The relationship between non-photochemical quenching of chlorophyll fluorescence and the rate of photosystem 2 photochemistry in leaves

It has been suggested previously that non-photochemical quenching of chlorophyll fluorescence is associated with a decrease in the rate of photosystem 2 (PS 2) photochemistry. In this study analyses of fluorescence yield changes, induced by flashes in leaves exhibiting different amounts of non-photochemical quenching of fluorescence, are made to determine the effect of non-photochemical excitation energy quenching processes on the rate of PS 2 photochemistry. It is demonstrated that both the high-energy state and the more slowly relaxing components of non-photochemical quenching reduce the rate of PS 2 photochemistry. Flash dosage response curves for fluorescence yield show that non-photochemical quenching processes effectively decrease the relative effective absorption cross-section for PS 2 photochemistry. It is suggested that non-photochemical quenching processes exert an effect on the rate of PS 2 photochemistry by increasing the dissipation of excitation energy by non-radiative processes in the pigment matrices of PS 2, which consequently results in a decrease in the efficiency of delivery of excitation energy for PS 2 photochemistry.     

Resolution of components of non-photochemical chlorophyll fluorescence quenching in barley leaves

Non-photochemical chlorophyll fluorescence quenching (qN) in barley leaves has been analysed by monitoring its relaxation in the dark, by applying saturating pulses of light. At least three kinetically distinct phases to qN recovery are observed, which have previously been identified (Quick and Stitt 1989) as being due to high-energy state quenching (fast), excitation energy redistribution due to a state transition (medium) and photoinhibition (slow). However, measurements of chlorophyll fluorescence at 77 K from leaf extracts show that state transitions only occur in low light conditions, whereas the medium component of qN is very large in high light. The source of that part of the medium component not accounted for by a state transition is discussed.Abbreviations ATP adenosine 5-triphosphate - DCMU 3[3,4-dichlorophenyl]-1,1 dimethylurea - pH trans-thylakoid pH gradient - F_o, F_m room-temperature chlorophyll fluorescence yield with all reaction centres open, closed - F_v variable fluorescence = F_m–F_o - LHC II Light harvesting complex II - PS I, PS II Photosystem I, II - P700, P680 primary donor in photosystem I, II - qP photochemical quenching of variable fluorescence - qN non-photochemical quenching of variable fluorescence - qN_e, qN_t, qN_i non-photochemical quenching due to high energy state, state transition, photoinhibition - qN_f, qN_m, qN_s components of qN relaxing fast, medium, slow - q_r quenching of r relative to the dark state - tricine N-tris[hydroxymethyl]methylglycine - r ratio of fluorescence maximum from photosystem II to that from photosystem I at 77 K     

The occurrence of rapidly reversible non-photochemical quenching of chlorophyll a fluorescence in cyanobacteria

Cyanobacteria have previously been considered to differ fundamentally from plants and algae in their regulation of light harvesting. We show here that in fact the ecologically important marine prochlorophyte, Prochlorococcus, is capable of forming rapidly reversible non-photochemical quenching of chlorophyll a fluorescence (NPQf or qE) as are freshwater cyanobacteria when they employ the iron stress induced chlorophyll-based antenna, IsiA. For Prochlorococcus, the capacity for NPQf is greater in high light-adapted strains, except during iron starvation which allows for increased quenching in low light-adapted strains. NPQf formation in freshwater cyanobacteria is accompanied by deep Fo quenching which increases with prolonged iron starvation.     

Non-photochemical quenching of Fo in leaves is emission wavelength dependent: consequences for quenching analysis and its interpretation

The effects of light-induced non-photochemical quenching on the minimal F_o, and variable F_v, fluorescence emissions at 690 and 730 nm in leaves were determined. Non-photochemical quenching of F_o, but not F_v, was found to be dependent upon the wavelength of emission, and was greater at 690 nm than at 730 nm. For emission at 730, compared to at 690 nm, approx. 30% of F_o was not affected by non-photochemical quenching processes in leaves of C3 plants; in maize leaves this was found to be approx. 50%. The data indicate that a substantial proportion of the pigments contributing to F_o emission at 730 nm are not quenched by light-induced, non-photochemical quenching processes and that there are large differences in the pigment matrices contributing to F_o and F_v emissions at 730 nm, compared to those at 690 nm. These findings have important implications for the accurate estimation and interpretation of non-photochemical quenching of fluorescence parameters and their use in the calculation of photochemical efficiencies in leaves. Measurements of fluorescence emissions at wavelengths above 700 nm are likely to give rise to significant errors when used for determinations of photochemical and non-photochemical quenching parameters.     

Action potential in a plant cell lowers the light requirement for non-photochemical energy-dependent quenching of chlorophyll fluorescence

This study deals with effects of membrane excitation on photosynthesis and cell protection against excessive light, manifested in non-photochemical quenching (NPQ). In Chara corallina cells, NPQ and pericellular pH displayed coordinated spatial patterns along the length of the cell. The NPQ values were lower in H⁺-extruding cell regions (external pH ∼ 6.5) than in high pH regions (pH ∼ 9.5). Generation of an action potential by applying a pulse of electric current caused NPQ to increase within 30-60 s. This effect, manifested as a long-lived drop of maximum chlorophyll fluorescence (F_m′), occurred at lower photosynthetic flux densities (PFD) in the alkaline as compared to acidic cell regions. The light response curve of NPQ shifted, after generation of an action potential, towards lower PFD. The release of NPQ by nigericin and the rapid reversal of action potential-triggered NPQ in darkness indicate its relation to thylakoid ΔpH. Generation of an action potential shortly after darkening converted the chloroplasts into a latent state with the F_m identical to that of unexcited cells. This state transformed to the quenched state after turning on weak light that was insufficient for NPQ prior to membrane excitation of the cells. The ionophore, A23187, shifted NPQ plots similarly to the action potential effect, consistent with a likely role of a rise in the cytosolic Ca²⁺ level in the action potential-induced quenching. The results suggest that a rapid electric signal, across the plasma membrane, might exert long-lived effects on photosynthesis and chlorophyll fluorescence through ion flux-mediated pathways.     

Light-harvesting chlorophyll a-b complex requirement for regulation of Photosystem II photochemistry by non-photochemical quenching

Recently, it has been suggested (Horton et al. 1992) that aggregation of the light-harvesting a-b complex (LHC II) in vitro reflects the processes which occur in vivo during fluorescence induction and related to the major non-photochemical quenching (qE). Therefore the requirement of this chlorophyll a-b containing protein complex to produce qN was investigated by comparison of two barley mutants either lacking (chlorina f2) or depressed (chlorina¹⁰⁴) in LHC II to the wild-type and pea leaves submitted to intermittent light (IL) and during their greening in continuous light. It was observed that qN was photoinduced in the absence of LHC II, i.e. in IL grown pea leaves and the barley mutants. Nevertheless, in these leaves qN had no (IL, peas) or little (barley mutants) inhibitory effect on the photochemical efficiency of Q_A reduction measured by flash dosage response curves of the chlorophyll fluorescence yield increase induced by a single turn-over flash During greening in continuous light of IL pea leaves, an inhibitory effect on Q_A photoreduction associated to qN developed as Photosystem II antenna size increased with LHC II synthesis. Utilizing data from the literature on connectivity between PS II units versus antenna size, the following hypothesis is put forward to explain the results summarized above. qN can occur in the core antenna or Reaction Center of a fraction of PS II units and these units will not exhibit variable fluorescence. Other PS II units are quenched indirectly through PS II-PS II exciton transfer which develops as the proportion of connected PS II units increases through LHC II synthesis.     

Influence of Manganese Toxicity on Photosynthesis in Ricebean (Vigna Umbellata) Seedlings

Influence of manganese (Mn) toxicity on photosynthesis in ricebean (Vigna umbellata) was studied by the measurement of gas exchange characteristics and chlorophyll fluorescence parameters. The net photosynthetic rate (P _N), transpiration rate (E), and stomatal conductance (g _s) were reduced with increasing Mn concentration in nutrient solution. The reduction in g _s and E was more pronounced at 6 d of Mn treatment. However, P _N declined at 2 d of Mn treatment implying that the reduction in photosynthesis was not due to the direct effect of Mn on stomatal regulation. Mn did not affect the maximum efficiency of photosystem 2 (PS2) photochemistry (F_v/F_m). A reduction in photochemical quenching (q_P) and excitation capture efficiency of open PS2 (F_v′/F_m′) with a concomitant increase in q_N was observed. This implies that reduced demand for ATP and NADPH due to the reduction in photosynthesis causes a down-regulation of PS2 photochemistry and thus a high pH gradient (increase in q_N) and limited electron transport (decreased q_P). This revised version was published online in August 2006 with corrections to the Cover Date.     

应用非光化学淬灭初始变化率作为浮游植物光保护能力指标的可行性

在非光化学淬灭(Non-Photochemical Quenching,NPQ)荧光诱导的早期(1 min),NPQ随时间的变化率(NPQ/t)可作为衡量浮游植物光保护能力的指标,与常用的传统指标NPQm(最大NPQ)相比,这一新指标具有更易标准化、测量效率更高且包含的信息更丰富的优点。通过实验测量和理论推导证明了NPQ/t的常数性质,并通过可控光照条件下的室内培养实验证明了应用NPQ/t反映浮游植物光保护能力的合理性和有效性:NPQ/t和NPQm、光保护色素浓度呈线性相关,高光强下培养的藻类其NPQ/t高于低光强下培养的藻类,不同藻种之间的NPQ/t:扁藻(Tetraselmis chui)牟氏角毛藻(Chaetoceros mulleri)叉鞭金藻(Dicrateria inornata)。这一方法解决了野外实测研究中传统指标难以满足结果标准化和快速测量等要求的问题,对于推动浮游植物光保护生态学研究具有重要意义。     

Identification of a slowly inducible zeaxanthin-dependent component of non-photochemical quenching of chlorophyll fluorescence generated under steady-state conditions in Arabidopsis

The induction and relaxation of non-photochemical quenching (NPQ) under steady-state conditions, i.e. during up to 90 min of illumination at saturating light intensities, was studied in Arabidopsis thaliana. Besides the well-characterized fast qE and the very slow qI component of NPQ, the analysis of the NPQ dynamics identified a zeaxanthin (Zx) dependent component which we term qZ. The formation (rise time 10-15 min) and relaxation (lifetime 10-15 min) of qZ correlated with the synthesis and epoxidation of Zx, respectively. Comparative analysis of different NPQ mutants from Arabidopsis showed that qZ was clearly not related to qE, qT or qI and thus represents a separate, Zx-dependent NPQ component.     

Are red leaves photosynthetically active?

At irradiances close to those representing a sunny day, red and green leaves of poinsettia (Euphorbia pulcherrima) showed only minor differences in their photosynthetic capacities despite the strong differences in their pigment composition. However, contrarily to green leaves, red leaves did not show inhibition of photosynthesis at high irradiances, because anthocyanins protected chloroplasts from photoinhibition.     

Induction of photochemical and non-photochemical quenching of chlorophyll fluorescence by low concentrations of m-dinitrobenzene

Chlorophyll fluorescence quenching induced by low concentrations of m-dinitrobenzene (DNB) is investigated. In intact spinach chloroplasts DNB causes photochemical and non-photochemical quenching. The two forms of quenching are distinguished by applying the saturation pulse method with a new type of modulation fluorometer. Half-maximal photochemical quenching is observed at about 3 micromolar DNB. It is inhibited by 3-(3,4 dichlorophenyl)-1, 1-dimethylurea (DCMU) and by 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB). Photochemical quenching by DNB leads to suppression of the I-P transient in a fluorescence induction curve. Upon application of saturating continuous light, the increase of fluorescence yield is separated into a photochemical and a thermal part. DNB causes suppression of only the slowest sub-component of the thermal part, in analogy to the action of Hill reagents. Simultaneous measurements of oxygen exchange rate and fluorescence reveal that a part of DNB induced quenching is accompanied by oxygen uptake. Most DNB-induced non-photochemical quenching is prevented by nigericin and, hence, can be considered energy-dependent quenching. The small component persisting in the presence of nigericin is identical to the one observed with methylviologen and other Hill reagents, likely to be due to static quenching by oxidized plastoquinone. The presented data confirm the original finding of Etienne and Lavergne (Biochim Biophys Acta 283: 268–278, 1972) that low concentrations of DNB selectively affect the thermal component of variable fluorescence. However, while these authors interpreted the quenching by a non-photochemical mechanism, the present investigation emphasizes a photochemical mechanism, in analogy to the effect of electron acceptors or mediators.Abbreviations DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCMU 3-(3,4-dichlorophenyl)-1, 1-dimethylurea - DNB m-dinitrobenzene - PGA 3-phosphoglycerate - PMS phenazinemethosulphate - PS I and PS II photosystems I and II     

Influence of photosynthetic photon flux densities before and during long-term chilling on xanthophyll cycle and chlorophyll fluorescence quenching in leaves of tomato (Lycopersicon hirsutum)

A high-altitude ecotype of tomato ( Lycopersicon hirsutum f. typicum Humb. and Bonpl.) has previously been shown to resist further loss of photosynthetic function after three to four days of chilling stress. This study examined the influence of PPFD prior to, and during chilling on the development of protective zeaxanthin and energy-dependent quenching mechanisms in this ecotype. Five-week-old tomato plants were acclimated to either low PPFD (60 μmol m⁻² s⁻¹) or high PPFD (550 μmol m⁻² S⁻¹) at 25/20°C (day/night) for three days, and then exposed to a temperature of 5/5°C and a PPFD of either 60 or 550 μmol m⁻² s⁻¹ for three days. The plants acclimated to low PPFD had lower Chl a/b ratio, and lower level of total Chl per leaf area, total xanthophyll cycle pool and β-carotene. The capacity of their photosynthetic system to resist photoinhibition and to recover photosynthetic function was also lower compared to that of the plants acclimated at high PPFD but exposed to the same chilling stress. In the plants chilled at low PPFD, energy-dependent quenching preceded the formation of zeaxanthin on the first day of chilling and there was an overall reduction in the conversion of violaxanthin to zeaxanthin as compared to the plants chilled at high PPFD. During the last day of chilling-induced photoinhibition, energy-dependent quenching in any of the treatments did not increase, but zeaxanthin levels increased continuously throughout the three days of chilling. Our results suggest that light-acclimation before chilling affects the capacity of the plants to resist chilling-induced photoinhibition. In addition, photoinhibitory quenching appears to be a major component for quenching excessive energy at the latter stage of long-term chilling.     

Continuous recording of photochemical and non-photochemical chlorophyll fluorescence quenching with a new type of modulation fluorometer

A newly developed fluorescence measuring system is employed for the recording of chlorophyll fluorescence induction kinetics (Kautsky-effect) and for the continuous determination of the photochemical and non-photochemical components of fluorescence quenching. The measuring system, which is based on a pulse modulation principle, selectively monitors the fluorescence yield of a weak measuring beam and is not affected even by extremely high intensities of actinic light. By repetitive application of short light pulses of saturating intensity, the fluorescence yield at complete suppression of photochemical quenching is repetitively recorded, allowing the determination of continuous plots of photochemical quenching and non-photochemical quenching. Such plots are compared with the time courses of variable fluorescence at different intensities of actinic illumination. The differences between the observed kinetics are discussed. It is shown that the modulation fluorometer, in combination with the application of saturating light pulses, provides essential information beyond that obtained with conventional chlorophyll fluorometers.     

Diurnal and Seasonal Changes in Photosynthesis and Photosystem 2 Photochemical Efficiency in Prosopis juliflora Leaves Subjected to Natural Environmental Stress

The plants of Prosopis juliflora growing in northern India are exposed to large variations of temperature, vapour pressure deficits (VPD), and photosynthetic photon flux density (PPFD) throughout the year. Under these conditions P. juliflora had two short periods of leaf production, one after the winter season and second after summer, which resulted in two distinct even aged cohorts of leaves. In winter with cold nights (2–8 °C) and moderate temperatures during the day, the plants showed high rates of photosynthesis. In summer the midday temperatures often reached <45 °C and plants showed severe inhibition of photosynthesis. The leaves of second cohort appeared in July and showed typical midday depression of photosynthesis. An analysis of diurnal partitioning of the absorbed excitation energy into photochemistry showed that a smaller fraction of the energy was utilised for photochemistry and a greater fraction was dissipated thermally, further the photon utilisation for photochemistry and thermal dissipation is largely affected by the interaction of irradiance and temperature. The plants showed high photochemical efficiency of photosystem 2 (PS2) at predawn and very little photoinhibition in all seasons except in summer. The photoinhibition in summer was pronounced with very poor recovery during night. Since P. juliflora exhibited distinct pattern of senescence and production of new leaves after winter and summer stress period, it appeared that the ontogenic characteristic together with its ability for safe dissipation of excess radiant energy in P. juliflora contributes to its growth and survival.     

Interactions between nitrogen fertilization and ozone in watermelon cultivar Reina de Corazones in open-top chambers. Effects on chlorophyll a fluorescence, lipid peroxidation, and yield

Watermelon (Citrillus lanatus) plants were grown for two consecutive years in open-top chambers with three different ozone concentrations (O₃-free air, O₃ ambient, and air with additional O₃; CFA, NFA, and NFA+O₃) and three nitrogen fertilizer concentrations [0, 14.0, and 29.6 g N per pot; N0, N1, and N2). There was an interaction between ozone and N fertilizer for the major parameters studied. O₃ and N2 treatments led to a significant decrease in maximum efficiency of photosystem 2 (PS2) photochemistry (F_v/F_m), and induced a significant decrease in the actual quantum yield of PS2 (Φ_PS2), due mainly to the increased closure of PS2 reaction centres (q_P) and to an increase in the non-photochemical quenching (NPQ). On the other hand, these plants exhibited an increased susceptibility to photoinhibition, which could be associated with an increased fraction of reduced Q_A. An increase in lipid peroxidation indicated that damage was occurring at the membrane levels. High N concentration enhanced the detrimental effects of ozone on the fluorescence parameter induction and lipid peroxidation. All these negative alterations led to a decreased yield.     

Light-dependent, chilling effects on phosphorylation of thylakoid proteins and consequences for associated photochemical activities in maize

When maize ( Zea mays L. cv. LG11) leaves are exposed to low temperatures and high light modifications to both photosystem 2 (PS2) and the light-harvesting chlorophyll a/b protein complex associated with photosystem 2 (LHC2) occur. This study examines the consequences of these modifications for phosphorylation of LHC2 and PS2 polypeptides and the associated changes in electron transport. Maize leaves were chilled at 5°C for 6 h under photon flux densities of 1 500 and 250 μmol m^-2 s^-1. Thylakoids were then isolated from the leaves and their abilities to phosphorylate LHC2 and PS2 polypeptides and modify electron transport activities were determined. Measurements of chlorophyll fluorescence induction in the thylakoids were also made. Thylakoids isolated from leaves chilled under high light and from leaves kept in the ambient growth environment had similar phosphoprotein profiles. However, polypeptide phosphorylation in thylakoids from the chilled leaves did not produce a decrease in PS2 electron transport. Chilling leaves under low light produced a decrease in the ability of isolated thylakoids to phosphorylate PS2, but not LHC2, polypeptides, which was not associated with any change in the phosphorylation-induced decrease in PS2 electron transport. Chilling under high, but not low, light appears to produce changes in membrane organisation that do not affect the ability of the thylakoids to phosphorylate PS2 and LHC2 polypeptides, but which do prevent the phosphorylation-induced decrease in excitation energy transfer from LHC2 to PS2.     

Effects of different nitrogen forms on photosynthetic rate and the chlorophyll fluorescence induction kinetics of flue-cured tobacco

Net photosynthetic rate (P _N) of tobacco plants grown with NH₄-N as the only N source was the lowest all the times, while P _N grown only with NO₃-N was the greatest until 22^nd day, and P _N grown with both NO₃-N and NH₄-N (1 : 1) was the greatest. Maximal photochemical efficiency of photosystem 2 (PS2), F_v/F_m, and actual quantum yield of PS2 under actinic irradiation (Φ_PS2) in plants grown with only NH₄-N were greatest at early stage and then decreased and were smaller than those of other treatments. Photochemical quenching coefficient (q_P) and non-photochemical quenching coefficient (q_NP) in the NH₄-N plants were the greatest at all times. Hence excessive NH₄-N can decrease not only photochemical efficiency but also the efficiency of utilization of photon energy absorbed by pigments for photosynthesis. Therefore, excessive NH₄-N is a hindrance to photosynthesis of flue-cured tobacco. On the other hand, tobacco cultured with an appropriate mixture of NO₃-N with NH₄-N can sufficiently utilize photon energy and increase the efficiency of energy transformation.     

Temporal characteristics of chlorophyll fluorescence quenching and dissolved oxygen concentration as indicators of photosynthetic activity in Chlorella emersonii

A simple chlorophyll fluorescence (CF) measuring system has been implemented to study temporal characteristics of chlorophyll fluorescence induction (CFI) in dark-adapted freshwater algal cultures of Chlorella emersonii. There were two different decay time constants describing the CF quenching: τ₀ (the faster) and τ₁ (the slower) with amplitudes A₀ and A₁, respectively. The relative amplitude of the faster quenching component decreased once the sample was subject to deprivation from dissolved oxygen (DO). The DO concentration of samples was monitored to validate the effects of deprivation from air contact for up to 7 d and to the effect of adding DCMU to the culture (herbicide for blocking electron transport of photosystem 2). CFI analysis and DO measurements showed that the relative amplitude of A₀ to (A₀ + A₁) and the DO concentration can be used as an indication of relative photosynthetic activity, thus allowing for the possibility to classify the physiological state of algal blooms into active and inactive states.     

Spectral analysis on origination of the bands at 437 nm and 475.5 nm of chlorophyll fluorescence excitation spectrum in Arabidopsis chloroplasts

Chlorophyll fluorescence has been often used as an intrinsic optical molecular probe to study photosynthesis. In this study, the origin of bands at 437 and 475.5 nm in the chlorophyll fluorescence excitation spectrum for emission at 685 nm in Arabidopsis chloroplasts was investigated using various optical analysis methods. The results revealed that this fluorescence excitation spectrum was related to the absorption characteristics of pigment molecules in PSII complexes. Moreover, the excitation band centred at 475.5 nm had a blue shift, but the excitation band at 437 nm changed relatively less due to induction of non‐photochemical quenching (NPQ). Furthermore, fluorescence emission spectra showed that this blue shift occurred when excitation energy transfer from both chlorophyll b (Chl b) and carotenoids (Cars) to chlorophyll a (Chl a) was blocked. These results demonstrate that the excitation band at 437 nm was mainly contributed by Chl a, while the excitation band at 475.5 nm was mainly contributed by Chl b and Cars. The chlorophyll fluorescence excitation spectrum, therefore, could serve as a useful tool to describe specific characteristics of light absorption and energy transfer between light‐harvesting pigments. Copyright © 2015 John Wiley & Sons, Ltd.     

Phycobilin/chlorophyll excitation equilibration upon carotenoid-induced non-photochemical fluorescence quenching in phycobilisomes of the cyanobacterium Synechocystis sp. PCC 6803

To determine the mechanism of carotenoid-sensitized non-photochemical quenching in cyanobacteria, the kinetics of blue-light-induced quenching and fluorescence spectra were studied in the wild type and mutants of Synechocystis sp. PCC 6803 grown with or without iron. The blue-light-induced quenching was observed in the wild type as well as in mutants lacking PS II or IsiA confirming that neither IsiA nor PS II is required for carotenoid-triggered fluorescence quenching. Both fluorescence at 660 nm (originating from phycobilisomes) and at 681 nm (which, upon 440 nm excitation originates mostly from chlorophyll) was quenched. However, no blue-light-induced changes in the fluorescence yield were observed in the apcE⁻ mutant that lacks phycobilisome attachment. The results are interpreted to indicate that interaction of the Slr1963-associated carotenoid with - presumably - allophycocyanin in the phycobilisome core is responsible for non-photochemical energy quenching, and that excitations on chlorophyll in the thylakoid equilibrate sufficiently with excitations on allophycocyanin in wild type to contribute to quenching of chlorophyll fluorescence.