Present address: Departments of Crop Sciences and Plant Biology, University of Illinois, Urbana, IL 61801, USA.
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 × giganteus
Article first published online: 1 DEC 2005
Plant, Cell & Environment
Volume 29, Issue 4, pages 720–728, April 2006
How to Cite
FARAGE, P. K., BLOWERS, D., LONG, S. P. and BAKER, N. R. (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 × giganteus. Plant, Cell & Environment, 29: 720–728. doi: 10.1111/j.1365-3040.2005.01460.x
- Issue published online: 1 DEC 2005
- Article first published online: 1 DEC 2005
- Received 6 July 2005; received in revised form 15 September 2005; accepted for publication 4 October 2005
- chlorophyll fluorescence;
- electron transport;
- gas exchange;
Two C4 plants, Miscanthus × giganteus and Cyperus longus L., were grown at suboptimal growth temperatures and the relationships between the quantum efficiencies of photosynthetic electron transport through photosystem II (PSII) (PSII operating efficiency; Fq′/Fm′) and CO2 assimilation (φCO2) in leaves were examined. When M. × giganteus was grown at 10 °C, the ratio of the PSII operating efficiency to φCO2 increased relative to that found in leaves grown at 14 and 25 °C. Similar increases in the Fq′/Fm′ : φCO2 occurred in the leaves of two C. longus ecotypes when the plants were grown at 17 °C, compared to 25 °C. These elevations of Fq′/Fm′ : φCO2 at low growth temperatures were not attributable to the development of anthocyanins, as has been suggested for maize, and were indicative of the operation of an alternative sink to CO2 assimilation for photosynthetic reducing equivalents, possibly oxygen reduction via a Mehler reaction, which would act as a mechanism for protection of PSII from photoinactivation and damage. Furthermore, in M. × giganteus grown at 10 °C, further protection of PSII was effected by a 20-fold increase in zeaxanthin content in dark-adapted leaves, which was associated with much higher levels of non-photochemical quenching of excitation energy, compared to that observed in leaves grown at 14 and 25 °C. These differences may explain the long growing season and remarkable productivity of this C4 plant in cool climates, even in comparison to other C4 species such as C. longus, which occur naturally in such climates.
light-saturated rate of CO2 assimilation
intercellular CO2 concentration in leaf
fluorescence level at any point between Fo′ and Fm′
maximal fluorescence level from leaves in light
minimal fluorescence level of leaves in light
difference in fluorescence between Fm′ and F′ (Fq′= Fm′ − F′)
variable fluorescence level of leaves in light (Fv′= Fm′ − Fo′)
photosystem II (PSII) operating efficiency (the quantum yield of PSII photochemistry for a leaf in light)
the PSII efficiency factor which relates to the ability to maintain PSII reaction centres in the open state
PSII maximum efficiency which estimates the maximum quantum efficiency of PSII under given light conditions
quantum yield of CO2 assimilation
Many C4 plants have high rates of photosynthesis and productivity, but these are normally realized only in high light and in humid, warm environments (Long 1999; Sage 1999). C4 species are relatively rare in cool temperate environments; their abundance being negatively correlated with latitude (Doliner & Jollife 1979; Teeri, Stowe & Livingstone 1980) and altitude (Tieszen et al. 1979; Rundel 1980). Chill-induced decreases in CO2 assimilation in C4 leaves have been associated with poor development of the photosynthetic apparatus (Nie & Baker 1991; Nie, Long & Baker 1992; Nie et al. 1995), decreases in the activities of Benson-Calvin cycle enzymes (Kingston-Smith et al. 1997) and photoinhibition of photosynthesis, involving both increased dissipation of excitation energy in the photosystem II (PSII) antennae and photodamage to PSII reaction centres (Ortiz-Lopez et al. 1990; Andrews, Fryer & Baker 1995; Fryer et al. 1995; Haldimann, Fracheboud & Stamp 1996). Under stress conditions, such as low temperatures, that reduce the capacity to assimilate carbon, photosynthetic electron flux from photosystem I (PSI) to oxygen can increase and result in increased production of potentially damaging superoxide, hydrogen peroxide and hydroxyl radicals (Asada 1999; Ort & Baker 2002). This would be consistent with the observation that maize leaves developing at low temperatures in the field exhibit large increases in the ratio of the quantum efficiencies of photosynthetic electron transport and CO2 assimilation compared to leaves grown at optimal growth temperatures (Fryer et al. 1998). Increasing the rate of oxygen reduction by PSI would serve to maintain non-cyclic photosynthetic electron transport and help to prevent total reduction of PSII electron acceptors, thus limiting photodamage to PSII reaction centres (Ort & Baker 2002). However, the increased production of reactive oxygen species resulting from photoreduction of oxygen that would occur in C4 leaves at low temperatures is potentially extremely damaging to thylakoid membrane components (Asada 1999). Levels of enzymes that can scavenge reactive oxygen species increase in Zea species at chilling temperatures, and this could be an important response of the leaves to minimize damage to thylakoid membranes (Jahnke, Hull & Long 1991; Massacci et al. 1995; Hodges et al. 1997; Fryer et al. 1998).
Two rhizomatous C4 perennials, the grass Miscanthus× giganteus and the sedge Cyperus longus L., grow well in cool, wet climates such as found in the UK. M. × giganteus, although a member of the Andropogoneae tribe, which includes sorghum and sugar cane, has been shown to maintain high quantum yields of CO2 assimilation under chilling conditions during the early growing season in the UK (Beale, Bint & Long 1996), achieving efficiencies of radiant energy conversion into biomass comparable to those reported for C4 species in warmer climates (Beale & Long 1995). C. longus exhibits rates of CO2 assimilation at temperatures below 10 °C that are comparable to those of C3 plants (Jones, Hannon & Coffey 1981) and was found to achieve similar productivity to C3 plants in eastern Ireland (Collins & Jones 1986). A previous study showed that growth at 14 °C had little effect on the photosynthetic potential of M. × giganteus relative to 25 °C, in sharp contrast to Zea mays (Naidu & Long 2004). In the present study, the effects of reduced growth and measuring temperatures on the photosynthetic characteristics of M. × giganteus and two C. longus ecotypes originating from regions with contrasting climates (Cornwall, UK and Trieste, Italy) are examined in the context of the relationship between PSII electron transport and CO2 assimilation.
MATERIALS AND METHODS
Plant material and growth conditions
The rhizomes of M. × giganteus originating from a single clone (Picoplant, Oldenburg, Germany) were planted in peat-based compost (F2, Levington Horticultural Ltd, Ipswich, UK) and grown in controlled environment chambers (Fitotron SGC066.CHX, Sanyo Gallenkamp PLC, Leicester, UK). Fertilization was provided by irrigating with Hoagland's nutrient solution (Arnon & Hoagland 1940). Day/night temperatures were either 25 °C/20 °C, 14 °C/12 °C or 10 °C/8 °C (hereafter referred to as plants grown at 25 °C, 14 °C or 10 °C). The water vapour pressure deficit was kept below 1 kPa. Photon flux density at leaf height was 700 µmol m−2 s−1 and photoperiod was 14 h.
Rhizomes of the Cornish and Trieste ecotypes of C. longus L. were grown essentially as described above for M. × giganteus, except that they were planted in cocofibre compost (Sinclair Horticulture, Lincoln, UK) and grown at day/night temperatures of 25 °C/20 °C or 17 °C/15 °C (hereafter referred to as plants grown at 25 °C or 17 °C). The Cornish ecotype was originally collected as rhizomes from Cornwall, UK (50°39′N; 4°07′W) and the Trieste ecotype collected as seed from plants in Trieste, Italy (45°,39′N; 13°46′E). Stands of both ecotypes were established outdoors in an experimental garden at the University of Essex in Colchester, UK and provided rhizomes for planting in the controlled environment studies.
The youngest, fully expanded leaves from both species, as judged by ligule emergence, from a minimum of three plants were used for all measurements.
Chlorophyll fluorescence and gas exchange
For leaves of M. × giganteus, rates of CO2 assimilation were measured with an open gas exchange system incorporating infrared CO2 and water vapour analysers (LI-6400, Li-Cor Inc, Lincoln, NE, USA). The water vapour pressure deficit was < 1 kPa and atmospheric CO2 concentration was 360 µmol mol−1. The leaf was illuminated by either a quartz halogen light source or red light-emitting diodes (656–680 nm; 6400-02, Li-Cor Inc.). CO2 assimilation by C. longus leaves was measured in an open gas exchange system incorporating infrared CO2 and water vapour analysers (Series 225/3, ADC BioScientific Ltd, Hoddesdon, UK) and a xenon light source (Xenophot, Osram, Munich, Germany). The φCO2 at any given photosynthetically active photon flux density (PPFD) was estimated by dividing the rate of CO2 assimilation (corrected for respiratory losses) by the rate at which quanta were absorbed (see below). Maximum quantum yields were determined from the linear region of the light response curve between PPFDs of 50 and 150 µmol m−2 s−1.
Chlorophyll fluorescence was measured simultaneously with the gas exchange measurements for leaves using a modulated fluorimeter (for M. × giganteus, an FMS1 instrument, Hansatech Ltd, Kings Lynn, UK; for C. longus, a PAM-2000 instrument, Heinz Walz GmbH, Effeltrich, Germany) essentially as described by Genty, Briantais & Baker (1989). The fluorescence parameters F′, Fm′, Fo′, Fq′ and Fv′ were determined as previously described (Baker & Rosenqvist 2004) and the ratios Fq′/Fm′, Fq′/Fv′ and Fv′/Fm′ were calculated.
Leaves were harvested at the end of the night period prior to being exposed to light. Pigments were extracted and separated chromatographically as previously described (Fryer et al. 1995).
M. × giganteus
The responses of CO2 assimilation to increasing PPFD in M. × giganteus leaves grown at day temperatures of 10, 14 and 25 °C when measured at 10 and 20 °C are shown in Fig. 1. Lowering the growth temperature from 25 to 14 °C had little effect on the photosynthetic response at both measuring temperatures, confirming previous reports of Naidu et al. (2003); Naidu & Long (2004). However, growth at 10 °C resulted in a marked reduction in both the maximum φCO2(Table 1) and the light-saturated rate of CO2 assimilation (Asat) when measured at 20 °C. This decrease in Asat with reduced growth temperature was unlikely to be attributable to stomatal effects as the mean ci in leaves grown at 10 and 25 °C were 52 and 57 µmol mol−1, respectively. When the measuring temperature was reduced from 20 to 10 °C, large decreases in Asat occurred in leaves grown at 14 and 25 °C (Fig. 1), which were not due to stomatal limitations as ci increased in both cases (data not shown). However, the reduction in Asat observed in leaves grown at 10 °C upon lowering of the measuring temperature was considerably less (Fig. 1). Lowering the measuring temperature had little effect on the maximum quantum yields of all leaves (Fig. 1).
|10 °C||14 °C||25 °C|
|Maximum φCO2||0.048 ± 0.004||0.057 ± 0.002||0.061 ± 0.004|
|Leaf absorptance (450–700 nm)||0.859 ± 0.01||0.808 ± 0.019||0.833 ± 0.011|
|Leaf absorptance (656–680 nm)||0.862 ± 0.02||0.882 ± 0.024||0.891 ± 0.013|
The relationship between linear electron transport and CO2 assimilation in these leaves was examined over the range of PPFDs used to determine the photosynthetic light-response curves by measuring the fluorescence parameter Fq′/Fm′, which estimates the operating efficiency of PSII electron transport (Genty et al. 1989), and the φCO2. Fq′/Fm′ has previously often been termed as φPSII or ΦPSII; however, this implies that it is an equivalent quantum efficiency of that for CO2 assimilation, which it is not. The φPSII or ΦPSII refers to the quantum yield for PSII photochemistry of photosynthetically active photons that are absorbed only by PSII and its associated antennae, and not by the leaf as is the case for φCO2 (Baker et al. 2001). Plots of Fq′/Fm′ against φCO2 for leaves grown at 10, 14 and 25 °C and measured at 10 and 20 °C are shown in Fig. 2. The relationships between Fq′/Fm′ and φCO2 were similar for leaves grown at 14 and 25 °C and measured at 10 and 20 °C. However, for leaves grown at 10 °C, this relationship was modified with increases in Fq′/Fm′ relative to φCO2 occurring over the PPFD range; this phenomenon was more apparent when the measurements were made at 20 °C compared to 10 °C. This change in the relationship between PSII electron transport and CO2 assimilation is similar to that previously reported in maize leaves which had grown at lower temperatures, and it has been attributed to an increased linear electron flow to an alternative electron acceptor, probably O2, relative to electron flow associated with CO2 assimilation (Fryer et al. 1998).
It has been suggested that accumulation of high levels of anthocyanins in leaves when plants are grown at low temperature can result in increases in the ratio of Fq′/Fm′ : φCO2 due to absorption of photosynthetically active radiation (PAR) between 500 and 640 nm by the anthocyanins (Pietrini & Massacci 1998). The absorption spectra of leaves grown at 10, 14 and 25 °C are shown in Fig. 3. Leaves grown at 10 °C were reddish-purple in colour and exhibited substantially increased absorptance between 520 and 600 nm, compared to leaves grown at 14 and 25 °C. However, for leaves grown at 10 °C and, to a lesser extent, leaves grown at 14 °C, absorptance between 630 and 680 nm was lower than for leaves grown at 25 °C. This can be attributed to the much lower chlorophyll contents (see Table 2). Consequently, total absorptances of PAR between 450 and 700 nm for leaves grown at 10, 14 and 25 °C were not markedly different (Table 1). In order to avoid any effects of anthocyanins on the Fq′/Fm′ : φCO2 ratio, measurements of Fq′/Fm′ and φCO2 were made using red light-emitting diodes whose major emission is between 656 and 680 nm. Only very small differences were found between the absorptances of leaves grown at 10, 14 and 25 °C for radiation between 656 and 680 nm (Table 1); consequently, the observed increases in Fq′/Fm′ : φCO2 induced by growth at 10 °C shown in Fig. 2 cannot be attributed to the chill-induced accumulation of anthocyanins. When Fq′/Fm′ : φCO2 was determined using PAR between 400 and 700 nm, the differences between leaves grown at 10 °C, compared to those grown at 14 and 25 °C were greater (data not shown). However, such increases cannot be attributed totally to changes in the relationship between electron transport and CO2 assimilation due to anthocyanins strongly absorbing some of the PAR in the leaves grown at 10 °C, but not in those grown at 14 and 25 °C.
|Pigment (µmol m−2)||Growth temperature|
|10 °C||14 °C||25 °C|
|Violaxanthin||4.77 ± 1.21||10.58 ± 1.29||10.22 ± 0.68|
|Antheraxanthin||4.42 ± 6.2||3.44 ± 4.8||5.8 ± 2.0|
|Zeaxanthin||16.78 ± 0.77||2.80 ± 0.37||0.83 ± 0.54|
|Neoxanthin||3.19 ± 0.29||3.18 ± 0.67||4.47 ± 0.12|
|β-carotene||8.38 ± 0.70||12.30 ± 1.30||17.12 ± 0.64|
|Lutein||19.48 ± 1.32||16.61 ± 1.54||20.87 ± 0.74|
|Total carotenoids||57.02 ± 3.45||48.91 ± 4.15||46.03 ± 0.93|
|Total chlorophyll||294 ± 17||420 ± 34||658 ± 46|
The Fq′/Fm′ is the product of Fv′/Fm′ and Fq′/Fv′ (Genty et al. 1989). Fv′/Fm′ is the maximum efficiency of PSII when all the PSII reaction centres are ‘open’ and is inversely related to non-photochemical quenching; Fq′/Fv′ is the PSII efficiency factor that relates the PSII maximum efficiency to the operating PSII efficiency and mathematically equates to the photochemical quenching coefficient, qP (Baker et al. 2001; Baker & Rosenqvist 2004). At a measuring temperature of 20 °C, the changes in Fv′/Fm′ and Fq′/Fv′ that occur as Fq′/Fm′ decreases with increasing PPFD were similar for leaves grown at 10, 14 and 25 °C (Fig. 4). In all leaves, the light-induced decreases in Fq′/Fm′ were accompanied by decreases in Fv′/Fm′ and Fq′/Fv′, indicating that light-dependent decreases in PSII efficiency were due to both increases in non-photochemical quenching and decreased photochemical quenching, as would be expected. Upon lowering the measuring temperature of the leaves to 10 °C, differences in the relationships between Fv′/Fm′, Fq′/Fv′ and Fq′/Fm′ were observed (Fig. 5) compared to the relationships observed at 20 °C (Fig. 4). At 10 °C, the initial decline in Fq′/Fm′, induced by increases in PPFD from 50 to 150 µmol m−2 s−1, was accompanied by decreases in both Fv′/Fm′ and Fq′/Fv′ for leaves grown at all temperatures, as was the case when leaves were measured at 20 °C. However, above 150 µmol m−2 s−1, light-induced decreases in Fq′/Fm′ of leaves grown at 14 and 25 °C were attributable primarily to decreases in Fq′/Fv′, with only small decreases in Fv′/Fm′ being observed (Fig. 5), indicating that decreases in the operating PSII efficiency were due primarily to decreases in the ability of the leaves to utilize the products of linear electron transport. This was not the case for leaves grown at 10 °C, where large decreases in Fv′/Fm′ continued to occur between 150 and 350 µmol m−2 s−1. Increasing PPFD above 350 µmol m−2 s−1 induced little change in Fv′/Fm′ and the light-induced decreases in Fq′/Fm′ were due primarily to decreases in Fq′/Fv′, as was occurring in the leaves grown at 14 and 25 °C in PPFDs above 150 µmol m−2 s−1.
The low Fq′/Fm′ in maize leaves grown at 14 °C have been associated with high levels of non-photochemical quenching and zeaxanthin content (Fryer et al. 1995). Analyses of carotenoid contents of M. × giganteus leaves at the end of the night period indicated that reduction of growth temperature resulted in an increase in total carotenoid content, which was primarily attributable to very large increases in antheraxanthin and zeaxanthin (Table 2). Although decreases in violaxanthin, neoxanthin and β-carotene contents occurred as growth temperature was reduced, leaves grown at 10 °C exhibited a 20-fold greater zeaxanthin content than the leaves grown at 25 °C (Table 2). The total xanthophyll cycle pigment content (violaxanthin + antheraxanthin + zeaxanthin) of leaves grown at 10 °C was 2.2-fold greater than that of leaves grown at 25 °C. Consequently, a considerably greater proportion of the xanthophyll cycle pigments is maintained in the dark throughout the night period as zeaxanthin in the leaves grown at 10 °C, which is consistent with the increased capacity for non-photochemical quenching in these leaves.
The responses of CO2 assimilation to increasing PPFD in leaves of the Cornish and Trieste ecotypes of C. longus grown at 17 and 25 °C and measured at 14 and 25 °C are shown in Fig. 6. No significant differences were observed in the maximum φCO2 for leaves at each growth temperature. At PPFDs above 400 µmol m−2 s−1, the Cornish ecotype exhibited greater CO2 assimilation rates irrespective of growth or measuring temperature, and reducing the measuring temperature from 25 °C to 14 °C significantly reduced the CO2 assimilation rates of both ecotypes. In contrast to M. × giganteus where growth at 14 °C had little effect on photosynthetic capacity relative to 25 °C, growth of both ecotypes of C. longus at 17 °C significantly lowered the light-saturated and light-limited rates of CO2 uptake relative to plants grown at 25 °C.
The relationships between Fq′/Fm′ and φCO2 for leaves of the two ecotypes grown at 17 and 25 °C and measured at 14 and 25 °C over the range of PPFDs used to determine the photosynthetic light-response are shown in Fig. 7. The relationships between PSII electron transport and CO2 assimilation were similar in all leaves grown at 25 °C, irrespective of measuring temperature or ecotype. However, different relationships were observed for leaves grown at 17 °C, with increases in Fq′/Fm′ relative to φCO2 occurring. The shift in this relationship in favour of Fq′/Fm′ is similar to that observed for M. × giganteus grown at 10 °C compared to 14 and 25 °C (Fig. 2) and that reported previously when maize leaves experience low growth temperatures (Fryer et al. 1998). Only very small differences (less than 4%) were observed in the absorptances of the leaves of each ecotype when grown at the two different temperatures (data not shown); consequently, the shift in Fq′/Fm′ relative to φCO2 when the leaves are grown at 17 °C, compared to 25 °C, cannot be attributed to accumulation of large amounts of anthocyanins.
The changes in Fv′/Fm′ and Fq′/Fv′ as Fq′/Fm′ decreases with increasing PPFD for leaves of the Trieste ecotype grown at 17 and 25 °C and measured at 14 and 25 °C are shown in Fig. 8. The relationships between Fv′/Fm′, Fq′/Fv′ and Fq′/Fm′ were similar for all leaves irrespective of growth or measuring temperature, with increases in both non-photochemical quenching and the inability to utilize the products of electron transport both contributing substantially to the decreases in the Fq′/Fm′ over the whole range of PPFDs. Similar results were obtained from leaves of the Cornish ecotype (data not shown).
Although both the C. longus Cornish ecotype and M. × giganteus are from habitats near the low temperature limits of the natural distributions of C4 plants, photosynthetic capacity in M. × giganteus is clearly far more tolerant of low temperature (Long 1999). The decrease in growth temperature from 25 to 17 °C markedly lowered the CO2 uptake rate of leaves of C. longus at all measurement temperatures and light levels, whereas the decrease in growth temperature from 25 to 14 °C in M. × giganteus had little effect. Both M. × giganteus and C. longus exhibit changes in the relationship between photosynthetic electron transport and CO2 assimilation when grown at low temperatures. Growth of M. × giganteus at 10 °C and of C. longus at 17 °C resulted in an increase in the ratio of Fq′/Fm′ to φCO2 (Figs 2 & 7), indicating that the quantum efficiency of electron flux through PSII relative to the quantum efficiency of CO2 assimilation had increased with reduced growth temperature and that linear electron transport to alternative electron acceptors other than CO2 must have increased. This phenomenon is similar to that previously reported for maize leaves growing at low temperatures in the field (Fryer et al. 1998), where it was suggested that oxygen reduction via a Mehler reaction was most probably the alternative electron sink to CO2 assimilation. It has been suggested that increases in the ratio of Fq′/Fm′ to φCO2 may be due to the low temperature stress-induced accumulation of anthocyanins, which increase absorption of PAR between 500 and 640 nm with a resultant lowering of φCO2 and increase in Fq′/Fm′ (Pietrini & Massacci 1998). Although M. × giganteus does accumulate anthocyanins at low growth temperatures, our measurements of Fq′/Fm′ and φCO2 were made using PAR between 656 and 680 nm, where anthocyanins do not absorb significantly. Neither C. longus nor the two maize cultivars LG11 and LG20.80, which were used in the previous field studies, exhibited any noticeable accumulation of anthocyanins when grown at low temperatures (data not shown). Consequently, the enhancement of linear electron flux through PSII relative to CO2 assimilation in M. × giganteus, C. longus and Z. mays might suggest that this is a characteristic acclimatory response of C4 plants, using the NADP-malic enzyme for decarboxylation, to low growth temperatures. It is evident that this phenomenon cannot be rapidly induced in leaves grown at moderate growth temperatures by simply exposing them to low temperatures and is associated with development and growth of leaves at low temperatures. In a maize crop, increases in the Fq′/Fm′ : φCO2 ratio during growth of leaves at low temperatures experienced in the field during the early growing season, compared to midsummer temperatures, was accompanied by an elevation of the activities of a range of enzymes (ascorbate peroxidase, glutathione reductase, monodehydroascorbate reductase, dehydroascorbate reductase and superoxide dismutase) and antioxidants (ascorbate and α-tocopherol) involved in the scavenging of reactive oxygen species (Fryer et al. 1998). Similarly, the activities of ascorbate peroxidase and superoxide dismutase and the ascorbate content of leaves of C. longus were also considerably increased when plants were grown at 13 °C compared to 25 °C (Blowers 1997). Such elevation of activities of enzymes and antioxidants would be consistent with the suggestion that the elevated Fq′/Fm′ : φCO2 ratio is due to increased electron flux to oxygen via a Mehler reaction, which would generate superoxide and hydrogen peroxide (Fryer et al. 1998). Increasing electron flux to oxygen in circumstances where CO2 assimilation is restricted will potentially allow the PSII electron acceptors to be maintained in a partially oxidized state, minimizing the possibility of photoinactivation and damage to PSII. However, this strategy can only be successful if the reactive oxygen species generated by the reduction of oxygen are rapidly scavenged, otherwise they will damage the thylakoid membrane and other cellular components.
The enhancement of Fq′/Fm′ : φCO2 that is observed with the growth of the C4 leaves at low temperatures is not dependent upon any modification to the relationships between Fv′/Fm′, Fq′/Fv′ and Fq′/Fm′. For M. × giganteus grown at 10, 14 and 25 °C and measured at 20 °C (Fig. 4) and grown at 14 and 25 °C and measured at 10 °C (Fig. 5), and for C. longus grown at 17 and 25 °C and measured at 14 or 25 °C (Fig. 8), the relationships between Fv′/Fm′, Fq′/Fv′ and Fq′/Fm′ were similar. Consequently, the decrease in the efficiency of light use for CO2 assimilation occurring at leaves grown at low temperatures appears not to be associated with any change in the way the PSII operating efficiency is regulated; the relationships between non-photochemical and photochemical quenching remain the same. However, in leaves of M. × giganteus grown and measured at 10 °C a marked change in this relationship is observed (Fig. 5). With increasing PPFD between c. 150–350 µmol m−2 s−1, there are greater decreases in Fv′/Fm′ and higher Fq′/Fv′ than are found in leaves grown at 14 and 25 °C, indicating that for a given Fq′/Fm′ at those PPFDs, the leaves grown at 10 °C have much higher rates of both non-photochemical and photochemical quenching. This increased non-photochemical quenching is presumably associated with increased quenching of excitation energy by zeaxanthin, as it is accompanied by a 20-fold increase in zeaxanthin content in dark-adapted leaves grown at 10 °C compared to leaves grown at 25 °C (Table 1). Non-photochemical quenching of excitation energy by high levels of zeaxanthin has been implicated previously as a major factor involved in the severe depression of photosynthetic efficiency of maize leaves grown at low temperature (Fryer et al. 1995). Intriguingly, when M. × giganteus leaves grown at 10 °C are measured at 20 °C similar increases in the depression of Fv′/Fm′ with increasing PPFD are not observed (Fig. 4), and the changes in Fv′/Fm′ and Fq′/Fv′ are similar to those observed in leaves grown at 14 and 25 °C. It is not clear why the leaves grown at 10 °C have higher rates of non-photochemical quenching at similar PSII operating efficiencies at 10 °C compared to 20 °C. This phenomenon may involve differences in the degree of thylakoid energization and associated energy-dependent quenching of excitation energy in the PSII antennae, and warrants further investigation.
M. × giganteus appears remarkable, even among C4 species native to cool climates, in its ability to grow at a temperature of 10 °C. C4 photosynthesis is potentially more efficient because ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) is isolated to the bundle sheath and is not in direct contact with the atmosphere, but rather receives its CO2 via the ‘CO2 concentrating mechanism’ represented by the photosynthetic C4 dicarboxylate cycle. However, this limits the maximum content of Rubisco that a C4 leaf can support, and this becomes a penalty by severely limiting the rate of carbon uptake at low temperature (Kubien et al. 2003; Kubien & Sage 2004). This restriction will also mean that the leaf is less able to dissipate absorbed excitation energy via carbon metabolism at low temperatures and will be potentially more vulnerable to photoinhibition. At 14 °C, M. × giganteus appears to tolerate low temperature by maintaining or increasing quantities of rate-limiting photosynthetic enzymes (Naidu et al. 2003; Naidu & Long 2004), but at 10 °C photoinhibitory damage is minimized by a massive increase in de-epoxidated xanthophylls and the induction of electron transport to an acceptor(s) other than CO2. This appears to underlie the remarkable capacity of this grass to grow in cool climates and greatly outyield other C4 species at these temperatures (Beale & Long 1995; Heaton, Voigt & Long 2004).
In conclusion, increases in the Fq′/Fm′ : φCO2 ratio in leaves of both M. × giganteus and C. longus when grown at low temperatures, as was previously found to be the case in Z. mays (Fryer et al. 1998), are indicative of a mechanism by which these C4 plants can protect PSII from photoinactivation and damage when the capacity for CO2 assimilation is reduced. The operation of this mechanism is not associated with significant changes in non-photochemical and photochemical processes involved with the quenching of excitation energy. A further photoprotective mechanism, identified in M. × giganteus when grown at 10 °C, but not at 14 °C, was associated with a large increase in the zeaxanthin content of leaves, which was maintained overnight in the dark and was associated with a large increase in the non-photochemical quenching of excitation energy.
This research was supported by funds from the Biotechnology and Biological Sciences Research Council. D.B. was the recipient of a research studentship from the Natural Environment Research Council. We thank Michael Fryer for the advice and assistance with the carotenoid analyses and Sue Corbett for providing a constant supply of M. × giganteus rhizomes.
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