Relationship between photosynthetic CO2 uptake rate and electron ...

Acta Physiol Plant (2014) 36:849–857 DOI 10.1007/s11738-013-1463-y

ORIGINAL PAPER

Relationship between photosynthetic CO2 uptake rate and electron transport rate in two C4 perennial grasses under different nitrogen fertilization, light and temperature conditions Shau-Lian Wong • Chung-Wei Chen • Meng-Yuan Huang • Jen-Hsien Weng

Received: 21 May 2013 / Revised: 21 November 2013 / Accepted: 9 December 2013 / Published online: 22 December 2013 Ó Franciszek Go´rski Institute of Plant Physiology, Polish Academy of Sciences, Krako´w 2013

Abstract We examined the feasibility of using chlorophyll fluorescence to estimate CO2 exchange (A) of C4 perennial grasses under different environmental as well as physiologic conditions using Pennisetum purpureum and Miscanthus floridulus, capable of year-round growth, to determine the association of electron transport rate (ETR) and A. The grasses were fertilized with three levels of nitrogen, and measurement involved the top two fully expanded leaves, with chlorophyll content 0.18–0.55 g m-2. Chlorophyll fluorescence, CO2 and H2O exchange were measured simultaneously at four seasonal temperatures (30–15 °C, September–January), six levels of photosynthetic photon flux density (PPFD) (0–2,000 lmol m-2 s-1) and two levels of relative humidity [60 % (15–30 °C) and 40 % (30 °C alone)]. Variables were recorded when A was stable. Most leaves with high chlorophyll content showed high A at the same PPFD and seasonal temperature. Despite a broad range of A obtained because of both stomatal and non-stomatal factors, ETR was still highly correlated linearly with net photosynthetic rates, when combining data for the same species for analysis. Thus, ETR could be used to assess the dynamic A of C4 perennial species through different seasons,

Communicated by L. A. Kleczkowski. S.-L. Wong C.-W. Chen Division of Botany, Taiwan Endemic Species Research Institute, Chichi, Nantou, Taiwan M.-Y. Huang Biodiversity Research Center, Academia Sinica, Taipei, Taiwan J.-H. Weng (&) Graduate Institute of Ecology and Evolutionary Biology, China Medical University, Taichung, Taiwan e-mail: [email protected]

even under varied light intensity, seasonal temperature, humidity, nitrogen fertilization and phenological stage. Keywords C4 Electron transport rate Light intensity Nitrogen fertilization Photosynthetic rate Seasonal temperature Abbreviations A CO2 fixation a Leaf absorption Ca Ambient CO2 concentration Ci Intercellular CO2 concentration ETR Electron transport rate f The fraction of absorbed quanta used by PSII UCO2 Photosynthetic rate per absorbed quantum UPSII Photosystem II efficiency gs Stomatal conductance N Nitrogen PN Net photosynthetic rate PPFD Photosynthetic photon flux density PSII Photosystem II RH Relative humidity

Introduction Photosynthesis, one of the major determinants of biomass production and terrestrial carbon budgets, is influenced by many environmental and physiological factors (Berry and Downton 1982). Understanding the biomass production of crops and vegetation under different conditions requires monitoring spatial and temporal variations in photosynthetic capacity. However, the traditional measurement of photosynthetic capacity, especially in the field, is limited

123

850

by cumbersome techniques used to measure leaf gas exchange. Recently, chlorophyll fluorescence quenching analysis has been found to be a fast, simple, non-invasive, and reliable method to assess changes in photosystem II (PSII) function under different environmental and physiological conditions (Rohaćˇek and Barta´k 1999; Maxwell and Johnson 2000). Among chlorophyll fluorescence variables, electron transport rate (ETR), calculated from the product of PSII efficiency and absorbed light, expresses the relative rate of electron transport through PSII (Krall and Edwards 1992). Electron flow through PSII is consumed mainly by carbon assimilation (Krall and Edwards 1992; Oberhuber and Edwards 1993). When CO2 fixation (A) is inhibited by certain environmental and physiological stresses, leaves may downregulate their PSII efficiency (UPSII), mainly through xanthophyll-dependent non-photochemical quenching to avoid damage caused by excess absorbed energy (Demmig-Adams et al. 1996; Kato et al. 2003; Adams et al. 2004). Thus, a linear correlation between ETR and A can be obtained for many C3 and C4 plants. However, in C3 plants, both CO2 fixation and photorespiration are major sinks for electrons from PSII. Therefore, with increased photorespiration, the ratio of ETR to A [or UPSII to photosynthetic rate per absorbed quantum ( UCO2 )] greatly increases with decreasing CO2 partial pressure (Krall and Edwards 1990; Cornic and Briantais 1991) or increasing temperature (Oberhuber and Edwards 1993; D’Ambrosio et al. 2006; Wong et al. 2012) as well as O2 partial pressure (D’Ambrosio et al. 2006; Ripley et al. 2007). In C4 plants, photorespiration is restricted, absorbed photons may be mostly used to drive the CO2 fixation (Krall and Edwards 1992). Numerous studies have concluded a strong linear relationship between ETR and A. This relationship appears to be stable under many conditions (Edwards and Baker 1993), even when merging data from different genotypes (Earl and Tollenaar 1998) or with differences in both CO2 partial pressure and temperature (Kakani et al. 2008) across a broad range of light intensity. However, the ETR–A relationships in C4 species may be complicated because of uncertain ETR based on fluorescence measurements: chlorophyll fluorescence may occur from both the mesophyll and bundle sheath cells (Romanowskaa et al. 2006). Moreover, in addition to A and photorespiration, electrons from PSII have several energy sinks, such as the water–water cycle (Asada 1999) and the cyclic electron flow within PSII (Miyake and Okamura 2003) as well as nitrogen (N) assimilation (Robinson 1990). Furthermore, CO2 leakage from bundle sheath cells is another possibility for electron usage in C4 plants and is influenced by many conditions (Kubien et al. 2003; Naidu and Long 2004; Yin et al. 2011b). Thus, the ETR–A (or

123

Acta Physiol Plant (2014) 36:849–857

UPSII–UCO2 ) relationship of C4 plants may vary by genotype (D’Ambrosio et al. 2003), leaf N (Khamis et al. 1990), growth season or growth conditions (Earl and Tollenaar 1998; Fryer et al. 1998; Baerlocher et al. 2004; Farage et al. 2006) or even conditions during measurement (Earl and Tollenaar 1998). Leaf N is an important factor affecting photosynthesis. Many C4 and C3 species show a strong positive correlation between A and leaf N content (Weng and Chen 1987; Makino and Osmond 1991; Weng and Hsu 2001; Lawlor 2002). Most leaf N is allocated to chloroplasts (Makino and Osmond 1991; Lawlor 2002), and lower A under limited N is often attributed to reduced chlorophyll content and enzyme activities (Verhoeven et al. 1997; Lawlor 2002; Cheng 2003; Huang et al. 2004; Netto et al. 2005; Zhao et al. 2005), which leads to decreased light absorption and light utilization by leaves (Verhoeven et al. 1997; Cheng et al. 2001). N deficiency also results in significant decreases in UPSII (Cheng et al. 2001) and increases the electron flux to alternative sinks (Kumagai et al. 2010). However, for C4 plants, the effect of leaf N on the ETR– A (or UPSII– UCO2 ) relationship has not been studied in detail. Khamis et al. (1990) noted a small decrease in UCO2 at any given UPSII in N-limited maize leaves. However, Cheng et al. (2001) noted that when leaves of apple (a C3 plant) were measured under non-photorespiratory conditions and quantum yield was expressed on an absorbed PPFD basis, a close UPSII– UCO2 relationship could be found for leaves with a wide range of N content. Eco-physiological studies require knowledge of photosynthetic characteristics of plants at different environmental and physiological conditions. Under well-water conditions, light intensity, temperature and leaf N status are important factors in photosynthesis. Chlorophyll fluorescence measurement is a fast and simple method for estimating photosynthesis. However, for C4 plants, the combined effects of leaf N, light intensity and seasonal temperature on the ETR–A relationship is not clear, especially for subtropical perennial C4 grasses. In addition, stomatal conductance (gs) is another important limiting factor in photosynthesis. Stomata prevent water loss and facilitate CO2 diffusion to mesophyll cells. To optimize water-use efficiency, gs and A may be closely related under many conditions, including changing light intensity (D’Ambrosio et al. 2003; Huxman and Monson 2003), temperature (Pittermann and Sage 2000, 2001) and leaf N status (Weng and Hsu 2001; Mohotti and Lawlor 2002). In this study, we used two perennial C4 grasses to elucidate the ETR–A relationship in varied seasons under different seasonal temperature, photosynthetic photon flux density (PPFD), and relative humidity. We also discuss the effects of stomatal and non-stomatal limitations.


Materials and methods Plant materials Two perennial C4 plants, Pennisetum purpureum (cv. A146), a cultivated herbage, and Miscanthus floridulus, a wild grass, were used as materials. The former was propagated from cuttings, and the latter was collected from the lowlands of central Taiwan. Plants were potted (38 cmdiameter) in a mixture of soil:vermiculite:sand (1:1:1), and placed outdoors to receive regular water and full sunlight in the nursery of the Endemic Species Research Institute, Chichi Township, Nantou County, Taiwan (23°490 N, 120°480 E, 250 m a.s.l.). To obtain leaf samples with different photosynthetic capacity, 0, 1 and 2 g N fertilizer was applied to pots at 4 and 2 weeks before measurement. During the growth period (May 2010–January 2011), the mean monthly temperature was 26.6–28.9 °C (May– August) and 28.5–14.8 °C (September–January) (data from the Endemic Species Research Institute). Measurements Measurements were carried out from September 2010 to January 2011. Pot-grown plants were exposed to outdoor sunlight before measurement. We measured the top two fully expanded leaves with different chlorophyll content [0.18–0.55 g m-2, determined by use of a chlorophyll meter (SPAD 502, Minolta Camera Co., Osaka, Japan)]. From 0930 to 1500 hours, photosynthesis, stomatal conductance and chlorophyll fluorescence were measured by use of a portable, open-flow gas exchange system (LI-6400, LI-COR, Lincoln, NE, USA) with an integrated fluorescence chamber head (LI-6400-40) stepwise from high to low levels (2,000; 1,200; 800, 400, 200 and 0 lmol m-2 s-1) of PPFD. The values of A, gs, ratio of intercellular to atmospheric CO2 concentration (Ci/Ca) and chlorophyll fluorescence variables were recorded when A was stable. Leaf temperature was maintained at 15, 20, 25 and 30 °C. Measurements were taken in the season with the climate temperature close to the leaf temperature. The 30 °C measurement was in September to October and the 25, 20 and 15 °C measurements were in November, December and January, respectively. Throughout the measurements, CO2 concentration in the chamber was kept at 350–400 lmol mol-1 (no control). Relative humidity (RH) of air entering the chamber was 60 % (for all four temperatures) and 40 % (for 30 °C only), which was controlled by passing temperature-controlled water. The actual PSII efficiency (DF/Fm0 ) was calculated as (Fm0 -F)/Fm0 . F and Fm0 are the actual and maximal levels of fluorescence during illumination, respectively; the

851

former was determined under each PPFD level, and the latter was determined by applying a 0.8-s pulse of saturating flashes of approximately 6,000 lmol quanta m-2 s-1. ETR was calculated as UPSII 9 PPFD 9 a 9 f (Maxwell and Johnson 2000). The variables a and f are the fractions of incident PPFD absorbed by the leaf and absorbed PPFD used by PSII, respectively. We used the mean leaf absorption of 0.84 for green leaves (Bjo¨rkman and Demmig 1987; see ‘‘Discussion’’ section), and 0.4 for f of C4 plants (Yin and Struik 2012). Statistical analysis We measured 5–10 (usually 6–7, Fig. 3) leaves from four pots for each species. Data were analyzed by linear regression by use of Sigma Plot v10.0.

Results We used P. purpureum measured at 30 and 15 °C and 60 % RH as an example for illustrating the light-response curves of PN, gs, ETR and Ci/Ca (Figs. 1, 2). Typically, PN, gs and ETR showed a hyperbolic increase with increasing PPFD, and leaves with higher chlorophyll content always had higher PN, gs and ETR, the closest relation being between ETR and PN. PN, gs, and ETR values were lower at low than high seasonal temperature in all leaves. The Ci/ Ca for both species decreased with increasing PPFD and stabilized somewhat at about 1,200 lmol m-2 s-1 PPFD with most treatments (Fig. 3). However, as compared with a given level of PPFD, low temperature and high RH produced high Ci/Ca for both species. Yet, under 30 °C and 60 % RH, even leaves with high chlorophyll content had higher gs but still showed lower Ci/Ca than leaves with low chlorophyll content. Although PN was influenced by many conditions, including stomatal and non-stomatal factors, ETR was still highly correlated with PN in both species when merging data from leaves with different chlorophyll content and measured under different PPFD, temperature and RH (Fig. 4a, c). However, both the slope and determination coefficient (r2) for the ETR–PN correlation were greater for P. purpureum than M. floridulus. In contrast, the gs–PN correlation was not as close as ETR–PN correlation for both species. At low temperature, many PN values were low in the gs–PN regression lines (Fig. 4b, d).

Discussion We examined the association of ETR and A for two C4 perennial grasses by merging data from varied seasons and

123

852


Fig. 1 Light-response curves for Pennisetum purpureum measured in September to October at 30 °C and 60 % relative humidity. PN represents net photosynthetic rate; ETR represents electron transport rate; gs represents stomatal conductance; Ci represents intercellular CO2 concentration; Ca represents atmospheric CO2 concentration;

closed and open symbols represent high (0.39–0.50 g m-2) and low (0.18–0.33 g m-2) chlorophyll content, respectively. Each point represents the value for one leaf. Numeric values in panel A are chlorophyll content for each leaf

different N fertilization, light intensity, seasonal temperature and RH. The top two fully expanded leaves with high chlorophyll content showed high PN at the same PPFD and seasonal temperature (Figs. 1, 2, and data not shown). This result might be due to both leaf chlorophyll content and photosynthetic characteristics being influenced by leaf N (Verhoeven et al. 1997; Lawlor 2002; Cheng 2003; Huang et al. 2004; Ding et al. 2005; Netto et al. 2005; Zhao et al. 2005). ETR is calculated as the product of PSII efficiency and absorbed light. a may vary by leaf chlorophyll content. Previously, we found that the correlation between a and SPAD value (Minolta SPAD-502 chlorophyll meter reading) could be fitted with the following equation (Jiang 2007):

a = 0.80–0.89), we found a similar association of PN and ETR regardless of use of a = 0.84 or 0.80–0.89 for calculating ETR (Fig. 5). Thus, we used a = 0.84 in the present study. Many reports have indicated that chlorophyll content always increases linearly with increasing leaf N (e.g., Cheng 2003; Netto et al. 2005; Zhao et al. 2005). Chlorophyll content can be detected easily and quickly by use of a chlorophyll meter (e.g., Netto et al. 2005). Thus, we used chlorophyll content measured by a chlorophyll meter as an indicator of leaf N. Photosynthesis is limited by both stomatal and nonstomatal factors. The former is associated with decreased leaf Ci caused by stomata closure, and the latter is associated with decrease in photochemical efficiency and CO2 fixation (Berry and Downton 1982; Brodribb 1996). Both A and gs decrease under N deficiency (Huang et al. 2004; Ding et al. 2005; Zhao et al. 2005). However, the response of Ci accompanying the decreases in A and gs are not uniform: such leaves may show higher Ci (Huang et al. 2004; Ding et al. 2005) or lower Ci (Zhao et al. 2005) than leaves with non-deficient N supply. We found similar

a ¼ 0:0034SPAD þ 0:7266

ð1Þ

We used the data shown in Fig. 1 to compare the association of PN and ETR calculated with the mean of a (0.84) (Bjo¨rkman and Demmig 1987) or with the result of Eq. 1. Even in leaves with a broad range of chlorophyll content (0.18–0.55 g m-2, SPAD = 22.4–49.5 and

123


853

and open symbols represent high (0.39–0.55 g m-2) and low (0.25–0.33 g m-2) chlorophyll content, respectively. Each point represents the value for one leaf. Numeric values in panel A are chlorophyll content for each leaf

Fig. 2 Light-response curves for Pennisetum purpureum measured in January at 15 °C and 60 % relative humidity. PN represents net photosynthetic rate; ETR represents electron transport rate; gs represents stomatal conductance; Ci represents intercellular CO2 concentration; Ca represents atmospheric CO2 concentration; closed

0.9

B Pennisetum purpureum

A Miscanthus floridulus

0.8 0.7

Ci/Ca

(6) 0.6

(6)

0.5

(6) (7) (10) (6)

0.4

(6) (7) (6) (5)

0.3 0.2

0

500

1000

1500

2000

0

500

1000

1500

2000

PPFD (µmol m s ) -2

Fig. 3 Effect of photosynthetic photon flux density (PPFD) on ratio of intercellular to atmospheric CO2 concentration (Ci/Ca) at different seasonal temperatures. Closed circles, closed triangles, open circles and open triangles represent at 60 % relative humidity and 15 °C (in January), 20 °C (in December), 25 °C (in November) and 30 °C (in

-1

September–October), respectively; inverted open triangles represents at 40 % relative humidity and 30 °C (in September–October). Data are mean ± SD; numeric values within the parentheses are sample size of each measurement

123

854

Acta Physiol Plant (2014) 36:849–857 40

Miscanthus floridulus

B

30

20

10

40

0 20 40 60 80 100 120 140 160 D C ETRSLC (µmol e m s ) -

30

-2

-1

10

y = 0.265 x -1.189 r 2 = 0.985*** 0

20

40

60

80

100 120 140 160 0.0

ETR (µmol e m s ) -

-2

-1

Fig. 4 The correlation of electron transport rate (ETR) and stomatal conductance (gs) with net photosynthetic rate (PN) for Miscanthus floridulus and Pennisetum purpureum. Closed circles, closed triangles, open circles and open triangles represent at 60 % relative humidity and 15 °C (in January), 20 °C (in December), 25 °C (in

-2

-1

P N (µmol CO2 m s )

40 y = 0.220x - 1.471 r 2 = 0.990***

30

20 y = 0.209 x - 1.075 r 2 = 0.992***

10 40

gs (mol H2O m-2 s-1)

20

0

60

80

100 120 140 160 180

ETR ( µmol e- m-2 s-1) Fig. 5 The association of electron transport rate (ETR) and net photosynthetic rate (PN) for Pennisetum purpureum at 60 % relative humidity and 30 °C. Data from Fig. 1 [SPAD value (Minolta SPAD502 chlorophyll meter reading) = 22.4–49.5]. Closed symbol and solid line vs. open symbol and dotted line—a = 0.84 and a ¼ 0:0034SPAD + 0:7266(a = 0.80–0.89), respectively, used for calculating ETR. Each point represents the value for each leaf measured under each level (0–2,000 lmol m-2 s-1) of photosynthetic photon flux density. ***Represents P \ 0.001

123

y = 88.664 x + 1.660 r 2 = 0.698***

y = 0.235 x - 0.633 r 2 = 0.952***

0

Pennisetum purpureum

PN (µmol CO2 m-2 s-1)

A

y = 106.751 x + 0.230 r 2 = 0.742*** 0.1

0.2

0.3

0.4

gs (mol H2O m-2 s-1) November) and 30 °C (in September–October), respectively; inverted open triangles represents at 40 % relative humidity and 30 °C (in September–October). Each point represents the value for each leaf measured under each level (0–2,000 lmol m-2 s-1) of photosynthetic photon flux density. ***Represents P \ 0.001

results. Under all conditions, most leaves with high chlorophyll content had high PN and gs (Figs. 1, 2 and data not shown). However, at 30 °C and 60 % RH, Ci/Ca was greater in leaves with low than high chlorophyll content (Fig. 1d). This finding might due to reduced carboxylation efficiency (Huang et al. 2004; Ding et al. 2005). In contrast, at 15 °C, all tested leaves showed high Ci/Ca, and the difference in Ci/Ca was less than at 30 °C. This finding might be due to key C4 enzymes, such as the cold liability of pyruvate Pi dikinase is sensitive to low temperature (Pittermann and Sage 2000; Kubien et al. 2003; Kakani et al. 2008) and leads to the carboxylation efficiency of all leaves being more limited by low temperature than gs (Pittermann and Sage 2000, 2001; Kubien et al. 2003; Kosćielniak and Biesaga-Kosćielniak 2006). As well, many PN values, obtained at low temperature, were low in the gs– PN regression lines (Fig. 4b, d). In addition, gs may be reduced in response to increased vapor pressure difference between leaf and air (Morison and Gifford 1983; Yong et al. 1997; Maherali et al. 2003). We showed that low RH could decrease Ci/Ca (Fig. 3).


For many C4 plants, including Miscanthus, A increases up to a Ci of about 200–250 lmol mol-1 (Ziska and Bunce 1997; Huxman and Monson 2003; Naidu and Long 2004). We measured photosynthesis under 350–400 lmol mol-1 CO2 and found that Ci/Ca could decrease to \0.5 when measured at 30 °C, especially under 40 % RH. Under these conditions, Ci was lower than the saturated portion of the A–Ci curve for many C4 plants. Thus, A might be limited by low Ci. In C4 or C3 plants under non-photorespiration (e.g., low O2 and saturated CO2), because A is a major sink for electrons from PSII, the allocation electron flow between A and other alternative sinks in C4 plants remains unchanged under many conditions. This situation explains the close association of ETR and A (or UPSII and UCO2 ) in many cases, including under changed Ci value (Krall and Edwards 1992; Oberhuber and Edwards 1993; Naidu and Long 2004; Farage et al. 2006; Kakani et al. 2008). In terms of the effect of N on the ETR–A (or UPSII– UCO2 ) association in C4 plants, maize leaves under low N showed a small decrease in UCO2 at any given UPSII (Khamis et al. 1990). In addition, the partitioning of electron flow between CO2 assimilation and photorespiration was not affected by leaf N content in leaves of apple (C3 fruit tree) (Cheng et al. 2001). However, probably because of PSII heterogeneity and changes in electron flow to sinks other than CO2 assimilation, the UPSII– UCO2 relationship under non-photorespiratory conditions was curvilinear when merging data from apple leaves with a wide range of leaf N content and measured at 87–1,990 lmol m-2 s-1 PPFD, low 2 % O2 and saturated CO2 (1,300 lmol mol-1) (Cheng et al. 2001). In the present study, we derived a broad range of leaf-scale photosynthetic rate from variation in N fertilizer and seasonal temperature, as well as PPFD and RH during measurement. Even when merging all data for the same species, PN still showed a close linear correlation with ETR (Fig. 4a, c). This result indicated no change in proportion of electron flow to CO2 assimilation and alternative sinks. The allocation of electron flow between A and other alternative sinks may be altered in low-temperature-grown C4 plants (Fryer et al. 1998; Farage et al. 2006). However, the threshold temperature for change in allocation electron flow varies by species. For example, PSII electron transport relative to A increased for 17 °C-grown Cyperus longus but for 10 °C-grown Miscanthus 9 giganteus (Farage et al. 2006). In the present study, the mean monthly temperature of the coldest month (January) remained near 15 °C. Probably, this temperature was not low enough to change the electron flow of the tested species. In addition, the ETR/A (or UPSII/ UCO2 ) ratio may increase with advancing growth for annual C4 crops (Earl and Tollenaar 1998) and winter-dormant perennial C4 grasses grown at high altitude (45°500 N) (Baerlocher et al. 2004). However, in the

855

lowlands of subtropical Taiwan, perennial C4 grasses can grow year-round. We measured the top two fully expanded leaves. Probably for these reasons, the ETR–A relation of the two tested species measured in different seasons was not influenced by phenological stages. Also, we found no change in ETR/A ratio with temperature for either species (Fig. 4a, c), which suggests no marked effects on CO2 leakage and dark respiration under light (Naidu and Long 2004). However, the slope for the ETR–PN correlation was higher for P. purpureum than M. floridulus (Fig. 4a, c). C4 maize cultivars have shown differences in the UPSII– UCO2 relationship, due to differences in allocation of electron flow between A and other alternative sinks (D’Ambrosio et al. 2003). As well, PN is determined by subtracting dark respiration from gross photosynthesis; thus, in addition to the effects of alternative sinks, the ETR–PN relation may be affected by dark respiration. According to Yin et al. (2011a), dark respiration under light is closely related to the intercept value at the PN-axis of the ETR–PN linear relationship. The absolute value of this intercept was higher for P. purpureum than M. floridulus (Fig. 4a, c), so P. purpureum had higher dark respiration under light than M. floridulus. Thus, the higher slopes for the ETR–A correlation in P. purpureum than M. floridulus was not due to low dark respiration. The ETR– PN relation was not influenced markedly by both gs and carboxylation efficiency within a species (Fig. 4a, c), and more electrons were not diverted to non-photochemical quencing in M. floridulus than P. purpureum (data not shown). The correlation coefficient between ETR and PN greater for P. purpureum than M. floridulus may be due to the lower proportion of electron flow to alternative sinks (Fryer et al. 1998; D’Ambrosio et al. 2003; Farage et al. 2006). From Fig. 4a, c and Yin et al. (2011a), the ratio of ETR to gross photosynthetic rate for P. purpureum and M. floridulus was about 3.8 and 4.3, respectively. The latter value was similar to that for both maize, an annual C4 crop (Earl and Tollenaar 1998) and Spartina alterniflora, a highaltitude-grown perennial C4 grass, in the early part of the growth season (Baerlocher et al. 2004), but lower than that for maize at the late grain-filling stage (Earl and Tollenaar 1998) and growth under controlled environmental conditions in a climate chamber (D’Ambrosio et al. 2003). However, the ETR/A value for M. floridulus was only 3.8, smaller than the theoretical minimum value (ca. 4, Edwards and Baker 1993). Because we used a lower flash strength (6000 lmol m-2 s-1) than saturating (Loriaux et al. 2013), to assess Fm0 , UPSII might have been under-measured for both tested species. In conclusion, we tested two perennial C4 grasses that can sustain year-round growth in subtropical Taiwan to determine the association of ETR and A. Despite a broad

123

856

range of A obtained because of both stomatal and nonstomatal factors caused by different light, humidity, seasonal temperature, phenological stage and N fertilization, ETR was still highly correlated linearly with net photosynthetic rates, when data from the same species were combined for analysis. Thus, ETR could be used to assess the dynamic A for some perennial C4 species through different seasons. However, the ETR–A correlation may vary by many conditions and may not apply to all C4 plants under all conditions. Author contribution SLW designed research, analysed data and wrote the manuscript; CWC carried out the laboratory experiments and analyzed the data; MYH contributed to the design of research; JHW designed and conducted research and wrote the manuscript.

References Adams WW III, Zarter CR, Ebbert V, Demmig-Adams B (2004) Photoprotective strategies of overwintering evergreens. Bioscience 54:41–49 Asada K (1999) The water–water cycle in chloroplasts: scavenging of active oxygens and dissipation of excess photons. Annu Rev Plant Phys Plant Mol Biol 50:601–639 Baerlocher MO, Campbell DA, Ireland RJ (2004) Developmental progression of photosystem II electron transport and CO2 uptake in Spartina alterniflora, a facultative halophyte, in a northern salt marsh. Can J Bot 82:365–375 Berry JA, Downton WJS (1982) Environmental regulation of photosynthesis. In: Govindjee (ed) photosynthesis, vol 2. Academic Press, London, pp 263–343 Bjo¨rkman O, Demmig B (1987) Photon yield of O2 evolution and chlorophyll fluorescence characteristics at 77 K among vascular plants of diverse origins. Planta 170:489–504 Brodribb T (1996) Dynamics of changing intercellular CO2 concentration (ci) during drought and determination of minimum functional ci. Plant Physiol 111:179–185 Cheng L (2003) Xanthophyll cycle pool size and composition in relation to the nitrogen content of apple leaves. J Exp Bot 54:385–393 Cheng L, Fuchigami LH, Breen PJ (2001) The relationship between photosystem II efficiency and quantum yield for CO2 assimilation is not affected by nitrogen content in apple leaves. J Exp Bot 52:1865–1872 Cornic G, Briantais JM (1991) Partitioning of photosynthetic electron flow between CO2 and O2 reduction in a C3 leaf (Phaseolus vulgaris L.) at different CO2 concentrations and during drought stress. Planta 183:178–184 D’Ambrosio N, Arena C, Virzo de Santo A (2003) Different relationship between electron transport and CO2 assimilation in two Zea mays cultivars as influenced by increasing irradiance. Photosynthetica 41:489–495 D’Ambrosio N, Arena C, Virzo de Santo A (2006) Temperature response of photosynthesis, excitation energy dissipation and alternative electron sinks to carbon assimilation in Beta vulgaris L. Environ Exp Bot 55:248–257 Demmig-Adams B, Adams WWIII, Barker DH, Logan BA, Bowlong DR, Verhoeven AS (1996) Using chlorophyll fluorescence to assess the fraction of absorbed light allocated to thermal dissipation of excess excitation. Physiol Plant 98:253–264

123

Acta Physiol Plant (2014) 36:849–857 Ding L, Wang KJ, Jiang GM, Biswas DK, Xu H, Li LF, Li YH (2005) Effects of nitrogen deficiency on photosynthetic traits of maize hybrids released in different years. Ann Bot 96: 925–930 Earl HJ, Tollenaar M (1998) Relationship between thylakoid electron transport and photosynthetic CO2 uptake in leaves of three maize (Zea mays L.) hybrids. Photosynth Res 58:245–257 Edwards GE, Baker NR (1993) Can CO2 assimilation in maize leaves be predicted accurately from chlorophyll fluorescence analysis? Photosynth Res 37:89–102 Farage PK, Blowers D, Long SP, Baker NR (2006) Low growth temperatures modify the efficiency of light use by photosystem II for CO2 assimilation in leaves of two chilling-tolerant C4 species, Cyperus longus L. and Miscanthus 9 giganteus. Plant Cell Environ 29:720–728 Fryer MJ, Andrews JR, Oxborough K, Blowers DA, Baker NR (1998) Relationship between CO2 assimilation, photosynthetic electron transport, and active O2 metabolism in leaves of maize in the field during periods of low temperature. Plant Physiol 116:571–580 Huang ZA, Jiang DA, Yang Y, Sun JW, Jin SH (2004) Effects of nitrogen deficiency on gas exchange, chlorophyll fluorescence, and antioxidant enzymes in leaves of rice plants. Photosynthetica 42:357–364 Huxman TE, Monson RK (2003) Stomatal responses of C3, C3–C4 and C4 Flaveria species to light and intercellular CO2 concentration: implications for the evolution of stomatal behaviour. Plant Cell Environ 26:313–322 Jiang CY (2007) The chlorophyll fluorescence and leaf reflectance spectra characteristics among sweet potato (Ipomoea batatas (L.) Lam) genotypes with various leaf colors. Master Thesis, Dept. of Life Sciences, National Chung-Hsing Univ., Taiwan Kakani VG, Surabhi GK, Reddy KR (2008) Photosynthesis and fluorescence responses of C4 plant Andropogon gerardii acclimated to temperature and carbon dioxide. Photosynthetica 46:420–430 Kato MC, Hikosaka K, Hirotsu N, Makino A, Hirose T (2003) The excess light energy that is neither utilized in photosynthesis nor dissipated by photoprotective mechanisms determines the rate of photoinactivation in photosystem II. Plant Cell Physiol 44:318–325 Khamis S, Lamaze T, Lemoine Y, Foyer C (1990) Adaptation of the photosynthetic apparatus in maize leaves as a result of nitrogen limitation: relationships between electron transport and carbon assimilation. Plant Physiol 94:1436–1443 Kosćielniak J, Biesaga-Kosćielniak J (2006) Photosynthesis and nonphotochemical excitation quenching components of chlorophyll excitation in maize and field bean during chilling at different photon flux density. Photosynthetica 44:174–180 Krall JP, Edwards GE (1990) Quantum yields of photosystem II electron transport and carbon dioxide fixation in C4 plants. Aust J Plant Physiol 17:579–588 Krall JP, Edwards GE (1992) Relationship between photosystem II activity and CO2 fixation in leaves. Physiol Plant 86:180–187 Kubien DS, von Caemmerer S, Furbank RT, Sage RF (2003) C4 photosynthesis at low temperature. A study using transgenic plants with reduced amounts of Rubisco. Plant Physiol 132:1577–1585 Kumagai E, Araki T, Ueno O (2010) Comparison of susceptibility to photoinhibition and energy partitioning of absorbed light in photosystem II in flag leaves of two rice (Oryza sativa L.) cultivars that differ in their responses to nitrogen-deficiency. Plant Prod Sci 13:11–20 Lawlor DW (2002) Carbon and nitrogen assimilation in relation to yield: mechanisms are the key to understanding production systems. J Exp Bot 53:773–787

Acta Physiol Plant (2014) 36:849–857 Loriaux SD, Avenson TJ, Welles JM, McDermitt DK, Eckles RD, Riensche B, Genty B (2013) Closing in on maximum yield of chlorophyll fluorescence using a single multiphase flash of subsaturating intensity. Plant Cell Environ 36:1755–1770 Maherali H, Johnson HB, Jackson RB (2003) Stomatal sensitivity to vapour pressure difference over a subambient to elevated CO2 gradient in a C3/C4 grassland. Plant Cell Environ 26:1297–1306 Makino A, Osmond B (1991) Effects of nitrogen nutrition on nitrogen partitioning between chloroplasts and mitochondria in pea and wheat. Plant Physiol 96:355–362 Maxwell K, Johnson GN (2000) Chlorophyll fluorescence—a practical guide. J Exp Bot 51:659–668 Miyake C, Okamura M (2003) Cyclic electron flow within PSII protects PSII from its photoinhibition in thylakoid membranes from Spinach chloroplasts. Plant Cell Physiol 44:457–462 Mohotti AJ, Lawlor DW (2002) Diurnal variation of photosynthesis and photoinhibition in tea: effects of irradiance and nitrogen supply during growth in the field. J Exp Bot 53:313–322 Morison JIL, Gifford RM (1983) Stomatal sensitivity to carbon dioxide and humidity: a comparison of two C3 and two C4 grass species. Plant Physiol 71:789–796 Naidu SL, Long SP (2004) Potential mechanisms of low-temperature tolerance of C4 photosynthesis in Miscanthus 9 giganteus: an in vivo analysis. Planta 220:145–155 Netto AT, Campostrini E, de Oliveira JG, Bressan-Smith RE (2005) Photosynthetic pigments, nitrogen, chlorophyll a fluorescence and SPAD-502 readings in coffee leaves. Sci Hort 104:199–209 Oberhuber W, Edwards GE (1993) Temperature dependence of the linkage of quantum yield of photosystem II to CO2 fixation in C4 and C3 plants. Plant Physiol 101:507–512 Pittermann J, Sage RF (2000) Photosynthetic performance at low temperature of Bouteloua gracilis Lag., a high-altitude C4 grass from the Rocky Mountains, USA. Plant Cell Environ 23:811–823 Pittermann J, Sage RF (2001) The response of the high altitude C4 grass Muhlenbergia montana (Nutt.) A.S. Hitchc. to long-and short-term chilling. J Exp Bot 52:829–838 Ripley BS, Gilbert ME, Ibrahim DG, Osborne CP (2007) Drought constraints on C4 photosynthesis: stomatal and metabolic limitations in C3 and C4 subspecies of Alloteropsis semialata. J Exp Bot 58:1351–1363 Robinson JM (1990) Nitrite photoreduction in vivo is inhibited by oxygen. Plant Physiol 92:862–865 Rohaćˇek K, Barta´k M (1999) Technique of the modulated chlorophyll fluorescence: basic concepts, useful parameters, and some applications. Photosynthetica 37:339–363

857 Romanowskaa E, Anna Dro_zaka A, Pokorskaa B, Shiellb BJ, Wojtek P, Michalskib WP (2006) Organization and activity of photosystems in the mesophyll and bundle sheath chloroplasts of maize. J Plant Physiol 163:607–618 Verhoeven AS, Demmig-Adams B, Adams WWIII (1997) Enhanced employment of the xanthophyll cycle and thermal energy dissipation in spinach exposed to high light and N stress. Plant Physiol 113:817–824 Weng JH, Chen CY (1987) Differences between Indica and Japonica rice varieties in CO2 exchange rates in response to leaf nitrogen and temperature. Photosynth Res 14:171–178 Weng JH, Hsu FH (2001) Gas exchange and epidermal characteristics of Miscanthus populations in Taiwan varying with habitats and nitrogen application. Photosynthetica 39:35–41 Wong SL, Chen CW, Huang HW, Weng JH (2012) Using combined measurements of gas exchange and chlorophyll fluorescence to investigate the photosynthetic light responses of plant species adapted to different light regimes. Photosynthetica 50:206–214 Yin X, Struik PC (2012) Mathematical review of the energy transduction stoichiometries of C4 leaf photosynthesis under limiting light. Plant Cell Environ 35:1299–1312 Yin X, Sun Z, Struik PC, Gu J (2011a) Evaluating a new method to estimate the rate of leaf respiration in the light by analysis of combined gas exchange and chlorophyll fluorescence measurements. J Exp Bot 62:3489–3499 Yin X, Sun Z, Struik PC, van der Putten PEL, van Ieperen W, Harbinson J (2011b) Using a biochemical C4 photosynthesis model and combined gas exchange and chlorophyll fluorescence measurements to estimate bundle-sheath conductance of maize leaves differing in age and nitrogen content. Plant Cell Environ 34:2183–2199 Yong JWH, Wongs SC, Farquhar GD (1997) Stomatal responses to changes in vapour pressure difference between leaf and air. Plant Cell Environ 20:1213–1216 Zhao D, Reddy KR, Kakani VG, Reddy VR (2005) Nitrogen deficiency effects on plant growth, leaf photosynthesis, and hyperspectral reflectance properties of sorghum. Europ J Agron 22:391–403 Ziska LH, Bunce JA (1997) Influence of increasing carbon dioxide concentration on the photosynthetic and growth stimulation of selected C4 crops and weeds. Photosynth Res 54:199–208

123