Dithiothreitol,an inhibitor of violaxanthin de-epoxidation,increases the susceptibility of leaves ofNerium oleander L. to photoinhibition of photosynthesis

Leaves ofNerium oleander L. plants, which had been previously kept in a shaded glasshouse for at least two months, were fed 1 mM dithiothreitol (DTT) through their petioles, either for 12h in darkness (overnight) or for 2h in low light (28 μmol photons·m⁻²·s⁻¹), in each case followed by a 3-h exposure to high light (1260 μmol photons·m⁻²·s⁻¹). During exposure to high light, violaxanthin became converted to zeaxanthin in control leaves, to which water had been fed, whereas zeaxanthin did not accumulate in leaves treated with DTT. Total carbon gain was not reduced by DTT during the photoinhibitory treatment. Exposure to high light led to a decrease in the photochemical efficiency of photosystem II, measured as the ratio of variable over maximum fluorescence emission,F _v/F _M, at both 298 K and 77K. The decrease was much more pronounced in the presence of DTT, mainly owing to a sustained increase in the instantaneous fluorescence,F _o. By contrast, in the control leaves,F _o determined immediately after the high-light treatment showed a transient decrease below theF _o value obtained before the onset of the photoinhibitory treatment (i.e. after 12 h dark adaptation), followed by a rapid return (within seconds) to this original level ofF _o during the following recovery period in darkness. Incubation of leaves with DTT led to large, sustained decreases in the photon-use efficiency of photosynthetic O₂ evolution by bright light, whilst the capacity of photosynthetic O₂ evolution at light and CO₂ saturation was less affected. In the control leaves, only small reductions in the photon yield and in the photosynthetic capacity were observed. These findings are consistent with previous suggestions that zeaxanthin, formed in the xanthophyll cycle by de-epoxidation of violaxanthin, is involved in protecting the photosynthetic apparatus against the adverse effects of excessive light.     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: effect of growth temperature on photoinhibition and recovery

Intact leaves of kiwifruit (Actinidia deliciosa (A. Chev.) C.F. Liang et A.R. Ferguson) from plants grown in a range of controlled temperatures from 15/10 to 30/25°C were exposed to a photon flux density (PFD) of 1500 μmol·m⁻²·s⁻¹ at leaf temperatures between 10 and 25°C. Photoinhibition and recovery were followed at the same temperatures and at a PFD of 20 μmol·m⁻²·s⁻¹, by measuring chlorophyll fluorescence at 77 K and 692 nm, by measuring the photon yield of photosynthetic O₂ evolution and light-saturated net photosynthetic CO₂ uptake. The growth of plants at low temperatures resulted in chronic photoinhibition as evident from reduced fluorescence and photon yields. However, low-temperature-grown plants apparently had a higher capacity to dissipate excess excitation energy than leaves from plants grown at high temperatures. Induced photoinhibition, from exposure to a PFD above that during growth, was less severe in low-temperature-grown plants, particularly at high exposure temperatures. Net changes in the instantaneous fluorescence,F ₀, indicated that little or no photoinhibition occurred when low-temperature-grown plants were exposed to high-light at high temperatures. In contrast, high-temperature-grown plants were highly susceptible to photoinhibitory damage at all exposure temperatures. These data indicate acclimation in photosynthesis and changes in the capacity to dissipate excess excitation energy occurred in kiwifruit leaves with changes in growth temperature. Both processes contributed to changes in susceptibility to photoinhibition at the different growth temperatures. However, growth temperature also affected the capacity for recovery, with leaves from plants grown at low temperatures having moderate rates of recovery at low temperatures compared with leaves from plants grown at high temperatures which had negligible recovery. This also contributed to the reduced susceptibility to photoinhibition in low-temperature-grown plants. However, extreme photoinhibition resulted in severe reductions in the efficiency and capacity for photosynthesis.     

Comparison of the effect of excessive light on chlorophyll fluorescence (77K) and photon yield of O2 evolution in leaves of higher plants

High-light treatments (1750–2000 mol photons m^–2 · s^–1) of leaves from a number of higher-plant species invariably resulted in quenching of the maximum 77K chlorophyll fluorescence at both 692 and 734 nm (F _M, 692 and F _M, 734). The response of instantaneous fluorescence at 692 nm (F _O, 692) was complex. In leaves of some species F _O, 692 increased dramatically in others it was quenched, and in others yet it showed no marked, consistent change. Regardless of the response of F _O, 692 an apparently linear relationship was obtained between the ratio of variable to maximum fluorescence (F _V/F _M, 692) and the photon yield of O₂ evolution, indicating that photoinhibition affects these two variables to approximately the same extent. Treatment of leaves in a CO₂–free gas stream containing 2% O₂ and 98% N₂ under weak light (100 mol · m^–2 · s^–1) resulted in a general and fully reversible quenching of 77K fluorescence at 692 and 734 nm. In this case both F _O, 692 and F _M, 692 were invariably quenched, indicating that the quenching was caused by an increased non-radiative energy dissipation in the pigment bed. We propose that high-light treatments can have at least two different, concurrent effects on 77K fluorescence in leaves. One results from damage to the photosystem II (PSII) reaction-center complex and leads to a rise in F _O, 692; the other results from an increased non-radiative energy dissipation and leads to quenching of both F _O, 692 and F _M, 692 This general quenching had a much longer relaxation time than reported for pH-dependent quenching in algae and chloroplasts. Sun leaves, whose F _V/F _M, 692 ratios were little affected by high-light exposure in normal air, suffered pronounced photoinhibition when the exposure was made under conditions that prevent photosynthetic gas exchange (2% O₂, 0% CO₂). However, they were still less susceptible than shade leaves, indicating that the higher capacity for energy dissipation via photosynthesis is not the only cause of their lower susceptibility. The rate constant for recovery from photoinhibition was much higher in mature sun leaves than in mature shade leaves, indicating that differences in the capacity for continuous repair may in part account for the difference in their susceptibility to photoinhibition.Abbreviations and symbols kDa kilodalton - LHC-II light-harvesting chlorophyll-protein complex - PFD photon flux density (photon fluence rate) - PSI, PSII photosystem I, II - F _O, F _M, F _V instantaneous, maximum, variable fluorescence emission - absorptance - _a photon yield of O₂ evolution (absorbed light) C.I.W.-D.P.B. Publication No. 925     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: Changes in susceptibility to photoinhibition and recovery during the growth season

Kiwifruit (Actinidia deliciosa (A. Chev.) C.F. Liang et A.R. Ferguson) plants grown in an outdoor enclosure were exposed to the natural conditions of temperature and photon flux density (PFD) over the growing season (October to May). Temperatures ranged from 14 to 21° C while the mean monthly maximum PFD varied from 1000 to 1700 mol · m^–2 · s^–1, although the peak PFDs exceeded 2100 mol · m^–2 · s^–1. At intervals, the daily variation in chlorophyll fluorescence at 692 nm and 77K and the photon yield of O₂ evolution in attached leaves was monitored. Similarly, the susceptibility of intact leaves to a standard photoinhibitory treatment of 20° C and a PFD of 2000 mol · m^–2 · s^–1 and the ability to recover at 25° C and 20 mol · m^–2 · s^–2 was followed through the season. On a few occasions, plants were transferred either to or from a shade enclosure to assess the suceptibility to natural photoinhibition and the capacity for recovery. There were minor though significant changes in early-morning fluorescence emission and photon yield throughout the growing season. The initial fluorescence, F_o, and the maximum fluorescence, F_m, were, however, significantly and persistently different from that in shade-grown kiwifruit leaves, indicative of chronic photoinhibition occurring in the sun leaves. In spring and autumn, kiwifruit leaves were photoinhibited through the day whereas in summer, when the PFDs were highest, no photoinhibition occurred. However, there was apparently no non-radiative energy dissipation occurring then also, indicating that the kiwifruit leaves appeared to fully utilize the available excitation energy. Nevertheless, the propensity for kiwifruit leaves to be susceptible to photoinhibition remained high throughout the season. The cause of a discrepancy between the severe photoinhibition under controlled conditions and the lack of photoinhibition under comparable, natural conditions remains uncertain. Recovery from photoinhibition, by contrast, varied over the season and was maximal in summer and declined markedly in autumn. Transfer of shade-grown plants to full sun had a catastrophic effect on the fluorescence characteristics of the leaf and photon yield. Within 3 d the variable fluorescence, F_v, and the photon yield were reduced by 80 and 40%, respectively, and this effect persisted for at least 20 d. The restoration of fluorescence characteristics on transfer of sun leaves to shade, however, was very slow and not complete within 15 d.Abbreviations and Symbols F_o, F_m, F_v initial, maximum, variable fluorescence - F_i F_v at t = 0 - F F_v at t = - PFD photon flux density - PSII photosystem II - leaf absorptance ratio - (_a photon yield of O₂ evolution (absorbed basis) - _{i a} at t = 0 - _a at t = We thank Miss Linda Muir and Amanda Yeates for their technical assistance in this study.     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: Recovery and its dependence on temperature

Recovery of photoinhibition in intact leaves of shade-grown kiwifruit was followed at temperatures between 10° and 35° C. Photoinhibition was initially induced by exposing the leaves for 240 min to a photon flux density (PFD) of 1 500 mol·m^-2·s^-1 at 20° C. In additional experiments to determine the effect of extent of photoinhibition on recovery, this period of exposure was varied between 90 and 400 min. The kinetics of recovery were followed by chlorophyll fluorescence at 77K. Recovery was rapid at temperatures of 25–35° and slow or negligible below 20° C. The results reinforce those from earlier studies that indicate chilling-sensitive species are particularly susceptible to photoinhibition at low temperatures because of the low rates of recovery. At all temperatures above 15° C, recovery followed pseudo first-order kinetics. The extent of photoinhibition affected the rate constant for recovery which declined in a linear fashion at all temperatures with increased photoinhibition. However, the extent of photoinhibition had little effect on the temperature-dependency of recovery. An analysis of the fluorescence characteristics indicated that a reduction in non-radiative energy dissipation and repair of damaged reaction centres contributed about equally to the apparent recovery though biochemical studies are needed to confirm this. From an interpretation of the kinetics of photoinhibition, we suggest that recovery occurring during photoinhibition is limited by factors different from those that affect post-photoinhibition recovery.Abbreviations and symbols F _o, F _m, F _v instantaneous, maximum, variable fluorescence - K _D, K _F, K _P, K _T rate constants for non-radiative energy dissipation, fluorescence, photochemistry, transfer to photosystem I - K(PI), k(R) rate constants for photoinhibition and recovery - PFD photon flux density - PSI, II photosystem I, II - _i photon yield of photosynthesis (incident light)     

The ratio of variable to maximum chlorophyll fluorescence from photosystem II,measured in leaves at ambient temperature and at 77K,as an indicator of the photon yield of photosynthesis

The response of a number of species to high light levels was examined to determine whether chlorophyll fluorescence from photosystem (PS) II measured at ambient temperature could be used quantitatively to estimate the photon yield of O₂ evolution. In many species, the ratio of the yield of the variable (F_V) and the maximum chlorophyll fluorescence (F_M) determined from leaves at ambient temperature matched that from leaves frozen to 77K when reductions in F_V/F_M and the photon yield resulted from exposure of leaves to high light levels under favorable temperatures and water status. Under conditions which were less favorable for photosynthesis, F_V/F_M at ambient temperature often matched the photon yield more closely than F_V/F_M measured at 77K. Exposure of leaves to high light levels in combination with water stress or chilling stress resulted in much greater reductions in the photon yield than in F_V/F_M (at both ambient temperature and 77K) measured in darkness, which would be expected if the site of inhibition was beyond PSII. Following chilling stress, F_V/F_M determined during measurement of the photon yield in the light was depressed to a degree more similar to that of the depression of photon yield, presumably as a result of regulation of PSII in response to greatly reduced electron flow.Abbreviations and Symbols F_o yield of instantaneous fluorescence - F_M yield of maximum fluorescence - F_V yield of variable fluorescence - PFD photon flux density (400–700 nm) - PSI (II) photosystem I (II) This work was supported by the Deutsche Forschungsgemeinchaft. W.W.A. gratefully acknowledges the support of Fellowships from the North Atlantic Treaty Organization and the Alexander von Humboldt-Stiftung. We also thank Maria Lesch for plant maintenance.     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: Effect of light during growth on photoinhibition and recovery

Photoinhibition of photosynthesis was induced in intact kiwifruit (Actinidia deliciosa (A. Chev.) C. F. Liang et A. R. Ferguson) leaves grown at two photon flux densities (PFDs) of 700 and 1300 mol·m^-2·s^-1 in a controlled environment, by exposing the leaves to PFD between 1000 and 2000 mol·m^-2·s^-1 at temperatures between 10 and 25°C; recovery from photoinhibition was followed at the same range of temperatures and at a PFD between 0 and 500 mol·m^-2·s^-1. In either case the time-courses of photoinhibition and recovery were followed by measuring chlorophyll fluorescence at 692 nm and 77K and by measuring the photon yield of photosynthetic O₂ evolution. The initial rate of photoinhibition was lower in the high-light-grown plants but the long-term extent of photoinhibition was not different from that in low-light-grown plants. The rate constants for recovery after photoinhibition for the plants grown at 700 and 1300 mol·m^-2·s^-1 or for those grown in shade were similar, indicating that differences between sun and shade leaves in their susceptibility to photoinhibition could not be accounted for by differences in capacity for recovery during photoinhibition. Recovery following photoinhibition was increasingly suppressed by an increasing PFD above 20 mol·m^-2·s^-1, indicating that recovery in photoinhibitory conditions would, in any case, be very slow. Differences in photosynthetic capacity and in the capacity for dissipation of non-radiative energy seemed more likely to contribute to differences in susceptibility to photoinhibition between sun and shade leaves of kiwifruit.Abbreviations and symbols F _o , F _m , F _v instantaneous, maximum, variable fluorescence - F _v /F _m fluorescence ratio - F _i =F _v at t=0 - F F _v at t= - K _D rate constant for photochemistry - k(F _p ) first-order rate constant for photoinhibition - k(F _r ) first-order rate constant for recovery - PFD photon flux density - PSII photosystem II - _i photon yield of O₂ evolution (incident light)     

Photoinhibition of photosynthesis in willow leaves under field conditions

Chlorophyll fluorescence of leaves of a willow (Salix sp.) stand grown in the field in northern Sweden was measured on several occasions during the growing season of 1987. For leaves that received mostly full daylight, the F _V/F _P ratio declined roughtly 15% in the afternoon on cloudless days in July (F _P is the fluorescence at the peak of the induction curve obtained at the prevailing air temperature after 45 min of dark adaptation, and F _V is variable fluoresence, F _V=F _P-F _O, where F _O is minimal fluorescence). There was no decrease in the F _V/F _P ratio on cloudy days, while the effect was intermediate on changeable days. In view of this light dependence, together with the fact that the decline in the F _V/F _P ratio was paralleled with an equal decline in the corresponding fluorescence ratio F _V/F _M at 77K, and a similar decline in the maximum quantum yield of O₂ evolution, it is suggested that the decline in the F _V/F _P ratio represents a damage in photosyntem II attributable to photoinhibition. Recovery of the F _V/F _P ratio in dim light following a decline on a cloudless day took 7–16 h to go to completion; the F _V/F _P ratio was fully restored the following morning. When all active leaves of a peripheral shoot were compared, the F _V/F _P ratio in the afternoon of a day of bright light varied greatly from leaf to leaf, though the majority of leaves showed a decline. This variation was matched by a pronounced variation in intercepted photon flux density. When leaves developed in the shade were exposed to full sunlight by trimming of the stand an increased sensitivity to photoinhibition was observed as compared to peripheral leaves. The present study indicates that peripheral willow shoots experienced in the order of 10–20% photoinhibition during an appreciable part of their life. This occurred even though the environmental conditions were within the optimal range of photosynthesis and growth.Abbreviations and symbols F _O minimum fluorescence - F _P fluorescence at the peak of the induction curve obtained at normal ambient temperatures - F _V variable fluorescence - F _M maximum fluorescence obtained at 77K - PPFD photosynthetic photon flux density     

Photoinhibition of photosynthesis in intact kiwifruit (Actinidia deliciosa) leaves: Effect of temperature

Photoinhibition of photosynthesis was induced in attached leaves of kiwifruit grown in natural light not exceeding a photon flux density (PFD) of 300 mol·m^-2·s^-1, by exposing them to a PFD of 1500 mol·m^-2·s^-1. The temperature was held constant, between 5 and 35° C, during the exposure to high light. The kinetics of photoinhibition were measured by chlorophyll fluorescence at 77K and the photon yield of photosynthetic O₂ evolution. Photoinhibition occurred at all temperatures but was greatest at low temperatures. Photoinhibition followed pseudo first-order kinetics, as determined by the variable fluorescence (F _v) and photon yield, with the long-term steady-state of photoinhibition strongly dependent on temperature wheareas the observed rate constant was only weakly temperature-dependent. Temperature had little effect on the decrease in the maximum fluorescence (F _m) but the increase in the instantaneous fluorescence (F _o) was significantly affected by low temperatures in particular. These changes in fluorescence indicate that kiwifruit leaves have some capacity to dissipate excessive excitation energy by increasing the rate constant for non-radiative (thermal) energy dissipation although temperature apparently had little effect on this. Direct photoinhibitory damage to the photosystem II reaction centres was evident by the increases in F _o and extreme, irreversible damage occurred at the lower temperatures. This indicates that kiwifruit leaves were most susceptible to photoinhibition at low temperatures because direct damage to the reaction centres was greatest at these temperatures. The results also imply that mechanisms to dissipate excess energy were inadequate to afford any protection from photoinhibition over a wide temperature range in these shade-grown leaves.Abbreviations and symbols fluorescence yield correction coefficient - F _o, F _m, F _v instantaneous, maximum, variable fluorescence - K _D, K _F, K _P, K _T rate constants for non-radiative energy dissipation, fluorescence, photochemistry, energy transfer to photosystem I - PFD photon flux density - PSI, II photosystem I, II - _i photon yield of photosynthesis (incident light)     

Photoinhibition of photosynthesis in intact bean leaves: role of light and temperature,and requirement for chloroplast-protein synthesis during recovery

Photoinhibition of photosynthesis was induced in intact leaves of Phaseolus vulgaris L. grown at a photon flux density (PFD; photon fluence rate) of 300 mol·m^-2·s^-1, by exposure to a PFD of 1400 mol·m^-2·s^-1. Subsequent recovery from photoinhibition was followed at temperatures ranging from 5 to 35°C and at a PFD of either 20 or 140 mol·m^-2·s^-1 or in complete darkness. Photoinhibition and recovery were monitored mainly by chlorophyll fluorescence emission at 77K but also by photosynthetic O₂ evolution. The effects of the protein-synthesis inhibitors, cycloheximide and chloramphenicol, on photoinhibition and recovery were also determined. The results demonstrate that recovery was temperature-dependent with rates slow below 15°C and optimal at 30°C. Light was required for maximum recovery but the process was light-saturated at a PFD of 20 mol·m^-2·s^-1. Chloramphenicol, but not cycloheximide, inactivated the repair process, indicating that recovery involved the synthesis of one or more chloroplast-encoded proteins. With chloramphenicol, it was shown that photoinhibition and recovery occurred concomitantly. The temperature-dependency of the photoinhibition process was, therefore, in part determined by the effect of temperature on the recovery process. Consequently, photoinhibition is the net difference between the rate of damage and the rate of repair. The susceptibility of chilling-sensitive plant species to photoinhibition at low temperatures is proposed to result from the low rates of recovery in this temperature range.Abbreviations and symbols Da Dalton - Fo, Fm, Fv instantaneous, maximum, variable fluorescence emission - PFD photon flux density - PSII photosystem II - photon yield C.I.W.-D.P.B. Publication No. 871     

Analysis of dark-relaxation kinetics of variable fluorescence in intact leaves

The dark-relaxation kinetics of variable fluorescence, F_v, in intact green leaves of Pisum stativum L. and Dolichos lablab L. were analyzed using modulated fluorometers. Fast (t_1/2 = 1 s) and slow (t_1/2 = 7–8 s) phases in f_v dark-decay kinetics were observed; the rate and the relative contribution of each phase in total relaxation depended upon the fluence rate of the actinic light and the point in the induction curve at which the actinic light was switched off. The rate of the slow phase was accelerated markedly by illumination with far-red light; the slow phase was abolished by methyl viologen. The halftime of the fast phase of F_v dark decay decreased from 250 ms in dark-adapted leaves to 12–15 ms upon adaptation to red light which is absorbed by PSII. The analysis of the effect of far-red light, which is absorbed mainly by PSI, on F_v dark decay indicates that the slow phase develops when a fraction of Q_A ^– (the primary stable electron acceptor of PSII) cannot transfer electrons to PSI because of limitation on the availability of P700⁺ (the primary electron donor of PSI). After prolonged illumination of dark-adapted leaves in red (PSII-absorbed) light, a transient. F_v rise appears which is prevented by far-red (PSI-absorbed) light. This transient f_v rise reflects the accumulation of Q_A ^– in the dark. The observation of this transient F_v rise even in the presence of the uncoupler carbonylcyanide m-chlorophenyl hydrazone (CCCP) indicates that a mechanism other than ATP-driven back-transfer of electrons to QA may be responsible for the phenomenon. It is suggested that the fast phase in F_v dark-decay kinetics represents the reoxidation of Q_A ^– by the electron-transport chain to PSI, whereas the slow phase is likely to be related to the interaction of Q_A ^– with the donor side of PSII.Abbreviations CCCP carbonylcyanide m-chlorophenylhydrazone - F_O initial fluorescence level - F_v variable fluorescence - P700 primary electron donor of PSI - PSI, II photosystem I, II - Q_A (Q_A ^–) Q_B (Q_B ^–) primary and secondary stable electron acceptor of PSII in oxidized (reduced) state Supported by grant B6.1/88 DST, Govt. of India.     

Estimation of the effect of photoinhibition on the carbon gain in leaves of a willow canopy

The occurrence of photoinhibition of photosynthesis in leaves of a willow canopy was examined by measuring the chlorophyll-a fluorescence ratio of F _V/F _M (F_M is the maximum fluorescence level of the induction curve, and F_V is the variable fluorescence, F _V=F _M–F ₀, where F₀ is the minimal fluorescence). The majority of the leaves situated on the upper parts of peripheral shoots showed an afternoon inhibition of this ratio on clear days. This was the consequence of both a decrease in F _M and a rise in F _O. In the same leaves the diurnal variation in intercepted photosynthetic photon flux density (PPFD) was monitored using leaf-mounted sensors. Using the multivariate method, partial least squares in latent variables, it is shown that the dose of PPFD, integrated and linearly weighted over the last 6-h period, best predicts photoinhibition. Photoinhibition occurred even among leaves that did not intercept PPFDs above 1000 mol·m^–2·s^–1. Exposure of leaves to a standard photoinhibitory treatment demonstrated that the depression in the F _V/F _M ratio was paralleled by an equal depression in the maximal quantum yield of CO₂ uptake and a nearly equal depression in the rate of bending (convexity) of the light-response curve of CO₂ uptake. As a result, the rate of net photosynthesis is depressed over the whole natural range of PPFD. By simulating the daily course in the rate of net photosynthesis, it is estimated that in the order of one-tenth of the potential carbon gain of peripheral willow shoots is lost on clear days as a result of photoinhibition. This applies to conditions of optimal temperatures. Photoinhibition is even more pronounced at air temperatures below 23° C, as judged from measurements of the F_V/F_M ratio on clear days: the afternoon inhibition of this ratio increased in a curvilinear manner from 15% to 25% with a temperature decrease from 23° to 14° C.Abbreviations and Symbols F_O minimum fluorescence - F_V variable fluorescence - F_M maximum fluorescence - PLS partial least squares in latent variables - PPFD photosynthetic photon flux density - VPD water vapour-pressure deficit This study was supported by the Swedish Natural Science Research Council. We are indebted to Dr. Jerry Leverenz (Department of Plant Physiology, University of Umeå, Sweden) for guidance with the modelling of the photosynthesis data.     

Photoinhibition, 77K chlorophyll fluorescence quenching and phosphorylation of the light-harvesting chlorophyll-protein complex of photosystem II in soybean leaves

When the capacity of leaves for orderly dissipation of excitation energy in photosynthesis is exceeded, one mechanism by which the excess energy appears to be dissipated is through a nonradiative decay process. This process is observed as a reversible quenching of chlorophyll fluorescence emission (77K) from both photosystem II and photosystem I which persists in darkness (Demmig and Björkman 1987, Planta 171, 171–184). Fluorescence quenching was induced in soybean (Glycine max (L.) Merr.) leaves by two methods: 1) changing the composition of the gas surrounding the leaf from normal air to 2% O₂, 0% CO₂ at a low, constant photon flux density (PFD=photon fluence rate), and 2) increasing the PFD in the presence of normal air. In either case the quenching was fully reversible after return to the original condition (low PFD, normal air). The half-time of the relaxation of the quenching was in the order of 30 min. Both treatments resulted in reversible dephosphorylation of the light-harvesting chlorophyll-protein complex of photosystem II (LHC-II). Treatment under photoinhibitory conditions (high PFD plus chloramphenicol) also caused dephosphorylation of LHC-II. Therefore, phosphorylation of LHC-II cannot account for the observed fluorescence quenching. In addition, our results indicate that in vivo a factor other than the redox state of the plastoquinone pool controls LHC-II phosphorylation. This factor may be pH, the pH gradient across the thylakoid membranes.Abbreviations and symbols CAP chloramphenicol - F_o, F_M, F_v instantaneous, maximumr variable fluorescence emission - LHC-II light-haryesting chlorophyll-protein complex of PSII - kDa kilodalton - pH pH gradient across the thylakoid membrane - PFD photon flux density (photon fluence rate) - PQ plastoquinone - PSI, PSII photosystem I, II - Q acceptor of PSII C.I.W.-D.P.B. Publication No. 926     

The relationship between carbon dioxide fixation and chlorophyll a fluorescence during induction of photosynthesis in maize leaves at different temperatures and carbon dioxide concentrations

The rate of CO₂ fixation (F_c) and 680 nm chlorophyll fluorescence emission (F680) were measured simultaneously during induction of photosynthesis in Zea mays L. leaves under varying experimental conditions in order to assess the validity of fluorescence as an indicator of in vivo photosynthetic carbon assimilation. Z. mays leaves showed typical Kautsky fluorescence induction curves consisting of a fast rise in emission (O to P) followed by a slow quenching via a major transient (S-M) to a steady-state (T). After an initial lag, net CO₂ assimilation commenced at a point corresponding to the onset of the S-M transient on the F680 induction curve. Subsequently, F_c and F680 always arrived at a steady-state simultaneously. Decreasing the dark-adaption period increased the rate of induction of both parameters. Alteration of leaf temperature produced anti-parallel changes in induction characteristics of F_c and F680. Reducing the CO₂ level to below that required for saturation of photosynthesis also produced anti-parallel changes during induction, however, at CO₂ concentrations tenfold greater than the atmospheric level the rate of F680 quenching from P to T was appreciably reduced without a similar change in the induction of F_c. Removal of CO₂ at steady-state produced only a small increase in F680 and a correspondingly small decrease in F680 occurred when CO₂ was re-introduced. The complex relationship between chlorophyll fluorescence and carbon assimilation in vivo is discussed and the applicability of fluorescence as an indicator of carbon assimilation is considered.Abbreviations F_c rate of CO₂ fixation - F680 fluorescence emission at 680 nm     

Photoinhibition at chilling temperature

The effects of moderate light at chilling temperature on the photosynthesis of unhardened (acclimated to +18° C) and hardened (cold-acclimated) spinach (Spinacea oleracea L.) leaves were studied by means of fluorescence-induction measurements at 20° C and 77K and by determination of quantum yield of O₂ evolution. Exposure to 550 mol photons·m^-2·s^-1 at +4° C induced a strong photoinhibition in the unhardened leaves within a few hours. Photoinhibition manifested by a decline in quantum yield was characterized by an increase in initial fluorescence (F _o) and a decrease in variable fluorescence (F _v) and in the ratio of variable to maximum fluorescence (F _V/F _M), both at 77K and 20° C. The decline in quantum yield was more closely related to the decrease in the F _V/F _M ratio measured at 20° C, as compared with F _V/F _M at 77K. Quenching of the variable fluorescence of photosystem II was accompanied by a decline in photosystem-I fluorescence at 77K, indicating increased thermal de-excitation of pigments as the main consequence of the light treatment. All these changes detected in fluorescence parameters as well as in the quantum yield of O₂ evolution were fully reversible within 1–3 h at a higher temperature in low light. The fast recovery led us to the view that this photoinhibition represents a regulatory mechanism protecting the photosynthetic apparatus from the adverse effects of excess light by increasing thermal energy dissipation. Long-term cold acclimation probably enforces other protective mechanisms, as the hardened leaves were insensitive to the same light treatment that induced strong inhibition of photosynthesis in unhardened leaves.Abbreviations F ₀ initial fluorescence - F _M maximum fluorescence - F _V variable fluorescence (F _M-F ₀ - PFD photon flux density - PS photosystem     

Photoinhibition of photosynthesis under natural conditions in ivy (Hedera helix L.) growing in an understory of deciduous trees

We investigated to what extent south-exposed leaves (E-leaves) of the evergreen ivy (Hedera helix L.) growing in the shadow of two deciduous trees suffered from photoinhibition of photosynthesis when leaf-shedding started in autumn. Since air temperatures drop concomitantly with increase in light levels, changes in photosynthetic parameters (apparent quantum yield, _i and maximal photosynthetic capacity of O₂ evolution, P_max; chlorophyll-a fluorescence at room temperature) as well as pigment composition were compared with those in north-exposed leaves of the same clone (N-leaves; photosynthetic photon flux density PPFD< 100 mol · m^–2 · s^–2) and phenotypic sun leaves (S-leaves; PPFD up to 2000 mol · m^–2 · s^–1).In leaves exposed to drastic light changes during winter (E-leaves) strong photoinhibition of photosynthesis could be observed as soon as the incident PPFD increased in autumn. In contrast, in N-leaves the ratio of variable fluorescence to maximum fluorescence (F_V/F_Mm) and _i did not decline appreciably prior to severe frosts (up to -12° C) in January. At this time, _i was reduced to a similar extent in all leaves, from about 0.073 mol O₂ · mol^–1 photons before stress to about 0.020. Changes in _i were linearly correlated with changes in f_v/f_m (r = 0.955). The strong reduction in F_V/F_M on exposure to stress was caused by quenching in F_M. The initial fluorescence (F₀), however, was also quenched in all leaves. The diminished fluorescence yield was accompanied by an increase in zeaxanthin content. These effects indicate that winter stress in ivy primarily induces an increase in non-radiative energy-dissipation followed by photoinhibitory damage of PSII. Although a pronounced photooxidative bleaching of chloroplast pigments occurred in January (especially in E-leaves), photosynthetic parameters recovered completely in spring. Thus, the reduction in potential photosynthetic yield in winter may be up to three times greater in leaves subjected to increasing light levels than in leaves not exposed to a changing light environment.Abbreviations and Symbols F₀, F_M initial and maximal fluorescence yield when all PSII centres are open and closed - F_V variable fluorescence (F_M-F₀) - P_max maximal photosynthetic capacity at 1000 umol · m^–2 · s^–1 PPFD and CO₂ saturation - PPFD photosynthetic photon flux density - _i apparent quantum yield of photosynthetic O₂ evolution - E-leaves, N-leaves shade leaves exposed, not exposed to drastic light changes during winter - S-leaves sun leaves from an open ivy stand Dedicated to Professor Otto Härtel on the occasion of his 80th birthdayThis work was supported by the Austrian Fonds zur Förderung der wissenschaftlichen Forschung.     

Photon yield of O2 evolution and chlorophyll fluorescence characteristics at 77 K among vascular plants of diverse origins

Photon yields of oxygen evolution at saturating CO₂ were determined for 44 species of vascular plants, representing widely diverse taxa, habitats, life forms and growth conditions. The photonyield values on the basis of absorbed light ( ^a) were remarkably constant among plants possessing the same pathway of photosynthetic CO₂ fixation, provided the plants had not been subjected to environmental stress. The mean ^a value ±SE for 37 C₃ species was 0.106±0.001 O₂·photon^-1. The five C₄ species exhibited lower photon yields and greater variation than the C₃ species ( ^a=0.0692±0.004). The ^a values for the two Crassulaceanacid-metabolism species were similar to those of C₃ species. Leaf chlorophyll content had little influence on ^a over the range found in normal, healthy leaves. Chlorophyll fluorescence characteristics at 77 K were determined for the same leaves as used for the photon-yield measurements. Considerable variation in fluorescence emission both at 692 nm and at 734 nm, was found 1) among the different species; 2) between the upper and lower surfaces of the same leaves; and 3) between sun and shade leaves of the same species. By contrast, the ratio of variable to maximum fluorescence emission at 692 nm (F_v/F_M, 692) remained remarkably constant (The mean value for the C₃ species was 0.832±0.004). High-light treatments of shade leaves resulted in a reduction in both ^a and the F_v/F_M, 692 ratio. The extent of the reductions increased with time of exposure to bright light. A linear relationship was obtained when ^a was plotted against F_v/F_M, 692. The results show that determinations of the photon yield of O₂ evolution and the F_v/F_M, 692 ratio can serve as excellent quantitative measures of photoinhibition of overall photosynthetic energy-conversion system and of photochemistry of photosystem II, respectively. This is especially valuable in field work where it is often impossible to obtain appropriate controls.Abbreviations and symbols CAM Crassulacean acid metabolism - PFD photon flux density (photon fluence rate) - PSI, PSII photosystem I, II - F_o, F_M, F_v instantaneous, maximum, variable fluorescence emission - absorptance - ^a photon yield (absorbed light) - ⁱ photon yield (incident light) C.I.W.-D.P.B. Publication No. 923     

Photosynthetic oxygen evolution and chlorophyll fluorescence in intact isolated chloroplasts on a solid support: the influence of orthophosphate

We devised recently a method to trap intact isolated chloroplasts on a solid support consisting of membrane filters made of cellulose nitrate (Cerovi et al., 1987, Plant Physiol. 84, 1249–1251). The addition of alkaline phosphatase to the reaction medium enabled continuous photosynthesis by spinach (Spinacia oleracea L.) chloroplasts to be sustained by hydrolysis of newly produced and exported triose phosphates and recycling of orthophosphate. In this system, simultaneous measurements of chlorophyll fluorescence and oxygen evolution were performed and their dependence on orthophosphate concentration was investigated. Optimal photosynthesis was obtained at a much higher initial orthophosphate concentration (2–4 mM) compared to intact chloroplasts in suspension. Secondary kinetics of chlorophyll fluorescence yield were observed and were shown to depend on the initial orthophosphate concentration.Abbreviations Chl chlorophyll - CSS intact isolated chloroplasts on solid support - ICS intact isolated chloroplasts in suspension - P_i orthophosphate - v rate of O₂ evolution - PPFD photosynthetic photon flux density The authors wish to thank Dr. Marijana Plesniar, from the University of Novi Sad, for stimulating discussions. This work was supported by the Fond for Science of the Republic of Serbia. Z.G.C.'s visit to the Robert Hill Laboratory was supported by the British Council and the University of Sheffield.     

Regulation of photosynthetic electron-transport in Phaseolus vulgaris L., as determined by room-temperature chlorophyll a fluorescence

The regulation of photosystem II (PSII) by light-, CO₂-, and O₂-dependent changes in the capacity for carbon metabolism was studied. Estimates of the rate of electron transport through PSII were made from gas-exchange data and from measurements of chlorophyll fluorescence. At subsaturating photon-flux density (PFD), the rate of electron transport was independent of O₂ and CO₂. Feedback on electron transport was observed under two conditions. At saturating PFD and low partial pressure of CO₂, p(CO₂), the rate of electron transport increased with p(CO₂). However, at high p(CO₂), switching from normal to low p(O₂) did not affect the net rate of photosynthetic CO₂ assimilation but the rate of electron-transport decreased by an amount related to the change in the rate of photorespiration. We interpret these effects as 1) regulation of ribulose-1,5-bisphosphatecarboxylase (RuBPCase, EC 4.1.1.39) activity to match the rate of electron transport at limiting PFD, 2) regulation of electron-transport rate to match the rate of RuBPCase at low p(CO₂), and 3) regulation of the electron-transport rate to match the capacity for starch and sucrose synthesis at high p(CO₂) and PFD. These studies provide evidence that PSII is regulated so that the capacity for electron transport is matched to the capacity for other processes required by photosynthesis, such as ribulose-bisphosphate carboxylation and starch and sucrose synthesis. We show that at least two mechanisms contribute to the regulation of PSII activity and that the relative engagement of these mechanisms varies with time following a step change in the capacity for ribulose-bisphosphate carboxylation and starch and sucrose synthesis. Finally, we take advantage of the relatively slow activation of deactivated RuBPCase in vivo to show that the activation level of this enzyme can limit the rate of electron transport as evidenced by increased feedback on PSII following a step change in p(CO₂). As RuBPCase as activated, the feedback on PSII declined.Abbreviations and symbols J_C electron-transport rate calculated from CO₂-assimilation measurements - J_F electron-transport rate calculated from fluorescence parameters - PFD photon-flux density - q_E energy-dependent quenching - PSII photosystem II - q_Q Q-dependent quenching - QY quantum yield - RuBPCase ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39) C.I.W. publication No. 1015