Title: Effects of N on plant response to heat-wave: a field study with prairie vegetation
Running title: Effects of N on plant response to heat-wave
Authors: Dan Wang1, Scott A. Heckathorn1, Kumar Mainali1, E. William Hamilton2
1
Dept. of Environmental Sciences, University of Toledo, Toledo, OH, USA
2
Dept. of Biology, Washington & Lee University, Lexington, VA, USA
Author for correspondence:
Dan Wang
Tel: 1-419-530-2925;
E-mail: [email protected]
1
1
2
Abstract
More intense, more frequent, and longer heat-waves are expected in the future due to
3
global warming, which could have dramatic ecological impacts. Increasing nitrogen (N)
4
availability and its dynamics will likely impact plant responses to heat stress and carbon (C)
5
sequestration in terrestrial ecosystems. This field study examined the effects of N availability
6
on plant response to heat-stress (HS) treatment in naturally-occurring vegetation. HS (5 days
7
at ambient or 40.5 ºC) and N treatments (± N) were applied to 16 1m2 plots in restored prairie
8
vegetation dominated by Andropogon gerardii (warm-season C4 grass) and Solidago
9
canadensis (warm-season C3 forb). Before, during, and after HS, air, canopy, and soil
10
temperature were monitored; net CO2 assimilation (Pn), quantum yield of photosystem II
11
(ФPSII), stomatal conductance (gs), and leaf water potential (Ψw) of the dominant species and
12
soil respiration (Rsoil) of each plot were measured daily during HS. One week after HS, plots
13
were harvested, and C% and N% were determined for rhizosphere and bulk soil, and above-
14
ground tissue (green/senescent leaf, stem, and flower). Photosynthetic N-use efficiency
15
(PNUE) and N resorption rate (NRR) were calculated. HS decreased Pn, gs, Ψw, and PNUE
16
for both species, and +N treatment generally increased these variables (±HS), but often
17
slowed their post-HS recovery. Aboveground biomass tended to decrease with HS in both
18
species (and for green leaf mass in S. canadensis), but decrease with +N for A. gerardii and
19
increase with +N for S. canadensis. For A. gerardii, HS tended to decrease N% in green
20
tissues with +N, while in S. canadensis, HS increased N% in green leaves. Added N
21
decreased NRR for A. gerardii and HS increased NRR for S. canadensis. These results
22
suggest that heat waves, though transient, could have significant effects on plants,
23
communities, and ecosystem N cycling, and N can influence the effect of heat waves.
24
25
Key words: heat stress, global climate change, N resorption rate, photosynthesis,
26
photosynthetic N-use efficiency.
2
1
Abbreviations
2
Cg: C% of green leaf; Cs: C% of senescent leaf; Cst, C% of stem; Cf, C% of flower; Ng: N%
3
of green leaf; Ns: N% of senescent leaf; Nst, N% of stem; Nf, N% of flower; gs, stomatal
4
conductance to water vapor; HS, heat stress; Ψw , leaf water potential; NRR, nitrogen
5
resorption rate; Pn, net photosynthesis; PNUE, photosynthetic N-use efficiency; ФPSII,
6
quantum yield of electron transport of photoystem II; SLA, specific leaf area, Rsoil, soil
7
respiration; Wa, aboveground biomass; Wf, biomass of flowers; Wg, biomass of green leaf;
8
Ws, biomass of senescent leaf; Wst, biomass of stem.
3
1
2
Introduction
Global mean surface temperatures have risen by 0.6 °C from 1900 to 2000, mainly
3
caused by increases in atmospheric CO2 and other greenhouse gases, and are projected to
4
increase by another 1.4-5.8 °C by year 2100 (Houghton et al. 2001; IPCC 2007). In addition
5
to rising mean annual temperatures, there will also be increases in the frequency, duration,
6
and severity of periods with exceptionally high temperatures (Wagner 1996). An increased
7
trend in the frequency of extreme heat stress events has been reported in various parts of the
8
world (Gaffen & Ross 1998; Gruza & Ran'kova 1999; Henderson & Muller 1997; Yan
9
2002). Thus, plants in the future will not only be exposed to higher mean temperatures, but
10
will also likely experience more frequent heat stress, which can greatly impact ecosystem
11
productivity (Ciais et al. 2005) and biodiversity (Thomas et al. 2004). An extreme stress
12
event is an episode in which the acclimatory capacities of an organism are substantially
13
exceeded (Gutschick & BassiriRad 2003). Extreme events, in spite of their ephemeral nature,
14
can cause shifts in the structure of plant communities. The environmental impacts from
15
extreme events can be significantly greater than those associated with mean increases (Karl
16
et al. 1997).
17
In addition to temperatures, human activities are increasing global N availability
18
(IPCC 2007). N availability is likely to affect plant, community, and ecosystem responses to
19
increasing heat stress, which will then impact ecosystem C sequestration. Understanding
20
effects of N on the responses of vegetation to heat stress requires insight into how stress
21
physiology and community structure interact. While the influence of plant N status on
22
response to acute heat stress has been previously examined, past studies have largely focused
23
on laboratory experiments examining physiological responses (Heckathorn et al. 1996a, b;
24
Lu & Zhang 2000). Further, because of the difficulties of imposing heat stress on naturally-
25
occurring vegetation, little experimental work has been conducted on response to acute heat
26
stress in field-grown plants (Morison & Lawlor 1999; Weis & Berry 1987). To date, there
27
have been only a handful of studies in which plant communities were exposed to extreme
28
high temperatures, and these focused on community processes (e.g., recolonization,
29
competition, invasion, and the role of species richness during extreme events) and were
30
conducted on grassland (Van Peer et al. 2004; White et al. 2001) or arctic species (Marchand
31
et al. 2006; Marchand et al. 2005). Also, N availability had significant effects on plant N-
4
1
relations responses to moderate warming (rather than acute heat stress) in a tallgrass prairie
2
(An et al. 2005). Thus, little is known as to how heat stress in general, and N interactions
3
with heat stress in particular, will affect natural plant communities. In this study, we
4
concentrate on physiological and growth responses of two dominant warm-season tall-grass
5
prairie species with contrasting photosynthetic pathways (a C4 grass and a C3 forb) in
6
experimental field plots receiving heat and N treatments.
7
C4 species typically have higher temperature optima for photosynthesis than C3
8
species (Sage & Monson 1999) as a consequence of lower photorespiration, which increases
9
with temperature. This may contribute to greater tolerance to heat waves for C4 species than
10
co-occurring C3 species (Coleman and Bazzaz 1992, Ehleringer et al. 1997, Wang et al.
11
2008). Thus, heat stress can potentially affect the relative distribution of C4 and C3 species. In
12
natural systems, the significance of climate warming for C4 vegetation can depend less on the
13
mean increase in global temperature, and more on the spatial and temporal variation of the
14
temperature increase (Sage & Kubien 2003). In New Zealand, for example, episodic heat
15
events inhibit C3 plants more than C4 grasses, and as a result, facilitate C4 grass invasion of
16
C3-dominated grasslands (White et al. 2000, 2001). On the other hand, because of their
17
greater nitrogen (N) investment in rubisco and photorespiratory enzymes, C3 plants have
18
lower N-use efficiency of photosynthesis (PNUE) than C4 plants (Li 1993; Sage & Pearcy
19
1987). The ecological consequences of greater PNUE in C4 species have been studied to only
20
a limited degree. For example, in grasslands, when soils are low in N, C4 grasses can be
21
superior competitors to C3 grasses and can dominate (Wedin & Tilman 1996). When soils
22
become N-enriched, the advantage in PNUE is offset and C3 species can match the
23
photosynthetic potential of C4 species, and thereby increase in cover. However, whether N
24
availability interacts with heat stress differently in C3 vs. C4 species remains to be
25
determined, but will have a bearing on the relative impact of global environmental change on
26
C3 and C4 species abundance and distribution.
27
To examine the influence of N on plant response to heat stress in naturally-occurring
28
mixed C3-C4 vegetation, we conducted a field study with the following three major
29
objectives: (1) to determine how heat stress affects the ecophysiological and morphological
30
variables of naturally-occurring co-dominant C4 and C3 species; (2) to determine the effect of
31
N on resistance and resilience of each species to heat stress; (3) to investigate how heat stress
5
1
affects plant C and N concentration, N-use efficiency, and N resorption rate. We predicted
2
that (1) heat stress will have a more pronounced negative effect on the C3 than the C4 species;
3
(2) supplemental N will help both the C3 and C4 species to better tolerate heat stress,
4
especially for the C3 species; and (3) heat stress will increase leaf N concentration and
5
decrease N-use efficiency, as a result of decreased leaf expansion and photosynthesis, but
6
more so for the C3 species.
6
1
2
Results
During heat stress (HS), air temperature in the heated plots was increased on average
3
to 40.5 ± 2.8 °C. During the five days of heat treatment, leaf temperature of A. gerardii and
4
S. canadensis in heated plots was higher than that in control plots, but returned to control
5
levels after heat stress (Fig 1, Table 1). Nitrogen treatment had no effect on leaf temperature
6
for A. gerardii, but for S. canadensis, plants with N treatment had a lower leaf temperature.
7
Soil temperature (at 10-cm) was not altered by heat or N treatment (not shown). C% of both
8
rhizosphere and bulk soil was not changed by heat stress, and N% of both rhizosphere and
9
bulk soil was not impacted by heat stress, but was increased by N treatment (P<0.01) (not
10
11
shown).
Leaf water potential (Ψw) was decreased for heated plants, but N had little effect on
12
Ψw; and Ψw was recovered one week post heat-stress (day 12) and was similar among
13
treatments (Fig 2, Table 1). Soil (=soil+root) respiration (Rsoil) was decreased by both heat
14
and N, but there was no interactive effect of N and heat, and post-heat-stress Rsoil was similar
15
for ±HS in +N plants but still lower in +HS vs. –HS plants with no added N (Fig 3, Table 1).
16
There was an overall negative effect of heat treatment on net photosynthesis (Pn). For both A.
17
gerardii and S. canadensis, Pn was significantly lower in heated plots than in control plots
18
during heat stress. N had no significant effect on Pn for A. gerardii, but for S. canadensis, N
19
increased Pn and there was a significant heat x N interaction (Fig 4, Table 1). Further, Pn
20
remained depressed one week after HS in +HS plants with added N, relative to un-heated
21
controls, but this was not observed in plants receiving no added N. Variation in stomatal
22
conductance to water vapor (gs) was a function of both heat and N. For A. gerardii and S.
23
canadensis, gs was lower in heated plots and higher at N-treated plots. There was also a
24
significant interactive effect of heat and N on gs for S. canadensis (Fig 4, Table 1). Also, gs
25
remained depressed one week after HS in +HS plants (more so in +N), relative to un-heated
26
controls, especially for S. canadensis. Quantum yield of electron transport (ФPSII) was not
27
decreased by heat stress for A. gerardii and S. canadensis, but N had a significant positive
28
29
effect on ФPSII for A. gerardii and S. canadensis (Fig 4, Table 1).
For A. gerardii, N treatment increased specific leaf area (SLA), but heat did not affect
30
SLA, while for S. canadensis, neither N or heat affected SLA (Fig 5, Table 2, 3). Total
31
aboveground biomass (Wa) was not significantly affected by heat or N in either species, but
7
1
there was a decrease in Wa with heat in both species (ANOVA including both species;
2
P=0.024), and increases in Wa with added N in S. canadensis and decreases in Wa with added
3
N in A gerardii (Fig 6, Table 2&3). Biomass of green leaf, stem, and senescent leaf was not
4
significantly altered by either heat or N treatment for both A. gerardii and S. canadensis.
5
However, flower biomass was increased significantly by N treatment for S. canadensis and
6
decreased by heat stress for A. gerardii. And for A. gerardii without N treatment, the percent
7
of senescent leaf was significantly higher in heated plots (18.2%) than at control plots
8
(12.3%).
9
Carbon concentration of plant tissue (C%) was altered by both heat and N treatment,
10
but the effects differed with species and among different plant parts (Fig 7, Table 2&3). For
11
A. gerardii, C% of N-treated plants was lower in green leaves and flowers, but higher in non-
12
heated senescent leaves. Heat decreased C% only in senescent leaves for A. gerardii with N
13
treatment. C% was not altered by either heat or N in green leaf, senescent leaf, stem, and
14
flower for S. canadensis. In general, nitrogen concentration (N%) was increased in N-treated
15
plant tissues (excluding flowers) for both A. gerardii and S. canadensis (Fig. 7, Table 2&3).
16
Heat had little effect on N% in A. gerardii, though there was a tendency for decreases in N%
17
in N-treated plants, while in S. canadensis, heat increased N% in green leaves, and for A.
18
gerardii, there was also an interactive effect of heat and N on N% in green leaves.
19
Photosynthetic nitrogen-use efficiency (PNUE) was significantly lower for S.
20
canadensis than A. gerardii and was decreased by heat for both A. gerardii and S. canadensis
21
(Fig 8, Table 1). In heated plots, plants with N treatment tended to have a higher PNUE for
22
A. gerardii, though the effect was not statistically significant. Further, recovery of PNUE was
23
incomplete after one week post-HS, relative to un-heated controls, for both species and N
24
levels. Nitrogen resorption rate (NRR) was decreased by N treatment, but not changed by
25
heat, for A. gerardii, while for S. canadensis, NRR was significantly higher for heated plants,
26
but was not different due to N treatment (Fig 9, Table 2&3).
8
1
Discussion
2
During heat stress, both A. gerardii (C4) and S. canadensis (C3) experienced
3
decreased Pn, gs, Ψw, PNUE, and soil (=soil+root) respiration decreased too; decreases in Pn,
4
gs, PNUE, and soil respiration were still evident one week after heat treatment ended (day
5
12). In general, N addition affected these physiological variables in both heated and
6
unheated-plants (increasing Pn, gs, Ψw, PNUE, but decreasing soil respiration). With few
7
exceptions, during heat stress, N did not alter the nature of the heat-stress effect on these
8
variables (i.e., there was no significant heat x N interaction). In contrast, after one week post-
9
heat-stress recovery, residual heat-stress-related decreases in Pn and gs were evident only
10
(Pn), or greater (gs), in high-N plants, and decreases in soil respiration were only evident in
11
low-N plants. Both species exhibited trends of decreasing aboveground biomass with heat
12
treatment, while added N tended to increase biomass in S. canadensis but decrease biomass
13
in A. gerardii. Carbon concentration (C%) of tissues was affected only by heat treatment in
14
A. gerardii (leaves and stems), while N% of green leaves in A. gerardii decreased with heat
15
stress (+N only) but increased with heat stress in S. canadensis. Lastly, heat treatment
16
increased N resorption rate (NRR) in S. canadensis, but not in A. gerardii, while added N
17
decreased NRR for A. gerardii, but not in S. canadensis.
18
Collectively, these results indicate the heat waves imposed here were of moderate
19
severity (as evidenced by the magnitude of HS effects), yet such moderate heat waves can
20
affect plant C and N relations and biomass growth and allocation, and the heat effects can
21
still be evident after one week of post-heat recovery. Further, many plant responses to the
22
heat treatment were influenced by N availability and differed between the C3 species, S.
23
canadensis, and the C4 species, A. gerardii. Specifically, these results suggest that in a future
24
warmer world with increasing N availability, S. canadensis may be affected less by heat
25
waves than A. gerardii , but in the absence of more N, the reverse may be true. It is also
26
worth noting that heat and N effects in this study may be smaller than likely to occur, as our
27
heat treatment was a single heat wave (and plants in Northwest Ohio experience ca. 3-5 heat
28
waves per summer) and our N treatment was initiated only 2 weeks prior to heat stress. Thus,
29
predictions of N effects on heat-stress responses and differences between C3 and C4 plants
30
based on this study may be conservative.
9
1
Any N-related influence on heat-induced changes in plant physiology, growth,
2
biomass allocation, and tissue C and N concentration, and any such differences in N x heat
3
effects between C3 and C4 species, will have important implications for plant herbivory and
4
decomposition, and thus, for ecosystem N and C dynamics. For example, heat-related
5
increases in tissue senescence and changes in C or N% will have a direct impact on herbivore
6
feeding preference and growth rate, and on litter quantity and quality, and hence on
7
decomposition rates. A shift in the ratio of C3:C4 species with increasing heat waves in the
8
presence/absence of higher N would have dramatic impact on ecosystem N or C cycling also,
9
as C4 foliage is characterized by a higher carbon-to-nitrogen (C:N) ratio and higher fiber
10
content than C3 foliage, thus contributing high C:N litter to soil organic matter, which results
11
in low N mineralization rates (Sage & Monson 1999; Wedin & Tilman 1996). High C:N
12
ratios also reduce decomposition rates, such that proportionally more N on a site may reside
13
in the soil organic matter pool (Aerts 1997); thus N availability often declines when a C4-
14
dominated sward replaces C3 vegetation (Reich et al. 2001).
15
Plants appear to be more susceptible to high day or night-time temperatures during
16
later flower-to-early seed developmental stages (Cross et al. 2003). Notably, in this study,
17
flower biomass (Wf) was significantly reduced for A. gerardii by heat stress, and this
18
decrease was somewhat smaller in high-N plants. Heat stress had no effect on Wf for S.
19
canadensis, suggesting that increases in heat waves in the future may affect the seed bank of
20
this plant community, which might affect community structure in the longer term. Heat-
21
stressed plants can compensate for decreases in flower production by producing later flowers
22
on existing inflorescences (Cross et al. 2003; Sato et al. 2000), but whether plants can still
23
compensate for decreased flower production after a late-growing-season heat stress remains
24
to be investigated.
25
The heat-treatment effects on plant C and N relations observed in this study differ
26
somewhat compared to results from previous studies. For example, while we observed
27
increased N% in green leaves with heat stress in S. canadensis, past studies applying long-
28
term warming observed decreases in N% of green leaves with warming (Tjoelker et al. 1999,
29
An et al. 2005). In contrast, other warming studies showed that elevated temperature
30
increased leaf N concentration due to enhanced soil N mineralization and increased plant N
31
uptake (Luomala et al. 2003; Nijs et al. 1996). We also observed unique effects on N
10
1
resorption (NRR) from senescing leaves in this study, compared to previous studies. Here,
2
NRR was decreased by N treatment but not changed by heat stress for A. gerardii; but for S.
3
canadensis, NRR was increased by heat stress but was not altered by N treatment. The
4
decreased NRR of A. gerardii due to N treatment in our study is consistent with the
5
observation that species from nutrient-poor environments often have a higher N resorption
6
rate than species from nutrient-rich environments (Aerts 1997). The partitioning of N
7
compounds between soluble and structural compounds is an important regulator of N
8
resorption (Norby et al. 2001; Yuan et al. 2005). The increased NRR of S. canadensis in the
9
heated plots might be caused by acceleration of the normal senescence and resorption
10
process. Warming has been observed to accelerate the senescence of leaves, such that
11
warmed plants completed the normal senescence and resorption process faster than those in
12
un-warmed controls, resulting in plant litter in the warmed plots either having a lower N
13
concentration or lower fraction of N in soluble compounds (Norby et al. 2000).
14
The present experiment showed that heat stress, though ephemeral, can potentially
15
modify community composition and impact ecosystem nutrient cycling, via effects on plant
16
growth and tissue N and C content. Further, increases in N availability may influence plant
17
response to heat stress; e.g., as slowing recovery of heat-related damage to photosynthesis,
18
and benefiting C3 species more than C4 species during heat stress. This study only examined
19
short-term plant responses to acute heat stress within one generation of perennial plants, but
20
the results indicate that the impact of acute heat stress on plant communities and ecosystems
21
should be studied more extensively, particularly in combination with other potentially-
22
interactive aspects of global environmental change (e.g., CO2, O3, and precipitation).
23
24
25
26
27
28
29
30
31
11
1
Materials and methods
2
3
4
Field site and treatments
The experiment site was located within restored prairie vegetation at the University of
5
Toledo’s Stranahan Arboretum (Toledo, Ohio, USA), which is located within the oak-
6
savannah glacial-sand ecosystem referred to as the “Oak Openings” region
7
(http://oakopen.org/). Andropogon gerardii (big bluestem), a warm-season C4 perennial
8
grass, and Solidago canadensis (goldenrod), a warm-season C3 perennial herbaceous dicot,
9
are the two dominant plant species in this field site. The experiment design was a 2x2
10
factorial (±heating x ±added N; with n=4 replicates per treatment combination), utilizing
11
16x1m2 randomly selected and assigned-treatments plots. Eight of the plots received heat
12
treatment for five days from 17 to 21 August 2006, and eight of the plots received added N
13
treatment (NH4NO3) applied twice (one and two weeks) before heat treatment at a rate of 5g
14
N/m2/year. Heat treatment was applied by using eight top-vented 1m3-chambers made with
15
transparent plastic attached to a wooden frame. A portable electric heater with a maximum
16
capacity of 1500W was installed in the chamber to increase air temperature and an electric
17
fan was used to circulate warm air inside the chamber. The target treatment temperature was
18
41 °C, which is 10 °C higher than average daytime temperature for August in Toledo, and 2-
19
3 °C higher than the typical maximum temperature in the summer season in this area. Heat
20
treatments were imposed during daytime for five days and for 10-h per day (8:00 am to 6:00
21
pm). Control plots were not covered by chambers and experienced ambient temperature. The
22
air temperature and leaf temperatures inside and outside the chambers were monitored using
23
data-loggers and fine-wire thermocouples during heat treatments; soil temperature at 10-cm
24
depth was measured with a thermometer.
25
26
Physiological Measurements
27
Before, during, and after heat stress, net photosynthesis, stomatal conductance to
28
water vapor, quantum yield of electron transport of photosystem II (PSII), and leaf water
29
potential were measured daily on randomly-chosen recently-expanded fully-lit leaves.
30
Steady-state net photosynthesis (Pn; net CO2 exchange) and stomatal conductance (gs) of
31
single leaves was measured with a portable photosynthesis system containing an infrared gas
12
1
analyzer (model 6400, LiCOR, Lincoln, NE, USA), equipped with a 250-mm3 leaf chamber
2
as in (Heckathorn et al. 1997). Measurements were made at ambient light and temperature
3
within one min of insertion of leaves into the cuvette (immediately after stabilization of CO2
4
and H2O fluxes). Quantum yield of PSII electron transport (ФPSII) was measured with a
5
pulse-amplitude-modulated (PAM) fluorometer (Model PAM 101/103, Walze, Germany), as
6
in Wang et al. (2008). A pressure chamber (Model 600, PMS Instruments Co., Corvallis,
7
Oregon) was used to measure midday leaf water potential (Ψw).
8
9
Biomass and C, N measurements
One-week after heat stress, 40x50 cm2 of each plot was harvested. The clipped plants
10
11
were sorted into different categories (green and senescent leaves, stems and flowers), oven-
12
dried at 65 °C for one week and weighed. N and C concentration for different plant parts, as
13
well as rhizosphere and bulk soil, was measured with a Perkin Elmer CHN Analyzer (Model
14
2400). N resporation rate (NRR) for each species was calculated by NRR = (Ng-Ns)/Ng,
15
where Ng was the green leaf N concentration and Ns was the senescent leaf N concentration.
16
Photosynthetic N-use efficiency (PNUE) was based on net photosynthesis per unit plant N.
17
18
19
20
Statistical analysis
Analyses were conducted within each species to determine whether the physiological
variables differed as a function of different treatments. For daily-measured variables like Ψw,
21
Pn, gs, PNUE, ФPSII, Rsoil, and leaf temperature, three-way analysis-of-variance (ANOVA)
22
(SAS 9.1) was used to test for significant effects of days, heat, N, and their interaction during
23
heat stress (day 1-day 5). A two-way ANOVA was used to test for significant effects of heat,
24
N, and their interaction on Pn, gs, PNUE, ФPSII, Rsoil, and leaf temperature after heat stress
25
(i.e., on day 12), and on biomass, C%, and N% of plants and soil. In order to test for species
26
effect, three-way ANOVA was conducted on biomass, C% and N%, with species, heat, N
27
and their interactions as independent factors. Days, heat, and N were all treated as fixed
28
effects.
13
1
Acknowledgements
2
We thank Daryl Moorhead, Sandra Stutzenstein, and Walter Schulisch for providing access
3
to experimental field sites, as well as for logistical support and assistance conducting the
4
experiment. We thank Jiquan Chen for providing us vehicles and experimental equipment.
5
Thanks to Sasmita Mishra and Rajan Tripathee for assistance with plant harvest and
6
processing plant materials. We are indebted to Rachel Henderson for her advice on carbon
7
and nitrogen analysis. We thank the reviewers for their helpful comments on this manuscript.
8
This research was supported by a grant from the National Science Foundation to SAH and
9
EWH.
14
1
References
2
Aerts R (1997). Nitrogen partitioning between resorption and decomposition pathways: a
3
trade-off between nitrogen use efficiency and litter decomposibility? Oikos 80: 603-606.
4
An YA et al. (2005). Plant nitrogen concentration, use efficiency, and contents in a tallgrass
5
prairie ecosystem under experimental warming. Glob. Change Biol. 11: 1733-1744.
6
Ciais P et al. (2005). Europe-wide reduction in primary productivity caused by the heat and
7
drought in 2003. Nature 437: 529-533.
8
Cross RH, Mckay SAB, Mchughen AG and Bonham-Smith PC (2003). Heat-stress
9
effects on reproduction and seed set in Linum usitatissimum L. (flax). Plant Cell Environ. 26:
10
1013-1020.
11
Gaffen DJ, Ross RJ (1998). Increased summertime heat stress in the US. Nature 396: 529-
12
530.
13
Gruza GV, Ran'kova EY (1999). Estimation of the climate response to changes in
14
greenhouse gas concentration from surface air temperature observations over Russia.
15
Izvestiya Akademii Nauk Fizika Atmosfery I Okeana 35: 742-749.
16
Gutschick VP, Bassirirad H (2003). Extreme events as shaping physiology, ecology, and
17
evolution of plants: toward a unified definition and evaluation of their consequences. New
18
Phytol. 160: 21-42.
19
Heckathorn SA, Coleman JS and Hallberg RL (1997). Recovery of net CO2 assimilation
20
after heat stress is correlated with recovery of oxygen-evolving-complex proteins in Zea
21
mays L. Photosynthetica 34: 13-20.
22
Heckathorn SA, Poeller GJ, Coleman JS and Hallberg RL (1996a). Nitrogen availability
23
alters patterns of accumulation of heat stress-induced proteins in plants. Oecologia 105: 413-
24
418.
25
Heckathorn SA, Poeller GJ, Coleman JS and Hallberg RL (1996b). Nitrogen availability
26
and vegetative development influence the response of ribulose 1,5-bisphosphate
27
carboxylase/oxygenase, phosphoenolpyruvate carboxylase, and heat-shock protein content to
28
heat stress in Zea mays L. Int. J. Plant Sci. 157: 546-553.
29
Henderson KG, Muller RA (1997). Extreme temperature days in the south-central United
30
States. Clim. Res. 8: 151-162.
15
1
Houghton J, Ding Y and Griggs D (2001). Climate change of 2001. The Scientific Basis.
2
Cambridge, UK: Cambridge University Press.
3
IPCC (2007). Climate Change 2007: The Physical Science Basis. Contribution of Working
4
Group I to the Fourth Assessment Report of the Intergovernmental Panel on Climate Change.
5
. Cambridge University Press, Cambridge.
6
Karl TR, Nicholls N and Gregory J (1997). The coming climate. Scientific American 276:
7
78-83.
8
Li MR (1993). Leaf photosynthetic nitrogen-use efficiency of C-3 and C-4 Cyperus species.
9
Photosynthetica 29: 117-130.
10
Lu CM, Zhang JH (2000). Photosystem II photochemistry and its sensitivity to heat stress
11
in maize plants as affected by nitrogen deficiency. J. Plant Physiol. 157: 124-130.
12
Luomala EM, Laitinen K, Kellomaki S and Vapaavuori E (2003). Variable
13
photosynthetic acclimation in consecutive cohorts of Scots pine needles during 3 years of
14
growth at elevated CO2 and elevated temperature. Plant Cell Environ. 26: 645-660.
15
Marchand FL, Kockelbergh F, Van De Vijver B, Beyens L and Nijs I (2006). Are heat
16
and cold resistance of arctic species affected by successive extreme temperature events? New
17
Phytol. 170: 291-300.
18
Marchand FL, Mertens S, Kockelbergh F, Beyens L and Nijs I (2005). Performance of
19
High Arctic tundra plants improved during but deteriorated after exposure to a simulated
20
extreme temperature event. Glob. Change Biol. 11: 2078-2089.
21
Morison JIL, Lawlor DW (1999). Interactions between increasing CO2 concentration and
22
temperature on plant growth. Plant Cell Environ. 22: 659-682.
23
Nijs I, Teughels H, Blum H, Hendrey G and Impens I (1996). Simulation of climate
24
change with infrared heaters reduces the productivity of Lolium perenne L in summer.
25
Environ. Exp. Bot. 36: 271-280.
26
Norby RJ, Cotrufo MF, Ineson P, O'neill EG and Canadell JG (2001). Elevated CO2,
27
litter chemistry, and decomposition: a synthesis. Oecologia 127: 153-165.
28
Norby RJ, Long TM, Hartz-Rubin JS and O'neill EG (2000). Nitrogen resorption in
29
senescing tree leaves in a warmer, CO2-enriched atmosephere. Plant Soil 224: 15-29.
16
1
Reich PB, Peterson DW, Wedin DA and Wrage K (2001). Fire and vegetation effects on
2
productivity and nitrogen cycling across a forest-grassland continuum. Ecology 82: 1703-
3
1719.
4
Sage RF, Kubien DS (2003). Quo vadis C-4? An ecophysiological perspective on global
5
change and the future of C4 plants. Photosynth. Res. 77: 209-225.
6
Sage RF, Monson RK (1999). C4 Plant Biology. Academic Press. Harcourt Brace &
7
Company, Publishers.
8
Sage RF, Pearcy RW (1987). The nitrogen use efficiency of C-3 and C-4 plants. 2. Leaf
9
nitrogen effects on the gas-exchange characteristics of Chenopodium album (L) and
10
Amaranthus retroflexus (L). Plant Physiol. 84: 959-963.
11
Sato S, Peet MM and Thomas JF (2000). Physiological factors limit fruit set of tomato
12
(Lycopersicon esculentum Mill.) under chronic, mild heat stress. Plant Cell Environ. 23: 719-
13
726.
14
Thomas CD et al. (2004). Extinction risk from climate change. Nature 427: 145-148.
15
Van Peer L, Nijs I, Reheul D and De Cauwer B (2004). Species richness and susceptibility
16
to heat and drought extremes in synthesized grassland ecosystems: compositional vs
17
physiological effects. Funct. Ecol. 18: 769-778.
18
Wagner D (1996). Scenarios of extreme temperature events. Clim. Change 33: 385-407.
19
Wedin DA, Tilman D (1996). Influence of nitrogen loading and species composition on the
20
carbon balance of grasslands. Science 274: 1720-1723.
21
Weis E, Berry JA (1987). Quantum efficiency of photosystem II in relation to energy-
22
dependent quenching of chlorophyll fluorescence. Biochimica Et Biophysica Acta 894: 198-
23
208.
24
White TA, Campbell BD, Kemp PD and Hunt CL (2000). Sensitivity of three grassland
25
communities to simulated extreme temperature and rainfall events. Glob. Change Biol. 6:
26
671-684.
27
White TA, Campbell BD, Kemp PD and Hunt CL (2001). Impacts of extreme climatic
28
events on competition during grassland invasions. Glob. Change Biol. 7: 1-13.
29
Yan YY (2002). Extreme temperature days in Hong Kong. Physical Geography 23: 476-491.
17
1
Yuan ZY, Li LH, Han XG, Huang JH and Wan SQ (2005). Foliar nitrogen dynamics and
2
nitrogen resorption of a sandy shrub Salix gordejevii in northern China. Plant Soil 278: 183-
3
193.
4
5
18
1
Fig. 1 Effects of heat wave (open symbols - heat stress; dark symbols - without heat stress)
2
and N (circle symbols - without N treatment; triangle symbols - with N treatment) on leaf
3
temperature of A. gerardii and S. canadensis. Day 1-5 refers to the five days during heat
4
treatment; day 12 refers to one week after the end of heat treatment when plants were in
5
recovery. C, CH, N, and NH indicate plants with no treatment, heat, N and heat*N treatment,
6
respectively. Values are means ± 1 SE; n=4.
7
8
Fig. 2 Effects of heat wave (open symbols - heat stress; dark symbols - without heat stress)
9
and N (circle symbols - without N treatment; triangle symbols - with N treatment) on leaf
10
water potential (Ψw) of A. gerardii and S. canadensis. Day1-5 refers to the five days during
11
heat treatment; day 12 refers to one week after the end of heat treatment when plants were in
12
recovery. C, CH, N, and NH indicate plants with no treatment, heat, N and heat*N treatment,
13
respectively. Values are means ± 1 SE; n=4.
14
15
Fig. 3 Effects of heat wave (open symbols - heat stress; dark symbols - without heat stress)
16
and N (circle symbols - without N treatment; triangle symbols - with N treatment) on soil
17
respiration (Rsoil). Day1-5 refers to the five days during heat treatment; day12 refers to one
18
week after the end of heat treatment when plants were in recovery. C, CH, N, and NH
19
indicate plants with no treatment, heat, N and heat*N treatment, respectively. Values are
20
means ± 1 SE; n=4. Inserted is F-Statistics from ANOVA.
21
22
Fig. 4 Effects of heat wave (open symbols - heat stress; dark symbols - without heat stress)
23
and N (circle symbols - without N treatment; triangle symbols - with N treatment) on net
24
photosynthesis (Pn), stomatal conductance (gs) and quantum yield of photosystem-II electron
25
transport (ФPSII) of A. gerardii and S. canadensis. Day1-5 refers to the five days during heat
26
treatment; day12 refers to one week after the end of heat treatment when plants were in
27
recovery. C, CH, N, and NH indicate plants with no treatment, heat, N and heat*N treatment,
28
respectively. Values are means ± 1 SE; n=4.
29
19
1
Fig. 5 Effects of heat wave and N on specific leaf area (SLA) for A. gerardii and S.
2
canadensis. Bars labeled by C, CH, N, and NH indicate plants with no treatment, heat, N and
3
heat*N treatment, respectively. Values are means ± 1 SD; n=4.
4
5
Fig. 6 Effects of heat wave and N on total above-ground, green leaves, stem, flower and
6
senescent leaves biomass for A. gerardii and S. canadensis. Bars labeled by C, CH, N, and
7
NH indicate plots with no treatment, heat, N and heat*N treatment, respectively. n=4.
8
9
Fig. 7 Effects of heat wave and N on C% and N% of green leaf (GL), stem (ST), flower (FL)
10
and senescent leaf (SL) for A. gerardii and S. canadensis. Bars labeled by C, CH, N, and NH
11
indicate plots with no treatment, heat, N and heat*N treatment, respectively. Values are
12
means ± 1 SD; n=4.
13
14
Fig. 8 Effects of heat wave (open symbols - heat stress; dark symbols - without heat stress)
15
and N (circle symbols - without N treatment; triangle symbols - with N treatment) on
16
photosynthetic nitrogen use efficiency (PNUE) of A. gerardii and S. canadensis. Day1-5
17
refers to the five days during heat treatment; day12 refers to one week after the end of heat
18
treatment when plants were in recovery. C, CH, N, and NH indicate plants with no treatment,
19
heat, N and heat*N treatment, respectively. Values are means ± 1 SE; n=4.
20
21
Fig. 9 Effects of heat wave and N on nitrogen resorption rate (NRR) for A. gerardii and S.
22
canadensis. Bars labeled by C, CH, N, and NH indicate plots with no treatment, heat, N and
23
heat * N treatment, respectively. Values are means ± 1 SD; n=4.
20
Table 1. Degrees of freedom (numerator and denominator df) and F-statistics from ANOVA on individual plant variables in response
to heat and nitrogen treatment in a restored prairie in Toledo, OH, with days (day1-day5), heat, and nitrogen and their interactions as
independent factors. F-statistics with a single asterisk indicated significance at P<0.10, whereas a double asterisk indicates P<0.05.
See text for abbreviations.
Variables
Factor
Days
Heat
N
Days*heat
Days*N
Heat*N
Days*heat*N
df
4,56
1,56
1,56
4,56
4,56
1,56
4,56
Tleaf
43.92**
386.51**
0.00
32.09**
2.15*
1.91
2.46**
A. gerardii
Pn
gs
35.44** 15.73** 4.57**
5.14** 41.90** 5.86**
3.01*
2.38
0.59
0.40
8.69** 0.95
0.28
0.26
1.21
0.88
0.56
0.58
0.11
1.59
1.89*
Ψw
ФPSII
PNUE
11.12** 15.53**
14.76**
0.17
1.53
2.71*
0.97
0.71
2.35** 0.36
2.00
2.56
1.61
0.53
Tleaf
43.38**
454.55**
4.61*
31.22**
0.71
0.40
4.42**
S. canadensis
Pn
gs
33.21** 13.54** 21.49**
4.27** 13.66** 57.77**
9.46** 21.30**
0.54
2.15*
4.13** 1.39
1.36
0.92
1.33
5.26**
3.33*
0.79
0.76
0.83
2.13*
Ψw
ФPSII
7.62**
0.03
9.03**
2.03*
0.77
0.28
1.86*
PNUE
12.81**
18.01**
1.43
3.77**
0.85
0.24
0.12
21
Table 2. Degrees of freedom (numerator and denominator df) and F-statistics from ANOVA on individual plant variables for A.
gerardii, in response to heat and nitrogen treatment in a restored prairie in Toledo, OH, with heat, N and their interactions as
independent variables. F-statistics with a single asterisk indicated significance at P<0.10, whereas a double asterisk indicates P<0.05.
See text for abbreviations.
Factor
Heat
N
Heat*N
df
1,12
1,12
1,12
Variables
SLA Wg Wst Wf
Ws
Wa
Cg
Ng
Cs
5.05**
1.23
0.99 0.02 0.06 3.45* 0.04 0.06 1.73
3.98* 0.59 0.08 0.17 1.33 0.68 6.29** 5.79** 4.67**
4.22* 3.99*
0.38 0.03 0.03 0.05 0.23 0.06 0.01
Ns
0.00
16.57**
0.07
Cst
1.24
0.42
0.04
Nst
0.35
2.78
0.96
Cf
0.04
14.91**
0.07
Nf
2.46
0.04
0.49
NRR
0.06
30.66**
0.64
22
Table 3. Degrees of freedom (numerator and denominator df) and F-statistics from ANOVA on individual plant variables for S.
canadensis, in response to heat and nitrogen treatment in a restored prairie in Toledo, OH, with heat, N and their interactions as
independent variables. F-statistics with a single asterisk indicated significance at P<0.10, whereas a double asterisk indicates P<0.05.
See text for abbreviations.
Factor
df
SLA
Heat
1,12 0.25
N
1,12 0.50
Heat*N 1,12 1.61
Wg
2.06
0.17
0.86
Wst
0.09
1.91
0.00
Wf
0.14
3.15*
0.97
Ws
0.68
0.27
0.17
Wa
0.27
1.63
0.12
Cg
0.75
0.02
0.11
Variables
Ng
Cs
12.64** 0.04
14.71** 0.53
0.36
1.37
Ns
0.26
6.73**
0.16
Cst
1.35
0.26
5.07**
Nst
8.46**
37.55**
15.68**
Cf
1.27
1.79
0.07
Nf
0.10
2.67
0.57
NRR
6.12**
0.01
0.01
23
Fig 1
40
Leaf temperature (° C)
30
A. gerardii
20
C
CH
N
NH
10
40
30
20
S. canadensis
10
1
2
3
4
5
12
During and after heat stress (days)
24
Fig 2
0.0
-.5
C
CH
N
NH
A. gerardii
Leaf water potential (mPa)
-1.0
-1.5
-2.0
0.0
-.5 S. canadensis
-1.0
-1.5
-2.0
1
2
3
4
5
12
During and after heat stress (days)
25
Soil respiration ( Rsoil; µmol CO2 m-2 s-1)
Fig 3
700
600
500
400
C
CH
N
NH
300
200
100
1
2
3
4
5
12
During and after heat stress (days)
26
transport (relative units)
Quantum yield of electron
Stomatal conductance
(gs, mol H2O m-2 s-1)
Net photosynthesis
(Pn, µmol CO2 m-2 s-1)
Fig 4
A. gerardii
S. canadensis
40
C
CH
N
NH
30
40
30
20
20
10
10
.4
S. canadensis
A. gerardii
.4
.3
.3
.2
.2
.1
.1
.7
.7
.6
.6
.5
.5
.4
.4
S. canadensis
A.gerardii
.3
1
2
3
4
5
12
.3
1
2
3
4
5
12
During and after heat stress (days)
27
Specific leaf area (m2/kg)
Fig 5
30
C
CH
N
NH
25
20
15
10
5
A. gerardii
S. canadensis
28
Fig 6
A. gerardii
120
100
80
60
Biomass (g)
40
20
S. canadensis
120
green leaf
stem
flower
senescent leaf
100
80
60
40
20
C
CH
N
NH
29
Fig 7
60
C%
60
S.canadensis
A. gerardii
50
50
40
40
30
30
2.5
2.5
A. gerardii
S. canadensis
N%
2.0
2.0
C
CH
N
NH
1.5
1.5
1.0
1.0
.5
.5
GL
SL
ST
FL
GL
SL
ST
FL
30
Photosynthetic nitrogen use efficiency (PNUE)
Fig 8
.8
A. gerardii
.6
.4
.2
.8
S. canadensis
.6
C
CH
N
NH
.4
.2
1
2
3
4
5
12
During and after heat stress (days)
31
N resorption rate (%)
Fig 9
60
C
CH
N
NH
45
30
15
A. gerardii
S. canadensis
32