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.     

Model for fluorescence quenching in light harvesting complex II in different aggregation states

Low-temperature (77 K) steady-state fluorescence emission spectroscopy and dynamic light scattering were applied to the main chlorophyll a/b protein light harvesting complex of photosystem II (LHC II) in different aggregation states to elucidate the mechanism of fluorescence quenching within LHC II oligomers. Evidences presented that LHC II oligomers are heterogeneous and consist of large and small particles with different fluorescence yield. At intermediate detergent concentrations the mean size of the small particles is similar to that of trimers, while the size of large particles is comparable to that of aggregated trimers without added detergent. It is suggested that in small particles and trimers the emitter is monomeric chlorophyll, whereas in large aggregates there is also another emitter, which is a poorly fluorescing chlorophyll associate. A model, describing populations of antenna chlorophyll molecules in small and large aggregates in their ground and first singlet excited states, is considered. The model enables us to obtain the ratio of the singlet excited-state lifetimes in small and large particles, the relative amount of chlorophyll molecules in large particles, and the amount of quenchers as a function of the degree of aggregation. These dependencies reveal that the quenching of the chl a fluorescence upon aggregation is due to the formation of large aggregates and the increasing of the amount of chlorophyll molecules forming these aggregates. As a consequence, the amount of quenchers, located in large aggregates, is increased, and their singlet excited-state lifetimes steeply decrease.     

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     

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.     

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     

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.     

On the relationship between the non-photochemical quenching of the chlorophyll fluorescence and the Photosystem II light harvesting efficiency. A repetitive flash fluorescence induction study

Plants respond to excess light by a photoprotective reduction of the light harvesting efficiency. The notion that the non-photochemical quenching of chlorophyll fluorescence can be reliably used as an indicator of the photoprotection is put to a test here. The technique of the repetitive flash fluorescence induction is employed to measure in parallel the non-photochemical quenching of the maximum fluorescence and the functional cross-section (sigma(PS II)) which is a product of the photosystem II optical cross-section a(PS II) and of its photochemical yield Phi(PS II) (sigma (PS II) = a(PS II) Phi(PS II)). The quenching is measured for both, the maximum fluorescence found in a single-turnover flash (F(M) (ST)) and in a multiple turnover light pulse (F(M) (MT)). The experiment with the diatom Phaeodactylum tricornutum confirmed that, in line with the prevalent model, the PS II functional cross-section sigma (PS II) is reduced in high light and restored in the dark with kinetics and amplitude that are closely matching the changes of the F(M) (ST) and F(M) (MT) quenching. In contrast, a poor correlation between the light-induced changes in the PS II functional cross-section sigma (PS II) and the quenching of the multiple-turnover F(M) (MT) fluorescence was found in the green alga Scenedesmus quadricauda. The non-photochemical quenching in Scenedesmus quadricauda was further investigated using series of single-turnover flashes given with different frequencies. Several mechanisms that modulate the fluorescence emission in parallel to the Q(A) redox state and to the membrane energization were resolved and classified in relation to the light harvesting capacity of Photosystem II.     

A quantitative determination of photochemical and non-photochemical quenching during the slow phase of the chlorophyll fluorescence induction curve of bean leaves

Estimations of the changes in the reduction-oxidation state of Photosystem II electron acceptors in Phaseolus vulgaris leaves were made during the slow decline in chlorophyll fluorescence emission from the maximal level at P to the steady-state level at T. The relative contributions of photochemical and non-photochemical processes to the fluorescence quenching were determined from these data. At a low photon flux density of 100 μmol · m^?2 · s^?1, non-photochemical quenching was the major contributor to the fluorescence decline from P to T, although large charges were observed in photochemical quenching immediately after P. On increasing the light intensity 10-fold, the contribution of photochemical processes to fluorescence quenching was markedly diminished, with nearly all the P-to-T fluorescence decline being attributable to changes in non-photochemical quenching. The possible factors responsible for changes in non-photochemical quenching within the leaves are discussed.     

Control of the light harvesting function of chloroplast membranes: the LHCII-aggregation model for non-photochemical quenching

Dissipation of excess excitation energy within the photosystem II light-harvesting antenna (LHCII) by non-photochemical quenching (NPQ) is an important photoprotective process in plants. An update to a hypothesis for the mechanism of NPQ [FEBS Letters 292, 1991] is presented. The impact of recent advances in understanding the structure, organisation and photophysics of LHCII is assessed. We show possible locations of the predicted regulatory and quenching pigment-binding sites in the structural model of the major LHCII. We suggest that NPQ is a highly regulated concerted response of the organised thylakoid macrostructure, which can include different mechanisms and sites at different times.     

Thermoluminescence as a probe of Photosystem II in intact leaves: Non-photochemical fluorescence quenching in peas grown in an intermittent light regime

We have measured thermoluminescence (TL) and chlorophyll fluorescence from leaves of peas grown under an intermittent light regime (IML) and followed changes in those leaves during greening. IML peas show low variable fluorescence and a certain capacity for reversible non-photochemical quenching. It has been suggested that reversible quenching may be caused by pH-dependent release of Ca²⁺ from Photosystem II (PS II) (Krieger and Weis (1992) Photosynthetica 27: 89–98). Under conditions in which reversible non-photochemical quenching occurs, a TL band at around 50 °C is observed, in the presence of DCMU, in IML leaves. A band in this temperature range has previously been observed in PS II depleted of Ca²⁺ (Ono and Inoue (1989) Biochimica et Biophysica Acta 973: 443–449). The 50 °C band disappears upon dark adaptation. In mature leaves, no significant band is seen at 50 °C. It is concluded that, in IML leaves, reversible quenching may be related to the release of Ca²⁺ from Photosystem II. However, it seems that in the mature system, under most conditions, such release does not contribute significantly to quenching     

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-induced fluorescence quenching in the light-harvesting chlorophyll a/b protein complex

Summary Irradiation of the principal photosystem II light-harvesting chlorophyll-protein antenna complex, LHC II, with high light intensities brings about a pronounced quenching of the chlorophyll fluorescence. Illumination of isolated thylakoids with high light intensities generates the formation of quenching centres within LHC II in vivo, as demonstrated by fluorescence excitation spectroscopy. In the isolated complex it is demonstrated that the light-induced fluorescence quenching: a) shows a partial, biphasic reversibility in the dark; b) is approximately proportional to the light intensity; c) is almost independent of temperature in the range 0–30°C; d) is substantially insensitive to protein modifying reagents and treatments; e) occurs in the absence of oxygen. A possible physiological importance of the phenomenon is discussed in terms of a mechanism capable of dissipating excess excitation energy within the photosystem II antenna.Abbreviations chla chlorophyll a - chlb chlorophyll b - F₀ fluorescence yield with reaction centers open - F_m fluorescence yield with reaction centres closed - F_i fluorescence at the plateau level of the fast induction phase - LHC II light-harvesting chlorophyll a/b protein complex II - PS II photosystem II - PSI photosystem I - Tricine N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine     

Light-induced quenching of chlorophyll fluorescence at 77 K in leaves,chloroplasts and Photosystem II particles

The light-induced chlorophyll (Chl) fluorescence decline at 77 K was investigated in segments of leaves, isolated thylakoids or Photosystem (PS) II particles. The intensity of chlorophyll fluorescence declines by about 40% upon 16 min of irradiation with 1000 μmol m⁻² s⁻¹ of white light. The decline follows biphasic kinetics, which can be fitted by two exponentials with amplitudes of approximately 20 and 22% and decay times of 0.42 and 4.6 min, respectively. The decline is stable at 77 K, however, it is reversed by warming of samples up to 270 K. This proves that the decline is caused by quenching of fluorescence and not by pigment photodegradation. The quantum yield for the induction of the fluorescence decline is by four to five orders lower than the quantum yield of Q_A reduction. Fluorescence quenching is only slightly affected by addition of ferricyanide or dithionite which are known to prevent or stimulate the light-induced accumulation of reduced pheophytin (Pheo). The normalised spectrum of the fluorescence quenching has two maxima at 685 and 695 nm for PS II emission and a plateau for PS I emission showing that the major quenching occurs within PS II. ‘Light-minus-dark’ difference absorbance spectra in the blue spectral region show an electrochromic shift for all samples. No absorbance change indicating Chl oxidation or Pheo reduction is observed in the blue (410–600 nm) and near infrared (730–900 nm) spectral regions. Absorbance change in the red spectral region shows a broad-band decrease at approximately 680 nm for thylakoids or two narrow bands at 677 and 670–672 nm for PS II particles, likely resulting also from electrochromism. These absorbance changes follow the slow component of the fluorescence decline. No absorbance changes corresponding to the fast component are found between 410 and 900 nm. This proves that the two components of the fluorescence decline reflect the formation of two different quenchers. The slow component of the light-induced fluorescence decline at 77 K is related to charge accumulation on a non-pigment molecule of the PS II complex. This revised version was published online in June 2006 with corrections to the Cover Date.     

pH dependent chlorophyll fluorescence quenching in spinach thylakoids from light treated or dark adapted leaves

The pH dependence of maximum chlorophyll fluorescence yield (F_m) was examined in spinach thylakoids in the presence of nigericin to dissipate the transthylakoid pH gradient. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) was present to eliminate photochemical quenching. Thylakoids were prepared from dark adapted leaves (dark thylakoids) or preilluminated leaves (light thylakoids). In the latter there had been approximately 50% conversion of the xanthophyll violaxanthin to zeaxanthin, while no conversion had occurred in the former. In the presence of a reductant such as ascorbate, antimycin A sensitive quenching was observed (half maximal quenching at 5 M), whose pH dependence differed between the two types of thylakoid. Preillumination of leaves resulted in more quenching at pH values where very little quenching was observed in dark thylakoids (pH 5–7.6). This was similar to activation of high-energy-state quenching (qE) observed previously (Rees D, Young A, Noctor G, Britton G and Horton P (1989) FEBS Lett 256: 85–90). Thylakoids isolated from preilluminated DTT treated leaves, that contained no zeaxanthin, behaved like dark thylakoids. A second form of quenching was observed in the presence of ferricyanide, that could be reversed by the addition of ascorbate. This was not antimycin A sensitive and showed the same pH dependence in both types of thylakoid. The former type of quenching, but not the latter, showed similar low temperature fluorescence emission spectra to qE, and was considered to occur by the same mechanism.Abbreviations DCMU 3(3,4-dichlorophenyl)-1,1-dimethylurea - DTT dithiothreitol - EDTA Ethylenediaminetetra-acetic acid - F₀ dark level fluorescence yield - F_m maximum fluorescence yield - F_v/F_m ratio of variable to total fluorescence yield - Hepes 4-(2-hydroxyethyl)1-piperazineethanesul-phonic acid - Mes 2-(N-morpholino) ethanesulfonate - pH transthylakoid pH gradient - PS I Photosystem I - PS II Photosystem II - Q_A primary stable electron acceptor of Photosystem II - qE high-energy-state fluorescence quenching     

Diurnal changes in photochemical efficiency,the reduction state of Q,radiationless energy dissipation,and non-photochemical fluorescence quenching in cacti exposed to natural sunlight in northern Venezuela

Summary Diurnal measurements of chlorophyll a fluorescence from cacti (Nopalea cochenillifera, Opuntia ficus-indica, and Opuntia wentiana) growing in northern Venezuela were used to determine photochemical fluorescence quenching related to the reduction state of the primary electron acceptor of PS II as well as non-photochemical fluorescence quenching which reflects the fraction of energy going primarily into radiationless deexcitation. The cladodes used in this study were oriented such that one surface received direct sunlight in the morning and the other one during the afternoon. Both surfaces exhibited large increases in radiationless energy dissipation from the photochemical system accompanied by decreases in PS II photochemical efficiency during direct exposure to natural sunlight. During exposure to sunlight in the morning, dissipation of absorbed light energy through photosynthesis and radiationless energy dissipation was sufficient to maintain Q, the primary electron acceptor for PS II, in a low reduction state. During exposure to sunlight in the afternoon, however, the reduction state of Q rose to levels greater than 50%, presumably due to a decrease in photosynthetic electron transport as the decarboxylation of the nocturnally accumulated malic acid was completed. Exposure to direct sunlight in the afternoon also led to more sustained increases in radiationless energy dissipation. Furthermore, the increases in radiationless energy dissipation during exposure of a water-stressed cladode of O. wentiana to direct sunlight were much greater than those from other well-watered cacti, presumably due to sustained stomatal closure and decreased rates of photosynthetic electron transport. These results indicate that the radiationless dissipation of absorbed light is an important process in these CAM plants under natural conditions, and may reflect a protective mechanism against the potentially damaging effects of the accumulation of excessive energy, particularly under conditions where CO₂ availability is restricted.Abbreviations CAM crassulacean acid metabolism - F _o instantaneous fluorescence emission - F _M maximum fluorescence emission - F _v variable fluorescence emission - K _D rate constant for radiationless energy dissipation in the antenna chlorophyll - PFD photon flux density - PS I photosystem I - PS II photosystem II - Q primary electron acceptor of photosystem II - q _NP non-photochemical fluorescence quenching - q _P photochemical fluorescence quenching - T _C cladode temperature     

Two forms of the Photosystem II D1 protein alter energy dissipation and state transitions in the cyanobacterium Synechococcus sp. PCC 7942

Synechococcus sp. PCC 7942 (Anacystis nidulans R2) contains two forms of the Photosystem II reaction centre protein D1, which differ in 25 of 360 amino acids. D1: 1 predominates under low light but is transiently replaced by D1:2 upon shifts to higher light. Mutant cells containing only D1:1 have lower photochemical energy capture efficiency and decreased resistance to photoinhibition, compared to cells containing D1:2. We show that when dark-adapted or under low to moderate light, cells with D1:1 have higher non-photochemical quenching of PS II fluorescence (higher q_N) than do cells with D1:2. This is reflected in the 77 K chlorophyll emission spectra, with lower Photosystem II fluorescence at 697–698 nm in cells containing D1:1 than in cells with D1:2. This difference in quenching of Photosystem II fluorescence occurs upon excitation of both chlorophyll at 435 nm and phycobilisomes at 570 nm. Measurement of time-resolved room temperature fluorescence shows that Photosystem II fluorescence related to charge stabilization is quenched more rapidly in cells containing D1:1 than in those with D1:2. Cells containing D1:1 appear generally shifted towards State II, with PS II down-regulated, while cells with D1:2 tend towards State I. In these cyanobacteria electron transport away from PS II remains non-saturated even under photoinhibitory levels of light. Therefore, the higher activity of D1:2 Photosystem II centres may allow more rapid photochemical dissipation of excess energy into the electron transport chain. D1:1 confers capacity for extreme State II which may be of benefit under low and variable light.Abbreviations D1 the atrazine-binding 32 kDa protein of the PS II reaction centre core - D1:1 the D1 protein constitutively expressed during acclimated growth in Synechococcus sp. PCC 7942 - D1:2 an alternate form of the D1 protein induced under excess excitation in Synechococcus sp. PCC 7942 - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - Fo minimal fluorescence in the dark-adapted state - Fo minimal fluorescence in a light-adapted state - F_M maximum fluorescence with all quenching mechanisms at a minimum, measured in presence of DCMU - F_M maximal fluorescence in a light-adapted state, measured with a saturating flash - F_Mdark maximal fluorescence in the dark-adapted state - F_V variable fluorescence in a light-adapted state (F_M-Fo) - PAM pulse amplitude modulated fluorometer - q_N non-photochemical quenching of PS II fluorescence - q_N (dark) q_N in the dark adapted state - q_P photochemical quenching of fluorescence     

Diurnal fluctuation in the composition of chlorophyll a/b light harvesting antenna of photosystem II in young wheat leaves

We investigated the diurnal fluctuation in the composition of the light harvesting chlorophyll a/b antenna of photosystem II in young wheat (Triticum aestivum) leaves grown under periodic day/night irradiation. By means of gel electrophoresis of the polypeptides of thylakoid membranes, we determined the amount of 25 kDa and 27 kDa polypeptides, which are the main components of the peripheral and inner antenna subpopulations, respectively. Our data show a preferential fluctuation in the amount of the 25 kDa protein relative to the 27 kDa polypeptide, in parallel to the fluctuation in the amount of chlorophyll a/b antenna of photosystem II, which suggests that the peripheral antenna plays a role in the diurnal adjustment of the antenna size.     

Oceanic picophytoplankton having a high abundance of chlorophyll b in the major light harvesting chlorophyll protein complex

Chl a-containing, very small unicellular, eukaryotic phytoplankton (picophytoplankton) often become the dominant organisms near the bottom of the euphotic zone in the ocean, where light is limited, not only in intensity (about 0.5% of the surface irradiance), but also in quality (dominant in blue to green wavelengths). We have isolated picophytoplankton from subsurface waters (from 75 to 150 m in depth) of the Kuroshio area near Japan. EM observations showed that a single chloroplast occupies a large part of the cytoplasm. Some of the isolates have a flagellum. The major photosynthetic pigments found in these isolates were chlorophyll a and b. The light-harvesting chlorophyll a/b complex (LHCP) was isolated from three clones of picophytoplankton, one flagellated form (NIBB8001) and two coccoid forms (94B8100A and 94B5100C) . More than 50% of the total chlorophylls were recovered in the major LHCP fraction. A common feature of the major LHCPs isolated from the three picophytoplankton clones was a high abundance of chlorophyll b: the ratios of chlorophyll a to b were about 0.8, 0.7 and 0.6 for the clones NIBB8001, 94B8100A and 94B5100C, respectively. These values were very low compared with those in chlorophyll a/b-binding LHCIIs in higher plants and in the major chlorophyll a/b-binding LHCPs in microalgae (higher than 1.0). The major LHCP apoproteins of NIBB8001 and 94B5100C contained one major polypeptide; the apparent molecular masses analyzed with SDS-PAGE were about 22 kDa and 27 kDa, respectively. The major LHCP apoprotein of 94B8100A had two major polypeptides having apparent molecular masses of about 23 and 25 kDa. None of the thylakoid proteins cross-reacted with an antibody raised against the LHC II apoprotein of spinach. It is suggested that the high abundance of chlorophyll b in picophytoplankton, together with a large chloroplast in a small cell, enable them to utilize the reduced light in their habitat.     

NPQ(T): a chlorophyll fluorescence parameter for rapid estimation and imaging of non‐photochemical quenching of excitons in photosystem‐II‐associated antenna complexes

In photosynthesis, light energy is absorbed by light‐harvesting complexes and used to drive photochemistry. However, a fraction of absorbed light is lost to non‐photochemical quenching (NPQ) that reflects several important photosynthetic processes to dissipate excess energy. Currently, estimates of NPQ and its individual components (q_E, q_I, q_Z and q_T) are measured from pulse‐amplitude‐modulation (PAM) measurements of chlorophyll fluorescence yield and require measurements of the maximal yield of fluorescence in fully dark‐adapted material (F_m), when NPQ is assumed to be negligible. Unfortunately, this approach requires extensive dark acclimation, often precluding widespread or high‐throughput use, particularly under field conditions or in imaging applications, while introducing artefacts when F_m is measured in the presence of residual photodamaged centres. To address these limitations, we derived and characterized a new set of parameters, NPQ_(T), and its components that can be (1) measured in a few seconds, allowing for high‐throughput and field applications; (2) does not require full relaxation of quenching processes and thus can be applied to photoinhibited materials; (3) can distinguish between NPQ and chloroplast movements; and (4) can be used to image NPQ in plants with large leaf movements. We discuss the applications benefits and caveats of both approaches.     

Rapid light-response curves of chlorophyll fluorescence in microalgae: relationship to steady-state light curves and non-photochemical quenching in benthic diatom-dominated assemblages

Rapid light-response curves (RLC) of variable chlorophyll fluorescence were measured on estuarine benthic microalgae with the purpose of characterising its response to changes in ambient light, and of investigating the relationship to steady-state light-response curves (LC). The response of RLCs to changes in ambient light (E, defined as the irradiance level to which a sample is acclimated to prior to the start of the RLC) was characterised by constructing light-response curves for the RLC parameters α _RLC, the initial slope, ETR_m,RLC, the maximum relative electron transport rate, and E _k,RLC, the light-saturation parameter. Measurements were carried out on diatom-dominated suspensions of benthic microalgae and RLC and LC parameters were compared for a wide range of ambient light conditions, time of day, season and sample taxonomic composition. The photoresponse of RLC parameters was typically bi-phasic, consisting of an initial increase of all parameters under low ambient light (E < 21–181 μmol m⁻² s⁻¹), and of a phase during which α _RLC decreased significantly with E, and the increase of ETR_m,RLC and E _k,RLC was attenuated. The relationship between RLC and LC parameters was dependent on ambient irradiance, with significant correlations being found between α _RLC and α, and between ETR_m,RLC and ETR_m, for samples acclimated to low and to high ambient irradiances, respectively. The decline of α _RLC under high light (Δα _RLC) was strongly correlated (P < 0.001 in all cases) with the level of non-photochemical quenching (NPQ) measured before each RLC. These results indicate the possibility of using RLCs to characterise the steady-state photoacclimation status of a sample, by estimating the LC parameter E _k, and to trace short-term changes in NPQ levels without dark incubation.