Photorespiration plays an important role in the regulation of photosynthetic electron flow under fluctuating light in tobacco plants grown under full sunlight

Plants usually experience dynamic fluctuations of light intensities under natural conditions. However, the responses of mesophyll conductance, CO2 assimilation, and photorespiration to light fluctuation are not well understood. To address this question, we measured photosynthetic parameters of gas exchange and chlorophyll fluorescence in tobacco leaves at 2-min intervals while irradiance levels alternated between 100 and 1200 μmol photons m−2 s−1. Compared with leaves exposed to a constant light of 1200 μmol photons m−2 s−1, both stomatal and mesophyll conductances were significantly restricted in leaves treated with fluctuating light condition. Meanwhile, CO2 assimilation rate and electron flow devoted to RuBP carboxylation at 1200 μmol photons m−2 s−1 under fluctuating light were limited by the low chloroplast CO2 concentration. Analysis based on the C3 photosynthesis model indicated that, at 1200 μmol photons m−2 s−1 under fluctuating light, the CO2 assimilation rate was limited by RuBP carboxylation. Electron flow devoted to RuBP oxygenation at 1200 μmol photons m−2 s−1 under fluctuating light remained at nearly the maximum level throughout the experimental period. We conclude that fluctuating light restricts CO2 assimilation by decreasing both stomatal and mesophyll conductances. Under such conditions, photorespiration plays an important role in the regulation of photosynthetic electron flow.


Introduction
In nature, plants grown in open habitats usually experience changes in light intensities because of clouds. Even on clear days, the leaves of understory plants are frequently exposed to short-term fluctuating light levels due to movements by the leaves and stems of other plants above them. To cope with fluctuating light conditions, plants must regulate their photosynthetic processes.
Under constant low light, most of the absorbed light energy can be used to drive photosynthesis, even when stomatal conductance (g s ) is reduced. Under constant high light, the rate of CO 2 assimilation (A n ) is maintained at an elevated level due to high g s and mesophyll conductance (g m ) (Yamori et al., 2010(Yamori et al., , 2011. Fluctuating light restricts both g s and CO 2 assimilation rate (Fay and Knapp, 1993;Kirschbaum et al., 1998). However, it is unclear how g m and photorespiration respond to those changes in irradiance.
Under natural conditions, g m is an important determinant of the CO 2 assimilation rate, especially at high light levels (Carriquí et al., 2015). Several environmental factors, such as water status and temperature, can affect g m Scafaro et al., 2011;Walker et al., 2013). For tobacco (Nicotiana tabacum) plants grown with adequate water and optimum temperature, g m is mainly dependent upon the growth light intensity (Yamori et al., 2010). Plants exposed to strong light have higher values of g m when compared with those grown under low light. When light levels are constant, g m does not appear to be dependent upon light intensity (Yamori et al., 2010). However, the effect of fluctuating light condition on g m is unclear. According to the model of Farquhar et al. (1980), CO 2 assimilation in C 3 plants is limited by either the carboxylation or the regeneration of ribulose-1,5-bisphosphate (RuBP). In tobacco, a model C 3 plant, the rate of CO 2 assimilation under high light is influenced by leaf nitrogen (N) content. CO 2 assimilation rate under high light tends to be limited by RuBP regeneration for plants grown at high N concentration (Yamori et al., 2010(Yamori et al., , 2011. However, that presumption is based on high values of g s and g m . Once g s and g m decrease because of environmental stresses such as drought, CO 2 assimilation rate is then partially constrained by RuBP carboxylation . Therefore, if fluctuating light levels restrict g m , then A n likely tends to be limited by RuBP carboxylation.
Photorespiration, an inevitable process in photosynthesis, plays a supporting role in photosynthetic CO 2 assimilation (Timm et al., 2012;Busch et al., 2013;Weber and Bauwe, 2013). This process is initiated by the oxygenation of RuBP, in which one molecule of glycolate-2-phosphate and one molecule of glycerate-3-phosphateare are produced (Ogren, 1984). Although glycolate-2-phosphate cannot be used by plants for biosynthetic reactions, and is also a potential inhibitor of chloroplast functioning (Anderson, 1971), it can be converted into glycerate-3-phosphate through the photorespiratory pathway (Leegood et al., 1995). When g s and g m are diminished, the decreased chloroplast CO 2 concentration increases the specificity of Rubisco to O 2 and then induces a rise in the rate of RuBP oxygenation (Wingler et al., 1999(Wingler et al., , 2000. Plants avoid those detrimental effects of glycolate-2phosphate and other photorespiratory intermediates by activating the photorespiratory pathway when chloroplast CO 2 concentration is low. In Arabidopsis thaliana plants grown under low irradiance, photorespiration plays a minor role in regulating photosynthetic electron flow after exposure to short-term fluctuating light (Kono et al., 2014). The growth light intensity significantly affects the development of the photorespiratory pathway (Huang et al., 2014). For example, plants such as tobacco grown under bright light have a greater capacity than those under low light (Huang et al., 2014). However, little is known about how the photorespiratory pathway functions in the acclimation to fluctuating light by plants grown under high light. Because this pathway is critical to the control of A n and photosynthetic electron flow (Takahashi et al., 2007;Timm et al., 2012;Huang et al., 2014), it is important that research focused on photosynthetic regulation under fluctuating light should include growth light intensity as an experimental variable.
In this study, we measured the photosynthetic parameters of gas exchange and chlorophyll fluorescence to investigate the responses of g m , A n , and photosynthetic electron flow to fluctuations in light levels. We also examined the limiting step of CO 2 assimilation and the role the photorespiratory pathway has in modulating photosynthetic electron flow under alternating light conditions. Our objective was to improve our understanding of how photosynthesis is regulated when sun-grown plants are exposed to changes in irradiance. The following questions were addressed: (1) Is g m restricted by fluctuating light? (2) What is the limiting step of A n under fluctuating light? and (3) Does the photorespiratory pathway play an important role in regulating photosynthetic electron flow under fluctuating light?

Plant Materials and Growing Conditions
Following seed germination, seedlings of tobacco cv. y87 were cultivated in phytotron for 7 weeks. Afterwards, they were grown in plastic pots in an open field at Kunming Institute of Botany, Yunnan, China (elevation 1900 m, 102 • 41 ′ E, 25 • 01 ′ N). During our experiment period (10 May to 24 June 2013), none of the plants experienced any water or nutrient stresses. The average temperature at Kunming was 20.9 • C in May and 20.6 • C in June. Fully expanded mature leaves on 13-week-old plants were used for photosynthetic measurements.

Analyses of Gas Exchange, Chlorophyll Fluorescence, and Mesophyll Conductance
Photosynthetic parameters for gas exchange and chlorophyll fluorescence were monitored with an open gas exchange system that incorporated infrared CO 2 and water vapor analyzers (Li-6400XT; Li-Cor Biosciences, Lincoln, NE, USA) and a 2-cm 2 measuring head (6400-40 Leaf Chamber Fluorometer; Li-Cor Biosciences). Measurements were made in a phytotron where relative air humidity (60%) and air temperature (25 • C) were controlled. The atmospheric CO 2 concentration was maintained at 400 µmol mol −1 by the Li-6400XT. To generate a light response curve, we initially exposed the mature leaves to strong irradiance (2000 µmol photons m −2 s −1 ) for 20 min to obtain steady, high levels of g s and CO 2 assimilation. Afterward, photosynthetic parameters were evaluated at 2-min intervals at photosynthetic photon flux densities (PPFDs) of 2000, 1600, 1200, 800, 500, 300, 200, 100, 50, 20, or 0 µmol photons m −2 s −1 . To investigate the responses of g s , g m , CO 2 assimilation rate, and photosynthetic electron flow to fluctuating light, we also evaluated those photosynthetic parameters under light levels that alternated every 2 min between 100 and 1200 µmol photons m −2 s −1 after dark-adaptation for 30 min. Photosynthetic induction curves were also developed at 1200 µmol photons m −2 s −1 after 30 min of darkness. Values for those parameters were recorded automatically by the Li-6400XT at 2-min intervals.
The CO 2 assimilation rate vs. chloroplast CO 2 concentration (C c ) was examined at 1200 µmol photons m −2 s −1 (von Caemmerer and Farquhar, 1981). For each A n /C c curve, the photosynthetic rate reached a steady state at 400 µmol mol −1 CO 2 , then decreased to a lower limit of 50 µmol mol −1 before increasing stepwise to an upper limit of 1600 µmol mol −1 . Each stepwise measurement was completed within 2-3 min. Using those A n /C c curves, we calculated the maximum rates of RuBP regeneration (J max ) and RuBP carboxylation (V cmax ) according to the method of Long and Bernacchi (2003).
The fluorescence parameters F o ′ , F m ′ , and F s were evaluated as previously described in Baker and Rosenqvist (2004). Here, F o ′ and F m ′ represented the minimum and maximum fluorescence after light-adaption, respectively. F s indicated the light-adapted steady-state fluorescence. The maximum quantum yield of PSII after light adaptation Genty et al., 1989). Total photosynthetic electron flow through PSII was calculated as J T = PSII × PPFD × L abs × 0.5 (Krall and Edwards, 1992), where L abs represented leaf absorbance and was assumed to be 0.85 for sun-grown tobacco leaves that receive high-nitrogen nutrition (Miyake et al., 2005). The constant of 0.5 was applied based on the assumption that photons were equally distributed between photosystem I (PSI) and PSII (Miyake et al., 2005). Following the assumption that the water-water cycle is not a major alternative electron sink when CO 2 assimilation is limited (Driever and Baker, 2011), we allocated the electron flow through PSII to RuBP carboxylation (J C ) and oxygenation (J O ). Values for J C and J O were estimated according to the method of Valentini et al. (1995): where A n was the net rate of CO 2 assimilation and R d represented the rate of mitochondrial respiration as measured after 30 min of dark-adaptation.
We recorded values for mesophyll conductance (g m ) at 1200 µmol photons m −2 s −1 after plants were exposed to either fluctuating or constant light for 60 min. For our comparisons, g m was also estimated at 1200 µmol photons m −2 s −1 in light response curves. Values for g m were estimated through a combination analysis of gas exchange and chlorophyll fluorescence, and according to the following equation Loreto et al., 1992;Warren and Dreyer, 2006;Yamori et al., 2010Yamori et al., , 2011: where A n was the net rate of CO 2 assimilation, C i was the intercellular CO 2 concentration, J T was total photosynthetic electron flow through PSII, R d was the rate of mitochondrial respiration, and Ŵ * was the CO 2 compensation point in the absence of daytime respiration (Farquhar et al., 1980;Brooks and Farquhar, 1985), with the latter assumed to be 32.2 at 25 • C (Long and Bernacchi, 2003). Using the estimated g m , we calculated the chloroplast CO 2 concentration with the following equation (Long and Bernacchi, 2003;Warren and Dreyer, 2006;Yamori et al., 2010Yamori et al., , 2011: where C i was the intercellular CO 2 concentration, A n was the net rate of CO 2 assimilation, and g m was mesophyll conductance. To identify the limiting step of CO 2 assimilation under fluctuating light, we applied the method of Yamori et al. (2010Yamori et al. ( , 2011 to determine C trans , the chloroplast CO 2 concentration at which the transition from RuBP carboxylation to RuBP regeneration occurred: were the Michaelis constants for CO 2 and O 2 , respectively (Farquhar et al., 1980), and were assumed to be 406.7 µmol mol −1 and 277 mmol mol −1 at 25 • C, respectively (Long and Bernacchi, 2003); J max was the maximum rate of RuBP regeneration; V cmax was the maximum rate of RuBP carboxylation; and Ŵ * was the CO 2 compensation point in the absence of daytime respiration. The limiting step of CO 2 assimilation was then determined by comparing the values of C c and C trans .

Statistical Analysis
The results were displayed as mean values of four independent measurements. We used One-Way ANOVA and SPSS 16.0 software (SPSS Inc., Chicago, IL, USA) to examine differences among treatments involving fluctuating vs. constant light. Those differences were considered significant at P < 0.05.
Fluctuating light conditions significantly restricted the opening of stomata. After plants were alternately exposed to 100 and 1200 µmol photons m −2 s −1 every 2 min for 60 min, g s was 0.18 mol m −2 s −1 (Figure 2A). However, when plants were illuminated at a constant 1200 µmol photons m −2 s −1 for 60 min, g s was 0.30 mol m −2 s −1 . After 60 min of fluctuating light, the CO 2 assimilation rate at 1200 µmol photons m −2 s −1 was 18.1 µmol CO 2 m −2 s −1 vs. 25.4 µmol CO 2 m −2 s −1 after exposure to 1200 µmol photons m −2 s −1 for 60 min ( Figure 2B). Those values for g s and A n differed significantly between constant and fluctuating-light treatments, demonstrating that the latter condition inhibited g s as well as CO 2 assimilation. This finding was consistent with those reported previously (Fay and Knapp, 1993;Kirschbaum et al., 1998).
By contrast, values for qP at 1200 µmol photons m −2 s −1 differed only slightly between the constant and fluctuating light treatments (Figure 3A), while F v ′ /F m ′ and PSII at 1200 µmol photons m −2 s −1 were significantly lower under fluctuating light (P < 0.001; Figures 3B,C). The parameter F v ′ /F m ′ represents the maximum efficiency of PSII when all reaction centers are "open, " and qP is the factor that relates maximum PSII efficiency to the operating PSII efficiency (Farage et al., 2006). Because PSII is the product of qP and F v ′ /F m ′ , the difference in PSII that we found between fluctuating light and constant light resulted from the change in F v ′ /F m ′ . These results suggested that although fluctuating light had little effect on the coefficient of PSII photochemical quenching, it induced a significant decline in the maximum efficiency of PSII. After 60 min of treatment, total electron flow through PSII (J T ) at 1200 µmol photons m −2 s −1 was significantly higher under constant illumination than under fluctuating light, i.e.,  (Figure 4C). Consequently, the ratio J O /J C at 1200 µmol photons m −2 s −1 was higher for plants treated with fluctuating light because of the lower value for J C (Figure 4D). These results indicated that fluctuations in irradiance levels suppressed photosynthetic electron flow, primarily by restricting electron flow devoted to RuBP carboxylation. By comparison, electron flow devoted to RuBP oxygenation was hardly affected by fluctuating light conditions.
After pooling the photosynthesis data collected at 1200 µmol photons m −2 s −1 under fluctuating light, we determined that g s was linearly and positively correlated with A n , J T , and J C ( Figures 5A,B). We found it interesting that J O was independent of g s (Figure 5B), which implied that RuBP carboxylation and RuBP oxygenation responded differently to g s . Under fluctuating light, J O remained at nearly the maximum level throughout the experimental period. In the initial stage of fluctuating light treatment, electron flow attributed to RuBP oxygenation contributed largely to the total electron transport through PSII ( Figure 5C).
To analyze the limiting step of CO 2 assimilation under fluctuating light, we examined the relationship between photosynthesis and chloroplast CO 2 concentration. Here, the ratio of the maximum rate of RuBP regeneration (J max ) to that of RuBP carboxylation (V cmax ) was 0.92, and the chloroplast CO 2 concentration at which the transition from RuBP carboxylation to RuBP regeneration occurred (C trans ) was 135 µmol mol −1 (Figure 6). After exposure to fluctuating light conditions for 60 min, g m at 1200 µmol photons m −2 s −1 was 0.21 mol m −2 s −1 , which was significantly lower than that found with light curves (0.29 mol m −2 s −1 ) or under constant light (0.28 mol m −2 s −1 ) (Figure 7). For the light curves, C c at 1200 µmol photons m −2 s −1 was 154 µmol mol −1 . After exposure to fluctuating light or constant light for 60 min, the value for C c was 105 or 131 µmol mol −1 , respectively. This indicated that fluctuating light not only decreased g s but also restricted g m , leading to a decline in C c . Because C c was significantly lower than C trans (P < 0.0001), the rate of CO 2 assimilation at 1200 µmol photons m −2 s −1 under fluctuating light was limited by RuBP carboxylation.
FIGURE 6 | Response of CO 2 assimilation rate (A n ) to incident chloroplast CO 2 concentration (C c ) at 25 • C and 1200 µmol photons m −2 s −1 . Maximum rates of RuBP regeneration (J max ) and RuBP carboxylation (V cmax ) were calculated according to method of Long and Bernacchi (2003). Chloroplast CO 2 concentration at which RuBP carboxylation transitions to RuBP regeneration (C trans ), as determined by method of Yamori et al. (2010); Yamori et al. (2011). Solid symbol, A n at atmospheric CO 2 concentration of 400 µmol mol −1 .
FIGURE 7 | Values for stomatal conductance (g s ), mesophyll conductance (g m ), and chloroplast CO 2 concentration (C c ) at 1200 µmol photons m −2 s −1 analyzed in light response curves, constant vs. fluctuating light levels after 60 min of treatment. Measurements were conducted at 25 • C and 400 µmol mol −1 CO 2 . Values are means ± SE (n = 4). For each treatment type, different letters indicate significant differences among light treatments (P < 0.05), based on Tukey's multiple comparison tests.

Discussion
The limiting step of A n is mainly determined by the relative values of C trans and C c . For tobacco plants supplied with high concentrations of nitrogen, photosynthesis tends to be limited by RuBP regeneration because C c is higher than C trans (Yamori et al., 2010(Yamori et al., , 2011. Such previous conclusions have been based on experiments that involved high levels of g s and g m under constant strong light. Although g s and A n can be significantly inhibited under fluctuating light, the limiting step of A n under such conditions has been unknown. Here, our results indicate that both g s and g m are significantly restricted under fluctuating light, leading to the decrease in C c . After exposure to fluctuating light for 60 min, C c was 105 vs. 135 µmol mol −1 for C trans . Those data provided evidence that, at 1200 µmol photons m −2 s −1 , the photosynthetic process is limited by RuBP carboxylation under fluctuating light. Meanwhile, the high activation of photorespiration contributed largely to the regulation of photosynthetic electron flow. Carriquí et al. (2015) have demonstrated that g m plays an important role in determining the CO 2 assimilation rate, especially at high light intensities. Nevertheless, the response of g m to light intensity remains controversial. For example, in sclerophylls such as Banksia integrifolia, B. serrata, and B. paludosa, the average g m under ambient CO 2 concentration is 22% lower at 500 than at 1500 µmol photons m −2 s −1 (Hassiotou et al., 2009). However, the average g m calculated for wheat leaves is not affected by light intensity (Tazoe et al., 2009). In tobacco leaves, g m is significantly lower at 250 than at 1000 µmol photons m −2 s −1 (Flexas et al., 2007). By contrast, Yamori et al. (2010) have shown that g m differs little between constant high light and constant low light in tobacco leaves. Our results indicated that g m at 1200 µmol photons m −2 s −1 under the fluctuating light was 25% lower than the level calculated at constant light of 1200 µmol photons m −2 s −1 (Figure 7). We believe that this difference was caused by the use of a low light regime (100 µmol photons m −2 s −1 ) in fluctuating light. We found that, under fluctuating light, g m was regulated by both high and low light levels; i.e., although the former induced an increase in g m , this effect could be partially reversed when plants were then exposed to reduced irradiance.
According to the photosynthesis model of Farquhar et al. (1980), CO 2 assimilation in C 3 plants is constrained by RuBP carboxylation and/or RuBP regeneration. Therefore, based on that model, the limiting step can be altered in two ways: (1) adjustments in the balance between the maximum rates of RuBP regeneration and RuBP carboxylation, or (2) changes in the chloroplast CO 2 concentration (Hikosaka et al., 2006;Yamori et al., 2011). For example, in research with tobacco plants, Yamori et al. (2011) have reported that the CO 2 assimilation rate at 380 µmol mol −1 CO 2 and 1500 µmol photons m −2 s −1 (A 380 ) depends upon the leaf-N content and is mainly determined by J max /V cmax . Furthermore, at high leaf-N content, A 380 is limited by RuBP regeneration due to the low ratio of J max /V cmax (Yamori et al., 2010(Yamori et al., , 2011. However, those conclusions have been drawn from experiments with plants that had high values for both g s and g m , and which did not consider the effects of fluctuating light levels. By comparison, our photosynthetic data for g s , A n , J T , and J max /V cmax ratio are very similar to those that describe the performance of plants grown with a high nitrogen supply (Yamori et al., 2011), indicating that plants grown with high N concentration were used in the present study. Furthermore, CO 2 assimilation rate at 1200 µmol photons m −2 s −1 under constant light was limited by RuBP regeneration. When plants were exposed to fluctuating light, the declines in g s and g m resulted in a decrease in C c . The low light regimes under fluctuating light decreased the Rubisco activation state (Yamori et al., 2012), which further restricted the Calvin cycle. The rate of CO 2 assimilation under high light during the fluctuating-light treatment tended to be limited by RuBP carboxylation. Fluctuating light has altered the limiting step of CO 2 assimilation in tobacco plants with high leaf-N content.
Previous studies with A. thaliana have investigated the roles of cyclic electron flow (CEF) and O 2 -dependent alternative electron sinks in regulating photosynthetic electron flow under fluctuating light (Suorsa et al., 2012;Kono et al., 2014). It is believed that CEF is essential for proper acclimation of PSI to such light condition (Suorsa et al., 2012). However, the contribution of photorespiration to photodamage under fluctuating light is small in Arabidopsis leaves sampled from plants exposed to low light (Kono et al., 2014). In tobacco, the capacity of the photorespiratory pathway is strongly influenced by the growth light intensity, with sun leaves up-regulating this pathway to control CO 2 assimilation and photosynthetic electron flow (Huang et al., 2014). However, it is unknown what role the photorespiratory pathway has in enabling plants normally grown under high light to adapt to fluctuating light conditions. Our data demonstrated that, when plants were exposed to fluctuating light, the reduction in C c meant that less electron flow could be devoted to RuBP carboxylation. However, we found that the flow devoted to RuBP oxygenation was completely and highly activated under such conditions. Suppression of CO 2 fixation can cause over-acidification of lumen in the thylakoid membrane, which then activates non-photochemical quenching (NPQ) to dissipate excess light energy harmlessly as heat Takahashi et al., 2007;Huang et al., 2012). At 1200 µmol photons m −2 s −1 , F v ′ /F m ′ were lower under fluctuating light than under constant light. Because F v ′ /F m ′ is inversely related to NPQ, this result was evidence of the higher activation of NPQ under fluctuating light. Furthermore, an increase in the proton gradient across the thylakoid membrane can limit linear electron flow (LEF) via cytochrome b6/f (Tikkanen and Aro, 2014). Consumption of photochemical energy, such as ATP and NADPH, through the photorespiratory pathway is thought to alleviate such over-acidification. Especially in the initial stage of our fluctuating-light period, electron flow that was consumed by the photorespiratory pathway largely contributed to the operation of LEF. Therefore, for tobacco plants grown under full sunlight, photorespiratory pathway would be essential for regulating photosynthetic electron flow under fluctuating light, even though our findings contradict a previous report concerning low-light-grown A. thaliana (Kono et al., 2014).
Although photorespiratory intermediates such as glycine and glycerate inhibit the Calvin cycle (Chastain and Ogren, 1989;Eisenhut et al., 2007;Timm et al., 2012), they can be converted to glycerate-3-phosphate through the photorespiratory pathway (Peterhansel and Maurino, 2011). This process is critical for photosynthesis and photoprotection (Takahashi et al., 2007). Under fluctuating light, a reduction in C c will accelerate RuBP oxygenation and, ultimately, the production of those intermediates. If the photorespiratory pathway is maintained at a low level under such conditions, the accumulation of those intermediates inhibits CO 2 assimilation as well as photosynthetic electron flow, causing acceleration of photodamage (Chastain and Ogren, 1989;Eisenhut et al., 2007;Takahashi et al., 2007). In plants with a high rate of CO 2 assimilation, rapid acceleration of photorespiratory pathway results in low glycine and glycerate contents (Timm et al., 2012). Therefore, to overcome those detrimental effects of photorespiratory intermediates, this pathway is highly activated under fluctuating light, which then benefits photosynthetic CO 2 assimilation and photosynthetic electron flow. In addition, the operation of this pathway is necessary for the regeneration of RuBP (Takahashi et al., 2007). To optimize photosynthetic CO 2 fixation, the rates of RuBP oxygenation and RuBP regeneration through photorespiratory pathway must be balanced. Therefore, under fluctuating light conditions, strong activation of the photorespiratory pathway accelerates RuBP regeneration, preventing a decrease in the RuBP pool and favoring the Calvin cycle.
In summary, our results provide evidence that, for sungrown tobacco leaves, fluctuating light conditions significantly decrease both stomatal and mesophyll conductances, as well as chloroplast CO 2 concentration. Consequently, the rate of CO 2 assimilation is limited by RuBP carboxylation under such conditions. Meanwhile, the photorespiratory pathway is highly activated to regulate photosynthetic electron flow and benefit photosynthetic CO 2 fixation. Thus, strong activation of this pathway is an important strategy by which sun-grown plants adapt to fluctuating light.