NL treatment.
The EC treatment increased stem CO2 efflux, whereas the
LL and NL treatments decreased it (Figure 3). Stem CO2
efflux increased about 130% in the EC treatment, with mean
values for the 3 days of pretreatment and the last 3 days of
treatment of 4.7 and 10.8 µmol m –2 s –1, respectively (P =
0.0181). Stem CO2 efflux increased by 10% in the first 12 h of
EC treatment and continued to increase until Day 5 of treatment. The LL treatment decreased stem CO2 efflux by about
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1834
WERTIN AND TESKEY
78% (P < 0.001). Averaging the measurements for the 3 days
of pretreatment and the last 3 days of treatment revealed a decrease in stem CO2 efflux from 6.4 to 1.4 µmol m –2 s –1 with a
decrease of 53% occurring within Day 1 of the LL treatment.
The NL treatment decreased stem CO2 efflux by 65% (P <
0.001), from a mean of 4.9 µmol m –2 s –1 for the 3 days of
pretreatment to 1.7 µmol m –2 s –1 for the last 3 days of treatment. Stem CO2 efflux decreased 21% on Day 1 of the NL
treatment and continued to decrease throughout the treatment.
Root CO2 efflux increased in response to the EC treatment
and decreased in response to the LL and NL treatments
(Figure 4). In EC saplings there was a small (16%) nonsignificant (P = 0.117) increase in root respiration from a pretreatment mean of 0.0232 to a mean of 0.0269 µmol CO2 gDM–1 s –1
in the final 3 days of treatment. The LL treatment decreased
root CO2 efflux by 74% (P < 0.001) from a mean of
25
Carbohydrates
Leaf and root soluble sugar and nonstructural carbohydrate
concentrations were affected by the treatments (Figure 5). The
EC treatment increased leaf soluble sugars by 20% (P =
0.0343), but had no significant effect on root soluble sugars or
leaf or root nonstructural carbohydrate concentration. In response to the LL treatment, leaf and root soluble sugar concen-
2.5
A. EC treatment
A
B
b
20
b
b
b/c
a
a
b/c
b/c
B
b
b
a/b
1.5
a
a/c
a
a
1.0
0.5
B. LL treatment
A
12
a
0.0
2.0
B
a
Rd (µmol m – 2 s – 1 )
Anet (µmol m – 2 s – 1 )
b
a/b a/b a/b
5
0
14
A. EC treatment
A
2.0
b
15
10
0.0408 µmol CO2 gDM–1 s –1 in the 3 days of pretreatment to a
mean of 0.0108 µmol CO2 gDM–1 s –1 during the last 3 days of
treatment (P < 0.001). Root CO2 efflux decreased 33% on Day
1 of the LL treatment. The NL treatment decreased root CO2
efflux 73% from a 3-day pretreatment mean of 0.0247 to a
mean of 0.0067 µmol CO2 gDM–1 s –1 for the final 3 days of
treatment (P < 0.001). Mean daily root CO2 efflux fell 40% on
Day 1 of the NL treatment.
10
b
8
b/c
6
b/c
b/c
b/c
b/c
c
4
B. LL treatment
A
a a/b
a
B
a/b
1.5
a/b
a/b
a/b
1.0
a/b
b
0.5
2
0
12
10
0.0
1.4 C. NL treatment
C. NL treatment
A
b
B
A
b
1.2
a/b
8
1.0
6
0.8
4
0.6
2
0.4
c
0
c
c
c
c
T3
T4
T5
a
B
a
a
b/c
b/c
b/c
c
c
T3
T4
T5
0.2
0.0
-2
P1 P2 P3
T1 T2
T6
T7
P1 P2 P3
Treatment day
T1 T2
T6
T7
Treatment day
Figure 1. Daily mean (+ SE, n = 3) net photosynthetic rate (Anet ) of
Populus deltoides saplings during the pretreatment (black bars) and
treatment (gray bars) periods in the (A) elevated CO2 (EC), (B)
low-light (LL) and (C) no-light (NL) treatments. Different uppercase
letters denote significantly different means for the 3 days of pretreatment and the last 3 days of treatment, and different lowercase letters
denote significantly different daily means (P < 0.05). P1–P3 denote
pretreatment days, and T1–T7 denote treatment days.
Figure 2. Daily mean (+ SE, n = 3) leaf respiration rate (Rd ) of Populus
deltoides saplings during the pretreatment (black bars) and treatment
(gray bars) periods in the (A) elevated CO2 (EC), (B) low-light (LL)
and (C) no-light (NL) treatments. Different uppercase letters denote
significantly different means for the 3 days of pretreatment and the
last 3 days of treatment, and different lowercase letters denote significantly different daily means (P < 0.05). P1–P3 denote pretreatment
days, and T1–T7 denote treatment days.
TREE PHYSIOLOGY VOLUME 28, 2008
CLOSE COUPLING OF WHOLE-PLANT RESPIRATION TO NET PHOTOSYNTHESIS
trations both decreased by about 50% (P = 0.008 and 0.0039,
respectively), and leaf nonstructural carbohydrate concentration decreased by 80% (P = 0.0029), however no significant
decrease in root nonstructural carbohydrates was observed
(P = 0.6411). The NL treatment significantly decreased the
concentrations of both leaf and root soluble sugars and carbohydrates. Leaf sugar concentration decreased 41% (P = 0.035)
and root sugar concentration decreased 34% (P = 0.031). Leaf
nonstructural carbohydrate concentration decreased 77% (P <
0.001) compared with a decrease of 52% in roots (P = 0.001).
Correlations between net photosynthesis and respiration
Leaf respiration rate and stem CO2 efflux correlated more
strongly with Anet than with carbohydrate status in saplings in
all treatments. Mean values of Rd and stem CO2 efflux for the
3 days of pretreatment and the last 3 days of treatment strongly
1835
correlated with mean values of Anet for the corresponding days
for saplings in all treatments (Rd: r 2 = 0.78, P < 0.001; stem
CO2 efflux: r 2 = 0.86, P < 0.001) (Figure 6). Weak correlations
between Rd measured on the final day of treatment and sucrose
concentration and starch concentration were observed (sucrose: r 2 = 0.36, P = 0.008; starch: r 2 = 0.47, P = 0.002) (Figure 7). Mean values of root respiration for the 3 days of
pretreatment and the last 3 days of treatment did not correlate
strongly with Anet (r 2 = 0.21, P = 0.08). Final day of treatment
values of root CO2 efflux correlated more closely with tissue
sucrose concentration than with Anet, but correlated poorly
with tissue starch concentration (sucrose: r 2 = 0.44, P = 0.003;
starch: r 2 = 0.11, P = 0.17).
Daily carbon gain (CDG ) and CUE were affected by changes
in Anet in a similar manner as whole-plant respiration (Figure 8). The EC treatment increased CDG by 23% from a 3-day
A. EC treatment
0.05 A. EC treatment
A
15
a/b
B
b
A
0.04
a/b
a
A
a
a
a
0.03
10
a/b
a/b
a
a/b
5
a
a/b
a/b
a
a
a
a
a
0.02
a/b
0
B. LL treatment
A
8
Root CO2 efflux (µmol gDM– 1 s – 1 )
Stem CO2 efflux (µmol m – 2 s – 1 )
0.01
B
a
a
a/b
6
b/c
b/c
4
c
2
0
c
c
C. NL treatment
A
6
c
0.00
A
a
a
B
a
0.04
b
0.03
b/c
0.02
c
c
c
0.01
0.00
B
c
C. NL treatment
a
a
a/b
B. LL treatment
0.05
A
B
0.03
a/b
a
a/b
b
4
0.02
b/c
c
c
c
2
c/d
c
c/d
0.01
d
0
P1 P2 P3
T1 T2
T3
T4
T5
T6
T7
Treatment day
0.00
P1 P2 P3
T1 T2
T3
T4
d
T5
T6
T7
Treatment day
Figure 3. Daily mean (+ SE, n = 3) stem CO2 efflux of Populus
deltoides saplings during the pretreatment (black bars) and treatment
(gray bars) periods in the (A) elevated CO2 (EC), (B) low-light (LL)
and (C) no-light (NL) treatments. Different uppercase letters denote
significantly different means for the 3 days of pretreatment and the
last 3 days of treatment, and different lowercase letters denote significantly different daily means (P < 0.05). P1–P3 denote pretreatment
days, and T1–T7 denote treatment days.
Figure 4. Daily mean (+ SE, n = 3) root CO2 efflux of Populus
deltoides saplings during the pretreatment (black bars) and treatment
(gray bars) periods in the (A) elevated CO2 (EC), (B) low-light (LL)
and (C) no-light (NL) treatments. Different uppercase letters denote
significantly different means for the 3 days of pretreatment and the
last 3 days of treatment, and different lowercase letters denote significantly different daily means (P < 0.05). P1–P3 denote pretreatment
days, and T1–T7 denote treatment days.
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1836
WERTIN AND TESKEY
pretreatment mean of 1.09 to a mean of 1.34 g C day –1 for the
last 3 days of treatment (P = 0.0178). There was an 80% increase in CDG on Day 1 of the EC treatment, followed by a decrease until Day 5 of treatment when it stabilized. The LL
treatment decreased CDG by 70% from a 3-day pretreatment
mean of 0.59 to a mean of 0.18 g C day –1 for the last 3 days of
treatment (P < 0.001) with CDG decreasing by 97% during Day
1 of the LL treatment. Although CDG increased on Day 2 of the
LL treatment, it remained low throughout the treatment. There
was a small (9%) increase in CUE on Day 1 of the EC treatment,
but CUE was not significantly different from the pretreatment
mean by Day 4 of treatment. The LL treatment decreased CUE
by 15% from a 3-day pretreatment mean of 0.64 to a mean of
0.54 on the last 3 treatment days (P = 0.046). Carbon-use efficiency decreased 88% on Day 1 of the LL treatment, before recovering dramatically on Day 2 of treatment. The CUE steadily
increased the last 4 days of treatment and did not appear to
have stabilized completely.
Diurnal patterns in net photosynthesis and respiration
The EC pretreatment and treatment Anet measurements displayed a significant diurnal pattern with post-illumination Anet
measurements being significantly higher than pre-dark measurements (P = 0.012 and P < 0.001, respectively) (Table 1).
Although not significant, a similar trend was observed for the
LL pretreatment and treatment Anet measurements (P = 0.79
and 0.35, respectively) and for the NL pretreatment Anet mea-
Leaf
A
30
Root
B
a
b
a
a
b
20
a
b
a
a
10
a
Discussion
Net photosynthesis and respiration
Changes in Anet in response to increased CO2 availability, decreased light availability, or elimination of light led to large
changes in Rd, stem CO2 efflux and root CO2 efflux. Changes
in respiration in all tissues and for all treatments generally occurred within the first day of treatment. Mean values of Rd and
stem CO2 efflux for the 3 days of pretreatment and the last
3 days of treatment strongly correlated to mean values of Anet
for the corresponding periods, whereas Rd measurements
made on the final day of treatment correlated less strongly
with leaf sucrose or carbohydrate concentration. Root CO2
efflux correlated more strongly with sucrose concentration
b
b
0
15
Ca
D
a
10
a
a
a
a
5
b
b
b
a
a
a
0
EC
LL
NL
EC
LL
NL
Treatment
Rd, Stem CO2 efflux (µmol m – 2 s – 1 )
Root CO2 efflux (µmol gDM– 1 s – 1 )
Nonstructural carbohydrate
concentration (% DM)
Soluble sugar
concentration (% DM)
40
surements (P = 0.47). Leaf respiration, measured three times
daily, showed a significant diurnal pattern in all treatments in
both the pretreatment and treatment phases. In each instance,
mid-light period measurements were significantly higher than
pre-illumination and post-dark measurements, whereas pre-illumination and post-dark Rd measurements were similar. Although not statistically significant, a diurnal trend in stem CO2
efflux was observed that was similar to the observed Rd diurnal
patterns.
There was a significant diurnal pattern in root CO2 efflux
(Figure 9). Root CO2 efflux was at a minimum early in the
light period, and increased steadily during the day, but peaked
before the end of the light period and then subsequently declined through the dark period. This diurnal pattern was observed for each sapling measured in all pretreatment phases
and during the EC and LL treatment periods. Maximum and
minimum daily root CO2 efflux was altered by the treatments,
increasing in response to the EC treatment and decreasing in
response to the LL treatment; however, the general diurnal
trend was preserved.
12
y = 1.01x – 4.72
r 2 = 0.86, P < 0.001
10
8
6
y = 0.0016x + 0.0089
r 2 = 0.21, P = 0.08
4
y = 0.117x + 0.0979
r 2 = 0.78, P < 0.001
2
0
6
Figure 5. Mean (+ SE, n = 3) concentrations on a dry mass (DM) basis
of soluble sugars, extracted from (A) leaves and from (B) roots, and
nonstructural carbohydrates, extracted from (C) leaves and (D) roots,
of Populus deltoides saplings in the elevated CO2 (EC), low-light
(LL) and no-light (NL) treatments during the pretreatment period
(black bars) and the last day of treatment (gray bars). Different letters
indicate a significant difference (P < 0.05) within a treatment.
8
10
12
14
16
Anet (µmol m – 2 s – 1 )
Figure 6. Scatter plot showing the relationship of net photosynthetic
rate (Anet ) with stem CO2 efflux (EC 䉭, LL 䉱, NL 䉱), leaf respiration
rate (Rd; EC 䊊, LL 䊉, NL 䊉) and root CO2 efflux (EC 䊐, LL 䊏,
NL 䊏). Treatment abbreviations: EC, elevated CO2 concentration;
LL, low-light; and NL, no light.
TREE PHYSIOLOGY VOLUME 28, 2008
Rd (µmol m – 2 s – 1 )
Root CO2 efflux (µmol gDM– 1 s – 1 )
CLOSE COUPLING OF WHOLE-PLANT RESPIRATION TO NET PHOTOSYNTHESIS
1837
2.5
2.0
y = 0.0029x – 0.0071
1.5 r 2 = 0.44, P = 0.003
y = 0.0188x + 0.464
r 2 = 0.47, P = 0.002
1.0
0.5
0.0
Figure 7. Scatter plots showing correlations of tissue (A) sucrose and (B) starch
concentrations with leaf respiration rate
(Rd; EC 䊊, LL 䊉, NL 䊉) and root CO2
efflux (EC 䊐, LL 䊏, NL 䊏). Treatment
abbreviations: EC, elevated CO2 concentration; LL, low light; and NL, no light.
y = 0.0425x + 0.104
r 2 = 0.36, P = 0.008
y = 0.0013x + 0.0107
r 2 = 0.11, P = 0.17
0
5
10
15
20
30
25
0
4
2
Sucrose concentration (% DM)
8
6
10
12
Starch content (% DM)
observations and indicate that Rd is strongly dependent on the
availability of current photosynthetic products.
The close coupling between Anet and Rd is further supported
by the observed diurnal patterns in Rd —mid-light period Rd
measurements were higher than pre-illumination Rd measurements. The post-dark Rd measurements, although generally
slightly higher than the predawn Rd measurements, were not as
high as the midday measurements. Thomas and Griffin (1994)
noted that Rd decreased during the first few hours of darkness
and then remained constant throughout the night. A similar
pattern may have occurred in our study, and we speculate that
this was associated with a decrease in immediately available
sucrose in the leaf as a result of increased leaf starch production or sucrose phloem loading to meet whole-plant carbon demand.
Current photosynthetic rates influenced respiration rates
throughout the plant. We found a strong influence of Anet on
stem CO2 efflux, with efflux rates changing within a day of
treatment application. Stem CO2 efflux for the 3 days of
pretreatment and for the last 3 days of treatment correlated
than with Anet; however, a strong diurnal pattern in root CO2
efflux was observed throughout the study suggesting that root
respiration was influenced by daily carbon limitations. Both
the strong statistical correlations between Anet and Rd and stem
CO2 efflux as well as the strong diurnal pattern observed in
root respiration and the time scale in which changes in respiration were observed in response to the treatment application
suggest that respiration in Populus saplings is strongly dependent on newly acquired photosynthetic products.
The increase in Anet in response to the EC treatment caused a
corresponding increase in Rd within the first day of treatment.
Similarly, the LL and NL treatments caused decreases in Anet
that were correlated with decreases in Rd within the first day of
treatment. As well, for each treatment there were strong correlations between Anet and Rd for the 3 days of pretreatment and
for the last 3 days of treatment. Previous studies have shown a
tight coupling between Anet and Rd, with Rd being highly dependent on the previous day’s Anet (Azcón-Bieto and Osmond
1983, Whitehead et al. 2004). Our measurements, which were
made under constant environmental conditions, support these
3.0
A. EC treatment
A
CDG (g day – 1 )
2.5
c
B. LL treatment
b
B
A
B
b
2.0
a/b
1.5
a
a
a/b
a/b
a/b
a/b
1.0
a
a
a/b
0.5
c
0.0
1.0
C. EC treatment
a
A
a
a
b
b/c
b/c
b/c /cb
D. LL treatment
b
b
a/b
A
A
a/b
0.8
a
a
a
B
a
a
CUE
b/c
0.6
a
a
a
a
T5
T6
a
0.4
b
0.2
0.0
P1 P2 P3
T1 T2
T3
T4
T5
T6
T7
P1
P2
P3
T1
T2
T3
T4
Treatment day
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
Figure 8. Mean (+ SE, n = 3)
daily carbon gain (CDG ) of
Populus deltoides saplings in the
(A) elevated CO2 (EC) and (B)
low-light (LL) treatments and
mean daily (+ SE, n = 3) carbon-use efficiency (CUE ) in the
(C) EC and (D) LL treatments.
Different uppercase letters denote
significantly different means for
the 3 days of pretreatment and the
last 3 days of treatment, and different lowercase letters denote
significantly different daily means
(P < 0.05). P1–P3 denote pretreatment days, and T1–T7 denote
treatment days.
1838
WERTIN AND TESKEY
Table 1. Mean (SE, n = 3) net photosynthetic rate (Anet ), leaf respiration rate (Rd ) and stem CO2 efflux of Populus deltoides saplings for each measurement period in the pretreatment and treatment phases of the elevated CO2 concentration (EC), low-light (LL) and no-light (NL) treatments.
Measurements were made 30 min before the light period (Pre-illum., Rd) or 1 h after the start of the light period (Post-illum., Anet and stem efflux),
midway through the light period (Mid-light, Rd and stem efflux), and 1 h before (Pre-dark, Anet and stem efflux) or after (Post-dark, Rd) the start of
the dark period. Different letters indicate significantly different means (P < 0.05) between measurements made on the same tissue for the same
treatment.
Treatment
Anet (µmol m –2 s –1)
Rd (µmol m –2 s –1)
Post-illum.
Pre-illum.
Pre-dark
EC Pretreatment 10.85 (0.20) a
Treatment
18.59 (0.41) a
Stem CO2 efflux (µmol m –2 s –1)
Mid-light
Post-dark
Post-illum.
Mid-light
9.95 (0.25) b 1.10 (0.03) a 1.43 (0.05) b 1.23 (0.03) a 4.71 (0.36) a 4.95 (0.35) a
15.17 (0.68) b 1.46 (0.08) a 1.91 (0.07) b 1.56 (0.06) a 7.68 (0.78) a 8.33 (0.83) a
Pre-dark
4.43 (0.34) a
8.23 (0.80) a
LL Pretreatment 10.18 (0.64) a
Treatment
6.36 (0.24) a
9.92 (0.71) a
6.06 (0.22) a
1.30 (0.08) a 1.80 (0.11) b 1.37 (0.08) a 6.48 (0.38) a 6.90 (0.46) a
0.85 (0.06) a 1.09 (0.57) b 0.1 (0.07) ab 3.36 (0.30) a 2.25 (0.39) a
5.78 (0.46) a
2.21 (0.37) a
NL Pretreatment
Treatment
9.23 (0.24) a
–
0.89 (0.03) a 1.26 (0.11) b 0.86 (0.04) a 4.81 (0.39) a 5.26 (0.35) a
–
–
–
–
–
4.62 (0.32) a
–
9.48 (0.25) a
–
Root CO2 efflux (µmol gDM– 1 s – 1 )
strongly with Anet in all treatments. Few previous studies have
shown such a tight coupling of Anet and stem CO2 efflux. Levy
et al. (1999) suggested that, because the internal CO2 concentration in sapling stems is so high, and has such a large effect
on stem CO2 efflux, daily variations in photosynthesis have no
noticeable effect on daily variation in stem CO2 efflux. We
found that photosynthetic rate had a major effect on stem CO2
efflux and this could be associated with several factors, including the size of the stem, rates of growth and cell division,
sucrose concentrations in the phloem, stem xylem CO2
concentration and stem carbohydrate stored reserves.
We observed a close coupling between Anet and root CO2
efflux in all treatments. The EC treatment increased root CO2
efflux within a day, and the LL and NL treatments caused significant decreases in root CO2 efflux on Day 1 of treatment.
Root CO2 efflux correlated more strongly with tissue sucrose
concentration then with starch concentration or Anet. Root CO2
efflux displayed a strong diurnal pattern suggesting that root
respiration, which regulates root growth, cellular maintenance
and biosynthesis, is subjected to daily carbohydrate limitation.
0.05
0.04
0.03
0.02
0.01
0.00
T1
T2
T3
Treatment day
Figure 9. Representative diurnal pattern of root CO2 efflux of one
Populus deltoides sapling over 3 days of pretreatment (P1–P3). Dark
periods are shaded.
Horwath et al. (1994) performed a 14CO2 pulse chase experiment with 2-year-old hybrid poplar saplings and measured 14C
efflux from the soil within 12 h, with the maximum rate occurring 2 days after pulse application. However, when large trees
are girdled it can take days (Högberg et al. 2001, Frey et al.
2006) to weeks (Andersen et al. 2005, Binkley et al. 2006) before a change in soil CO2 efflux is detected, and the magnitude
of the response was much less than observed in our study.
Measurements of soil respiration made on trees in the forest
include CO2 efflux from root respiration, microbial and fungal
respiration, and organic matter decomposition which may
dampen the magnitude of the response. The discrepancy between the observations of Högberg et al. 2001 and Frey et al.
2006 and our findings suggests that large trees have proportionately more stored carbohydrate reserves in their root
systems than the rapidly growing saplings that we studied.
The consistent diurnal pattern of increasing root respiration
during most of the light period indicated that the sapling root
systems were experiencing daily carbohydrate limitation,
which was greatest after the dark period, and least near the end
of the light period. This finding provides further evidence that
Anet had an almost immediate (within hours) influence on
whole-plant respiration. Few previous studies have observed
diurnal patterns in root CO2 efflux. Kuzyakov and Cheng
(2001) reported a diurnal pattern in soil efflux in wheat growing in a temperature-controlled environment, with efflux increasing after illumination by about 25–50% compared with
that in the dark. They concluded, as we have, that root respiration is controlled by newly acquired photosynthetic products.
However, Betson et al. 2007 observed no diurnal pattern in soil
respiration in girdled or ungirdled plots of trees, suggesting
that large trees have significant carbohydrate reserves that
mitigate daily carbon limitations.
Carbohydrates
At the end of the EC treatment, leaf soluble sugar concentration had significantly increased over pretreatment values.
Tjoelker et al. (1998) reported that five boreal tree species
grown in elevated CO2 concentrations had higher starch con-
TREE PHYSIOLOGY VOLUME 28, 2008
CLOSE COUPLING OF WHOLE-PLANT RESPIRATION TO NET PHOTOSYNTHESIS
centrations in leaves and roots compared with plants grown in
ambient CO2 concentration. We found no such large increases
in response to EC as observed by Tjoelker et al. (1998), suggesting that increases in tissue respiration consumed some of
the additionally available photosynthetic products. This notion
is supported by the finding that CDG decreased to near pretreatment values by the end of the EC treatment period.
After six days of LL treatment, leaf soluble sugar and carbohydrate concentrations were significantly lower than pretreatment values. A similar pattern was observed by Marenco et al.
(2001), who noted that leaf sugar and starch concentrations
were higher in fully sunlit trees than in shaded trees. Kull and
Niinemets (1998) concluded that leaf total nonstructural carbohydrate concentrations are a function of the light environment of the leaf. Our findings can be explained by the greatly
reduced photosynthetic rate in LL saplings with no concurrent
decrease in stem or root carbon demand. A significantly larger
fraction of the available leaf photosynthate was probably
transported to other plant tissues, leading to a decrease in
available sucrose and subsequently in Rd.
There was a significant decrease in root soluble sugar at the
end of the LL treatment, suggesting that when Anet decreased
the saplings were unable to meet current root carbon demands,
thus causing decreases in root CO2 efflux. Both the immediate
(within the first treatment day) decrease in root CO2 efflux and
the strong diurnal pattern in root CO2 efflux suggest that root
respiration of the study saplings was regulated by daily carbon
availability. The responses to the LL treatment were accentuated in the NL saplings where significant decreases in leaf and
root carbohydrate and soluble sugar were measured. However,
despite significantly depressed Rd and stem and root CO2
efflux at the end of the NL treatment, there were still substantial amounts of sucrose and carbohydrates in the plant tissue. It
is possible that the plants decreased respiration rates when Anet
was reduced or completely inhibited, thereby preserving a carbohydrate pool for an extended period, which would allow survival until environmental conditions become more favorable.
In conclusion, we observed significant changes in wholeplant respiration when Anet was altered. Changes in respiration
generally occurred within the first day of treatment, indicating
that respiration throughout the plant was carbohydrate limited.
Rapid changes in leaf respiration and stem and root CO2 efflux
after treatments were imposed also indicated that respiration
in rapidly growing saplings may vary daily depending on current photosynthetic rates. The strong diurnal pattern in root
CO2 efflux suggested that the saplings experienced daily carbon limitations even under conditions of high photosynthetic
carbon gain.
Acknowledgments
We thank Mary Anne McGuire for help and input on the project set up
and equipment usage. We thank Drs. Scott Merkle and Jeff Dean for
assistance with carbohydrate analysis, and Eboni Hall for assistance
in growing the plant material. This work was supported by a grant
from the United States Department of Energy NICCR Program (Grant
07-SC-NICCR-1060).
1839
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